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Part 1 : Laboratory Principles 20 Gastrointestinal Disease,334

1 Laboratory Essentials, 1 21 Diseases of the Liver,346

2 Practical Laboratory Safety,25 22 Pancreatic Diseases and Disorders,363

3 Principles of Laboratory Instrumentation, 41 23 Endocrinology,379

4 lmmunoassays,78 24 Kidney and Urinary Tract Diseases,412

5 Molecular Diagnostics, 97 25 Reproductive Diseases and Disorders,432

6 Automation in the Laboratory,114 26 Pregnancy, 446

7 Laboratory Quality Management Systems,124 27 Bone,Joint,and Skeletal Muscle Diseases,458

8 Enzymes,156 28 Nervous System Diseases,474

29 Skin, Hair,and Nail Diseases,488

Part 2: Pathophysiology and Analytes
30 Eye and Ear Diseases,497
9 Clinical Chemistry and Disease,171
31 Nutritional and Metabolic Diseases,504
10 Cell Injury and Its Relationship to Disease,179
32 Immune System Diseases,529
11 Inflammation,187

12 Body Fluids and Electrolytes,204

Part 3: Other Aspects of Clinical Chemistry

13 Blood Gases and Acid-Base Balance,219 33 Therapeutic Drug Monitoring,552

14 Blood Diseases,234 34 Toxicology,584

15 Proteins, 251 35 Transplantation,616

16 Cancer and Tumor Markers,268 36 Emergency Preparedness,634

17 Blood Vessel Diseases,289 Glossary,661

18 Heart Disease,306 Answer Key,683

19 Respiratory Diseases,318 Index,707

 Clinical Chemistry
Fundamentals and Laboratory
Techniques
Clinical Chemistry
Fundamentals and Laboratory Techniques

Author

Donna Larson, EdD, MT (ASCP), DLM

Vice President for Academic and Student Affairs
Clatsop Community College
Astoria, Oregon

Consulting Editors

Joshua Hayden, PhD, DABCC

Assistant Professor of Pathology and Laboratory Medicine
Weill Cornell Medical College
Director, Toxicology and Therapeutic Drug Monitoring
Assistant Director, Central Laboratory
New York Presbyterian Hospital-Cornell Campus
New York, New York

Hari Nair, PhD, DABCC

Technical Director
Boston Heart Diagnostics
Framingham, Massachusetts

ELSEVIER
ELSEVIER
325 1 Riverporr Lane
St. Louis, Missouri 63043

CLINICAL CHEMISTRY: FUNDAMENTALS AND

LABORATORY TECHNIQUES ISBN: 978- 1 -4 5 57-42 1 4 - 1

Copyright© 2017 by Elsevier, Inc. All rights reserved.

No part of this publication may b e reproduced or rransmirred i n any form or b y any means, electronic or mechanical,
including photocopying, recording, or any information storage and retrieval system, without permission in writing
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Agency, can be found at our website: www.elsevier.com/permissions.

This book and the individual contributions contained in it are protected under copyright by the Publisher (other than
as may be noted herein) .

Notices

Knowledge and best practice in this field are constantly changing. As new research and experience broaden our
understanding, changes in research methods, professional practices, or medical treatment may become necessary.
Practitioners and researchers must always rely on their own experience and knowledge in evaluating and using
any information, methods, compounds, or experiments described herein. In using such information or methods
they should be mindful of their own safety and the safety of others, including parties for whom they have a profes­
sional responsibility.
With respect to any drug or pharmaceutical products identified, readers are advised to check the most current
information provided (i) on procedures featured or (ii) by the manufacturer of each product to be administered,
to verify the recommended dose or formula, the method and duration of administration, and contraindications.
It is the responsibility of practitioners, relying on their own experience and knowledge of their patients, to make
diagnoses, to determine dosages and the best treatment for each individual patient, and to take all appropriate
safety precautions.
To the fullest extent of the law, neither the Publisher nor the authors, contributors, or editors, assume any liabil­
ity for any injury and/or damage to persons or property as a matter of products liability, negligence or otherwise,
or from any use or operation of any methods, products, instructions, or ideas contained in the material herein.

Library of Congress Cataloging-in-Publication Data

Clinical chemistry : fundamentals and laboratory techniques I edited by Donna Larson ; consulting editors,
Larson, Donna, editor. I Hayden, Joshua Uoshua A.) , editor. I Nair, Hari, editor.

Joshua Hayden, Hari Nair.

Clinical chemistry (Larson)

Includes bibliographical references and index.

St. Louis, Missouri : Elsevier/Saunders, [20 1 7] 1

LCCN 20 1 5 0440741 ISBN 978 1 4 5 5742 1 4 1 (paperback : alk. paper)

I MESH: Clinical Chemistry Tests.
LCC RB40 I NLM QY 90 I DOC 6 1 6. 07/5 6--dc23 LC record
available at http :/ /lccn.loc.gov/20 1 5044074

Executive Content Strategist: Kellie White

Content Development Manager: Jean Sims Fornango
Content Development Specialist: Beth LoGiudice, Spring Hollow Press
Publishing Services Manager: Catherine Jackson
Senior Project Manager: Daniel Fitzgerald
Designer: Margaret Reid

Working together

Printed in Canada
IJ!Jr- W\\"W
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to grow libraries in
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<:om • www. hooka1J org
Last digit is the print number: 9 8 7 6 5 4 3 2
 To my mom and dad, Donald and Barbara Bedard (I wish they could have been here
to see this); to my husband, Earl, and my son, Adrian, for their love and support; to my
sister and her family for their warmth and love; to the Allards for their support during
my clinical year and college years; and to all the friends and colleagues I worked with at
Wentworth-Douglass Hospital (NH), 509th Strategic Hospital (NH), RAF Lakenheath
Regional Hospital (UK), Winston-Salem State University (NC), Mt Hood Community
College (OR), Portland Community College (OR), and Clatsop Community College (OR).
Donna Larson
I appreciate the opportunity Elsevier provided for me to
write the first edition of this clinical chemistry book for
medical laboratory technology students. The process was
exciting, exhausting, challenging, and an educational expe­
rience like no other. I would like to thank the contributors
for their hard work to help make this book possible.
I would like to thank the Elsevier staff for the assistance,
guidance, encouragement, and experience that they shared
with me throughout the development of the book. Thank
you to Ellen Wurm-Cutter, who helped me through the
proposal and beginning stages of manuscript development.
vi
A big thank you to Kellie White, Jean Sims Fornango, and
Beth LoGiudice for joining the team and seeing this proj­
ect through to completion. The final product has been a
long time coming. My Thursday mornings will never be
the same! Thanks also to Dan Fitzgerald and his team for
putting everything together in a beautiful full-color book.
Everyone was understanding, patient, compassionate,
empathetic, and truly amazing.
Donna Larson
Sheryl Berman, PhD
Division Dean of Health Professions
Lane Community College
Eugene, Oregon
Jimmy L. Boyd, CLS (NCA), MS/MHS
Assistant Professor, Department Head
Medical Laboratory Technology
Arkansas State University, Beebe
Beebe, Arkansas
Craig Foreback, PhD
Senior Consultant
Clear Medical Solutions, LLC
Bradenton, Florida
Senior Lecturer Emeritus
University of Wisconsin School of Medicine
and Public Health
Madison, Wisconsin
Danielle Fortuna, MD
Department of Pathology, Anatomy, and
Cell Biology
Sidney Kimmel Medical College
Thomas Jefferson University
Philadelphia, Pennsylvania
Thomas Kampfrath, PhD, DABCC
Clinical Biochemist
Santa Clara Valley Medical Center
Department of Pathology and Laboratory Medicine
San Jose, California
Laura J. McCloskey, PhD
Department of Pathology, Anatomy, and Cell Biology
Sidney Kimmel Medical College
Thomas Jefferson University
Philadelphia, Pennsylvania
M. Laura Parnas, PhD, DABCC, FACB
Director of Clinical Science
Sutter Health Shared Laboratory
Livermore, California
John W. Ridley, PhD, RN, MT (ASCP)
Formerly, Director of Medical Laboratory Technology
West Central Technical College
Waco, Georgia
Laird C. Sheldahl, PhD
Instructor, Anatomy and Physiology, Biology
Mount Hood Community College
Gresham, Oregon
Douglas F. Stickle, PhD, DABCC, FACB
Department of Pathology, Anatomy, and Cell Biology
Sidney Kimmel Medical College
Thomas Jefferson University
Philadelphia, Pennsylvania
Zi-Xuan Wang, PhD
Department of Pathology, Anatomy, and Cell Biology
Sidney Kimmel Medical College
Thomas Jefferson University
Philadelphia, Pennsylvania
vii
This page intentionally left blank
Keith Bellinger, PBT (ASCP) Amy Gatautis, MBA, MT (ASCP), SC
Medical Technologist Program Director, Medical Laboratory Technology
The United States Department of Veterans Affairs New Cuyahoga Community College
Jersey Health Care System Cleveland, Ohio
East Orange, New Jersey
Assistant Professor, Phlebotomy Kristine Hayes, MAT, MLS (ASCP)
Rutgers-The State University of New Jersey MLT and Phlebotomy Program Coordinator
Newark, New Jersey Moberly Area Community College
Moberly, Missouri
Stephanie Bielas, PhD
Assistant Professor of Human Genetics Candy Hill, MEd, MT (ASCP)
University of Michigan CLT Program Coordinator
Ann Arbor, Michigan Jefferson State Community College
Birmingham, Alabama
Jimmy L. Boyd, CLS (NCA), MS/MHS
Assistant Professor, Department Head Lorri Huffard, PhD, MT (ASCP), SBB
Medical Laboratory Technology Dean, Science & Health Programs
Arkansas State University, Beebe Wytheville Community College
Beebe, Arkansas Wytheville, Virginia

Russell Cheadle, MS, MLS (ASCP) Phyllis Ingham, EdD, MEd, MT (ASCP)
Professor, Clinical Laboratory Technology Director Clinical Laboratory Technology Program
Macomb Community College West Georgia Technical College
Warren, Michigan Waco, Georgia

Cathy Crawford, BS, MT (ASCP) Stephen M. Johnson, MS, MT (ASCP)

Clinical Courses Instructor and MLT Teaching Assistant Program Director
Mount Aloysius College Saint Vincent Health Center School of Medical
Cresson, Pennsylvania Technology
Erie, Pennsylvania
Karen M. Escolas, EdD, MT (ASCP)
Chair, Department of Medical Laboratory Technology Haywood Joiner Jr., EdD, MT (ASCP)
Farmingdale State College, State University of New York Chair, Department of Allied Health
Farmingdale, New York Louisiana State University at Alexandria
Alexandria, Louisiana
Roger Fortin, MS, MBA, MLS (ASCP)
Program Director Stephanie Jordan, BS, MLS (ASCP), CM
Bunker Hill Community College Assistant Professor
Charlestown, Massachusetts Pierpont Community and Technical College
Fairmont, West Virginia
Trent Freeman, MA, BS, MLS (ASCP)
Education Coordinator Jeffrey Josifek, MS, MLS (ASCP), CLS (NCA)
Medical Education and Training Campus Department of Medical Laboratory Technology
The George Washington University Portland Community College
Fort Sam Houston, Texas Portland, Oregon
ix
 Reviewers

Minh Kosfeld, PhD, MLT (ASCP) Ryan Rowe, MLS (ASCP)

Assistant Professor Weber State University
Department of Biomedical Laboratory Science Ogden, Utah
Doisy College of Health Sciences
St. Louis University Mary Sadlowski, MT (ASCP)
St. Louis, Missouri Medical Technologist
Greater Baltimore Medical Center and Community
Marc L. Meyers, MBA, MT (ASCP) College of Baltimore County
PM Laboratory Coordinator Towson, Maryland
Centegra Clinical Laboratories
McHenry, Illinois Cheryl Selvage, MS, MT (ASCP)
Associate Professor
Constance Moore, MS, MT (ASCP) Lorain County Community College
Program Director, Laboratory Sciences Elyria, Ohio
Eastern Gateway Community College
Steubenville, Ohio Anita Marie Smith, MT (AMT), MBA
Laboratory Administrative Director
Richard C. Mroz Jr., DA, MS, BSMT, MT (ASCP) Moberly Regional Medical Center
MLT Program Director Moberly, Missouri
Fortis Institute
Fort Lauderdale, Florida Angela Sparkman, MEd, MT (ASCP)
Program Director, Assistant Professor of the Medical
Dawn Nelson, MA, MT (ASCP) Laboratory Technology Program
MLT Program Director Ivy Tech Community College
Florence-Darlington Technical College Sellersburg, Indiana
Florence, South Carolina
Andrea Thompson, BS, MLT (ASCP)
Kathleen C. Perlmutter, MBA, MT (ASCP) MLT Instructor
Phlebotomy Coordinator, MLT Faculty Barton Community College
Montgomery County Community College Great Bend, Kansas
Blue Bell, Pennsylvania
Dionne M. Thompson, MSE, MT (ASCP)
Jennifer D. Perry, MS, BSMT (ASCP) MLT Program Director/Instructor
Associate Professor and Chairperson Three Rivers College
Clinical Laboratory Sciences Department Poplar Bluff, Missouri
Marshall University
Huntington, West Virginia

Ellen F. Romani, AAS (MLT), MS

Department Chair
Medical Laboratory Technology/Phlebotomy/Therapeutic
Massage
Spartanburg Community College
Spartanburg, South Carolina
Clinical Chemistry: Fundamentals and Laboratory Techniques
is a comprehensive, readable, and student-friendly text for
2-year medical laboratory technology programs. The text­
book has a full-color design along with detailed illustrations
and diagrams to help students with complex chemistry
concepts. Pathophysiologic concepts are included to help
students understand the clinical relevance of clinical chem­
istry assays.
Purpose and Organization
As I look back at my journey in clinical laboratory science,
I cannot help but marvel at how laboratory test methods
rapidly changed over the course of the 20th century and
into the 2 1 st century. While researching my dissertation,
The Structure of Knowledge in Clinical Laboratory Science, I
was amazed to read articles in laboratory journals ( 1 940s)
concerning how to build a better cage for laboratory ani­
mals. (Pregnancy tests during that time used rabbits to
determine whether a woman was pregnant.) The radioim­
munoassays that were popular in the 1 970s and into the
1 980s were largely replaced by colorimetric immunoassays
in the late 1 980s and 1 990s. Looking back, there was always
new information in the expanding discipline of clinical lab­
oratory science. The more the knowledge base expands, the
more the students are asked to learn. This is especially true
of medical laboratory technology (MLT) students.
MLT students have a mere 2 years to learn all the clinical
laboratory science (CLS) knowledge (with few prerequisite
and general education courses) on which to build a solid
knowledge foundation. Pieces from various disciplines are
incorporated or embedded in their CLS . When writing this
book, I envisioned a clinical chemistry book that would
incorporate j ust-in-time learning concepts for which the
material would be fortified with additional material when
needed. Building on this approach, Part 1 , Laboratory Prin­
ciples, covers laboratory principles, safety, quality assur­
ance, and other fundamentals of laboratory techniques.
The concepts are essential for anyone working in a clini­
cal laboratory, and this section provides a good reference
for beginning MLT students. For example, the students
do not take a statistics course, but statistical concepts and
calculations are included in Chapter 7, Laboratory Qual­
ity Management Systems. Quality management methods,
including the applications of Westgard rules for control
charts and the calculation of the mean, mode, and standard
deviation, are explained and practiced in that chapter.
Part 2, Pathophysiology and Analytes, covers the dis­
eases, broken down by body system, that are commonly
diagnosed through chemical tests. Each chapter in this sec­
tion contains information about anatomy and physiology
of a specific body system, disease mechanisms of common
conditions that require clinical chemistry testing, and how
laboratory results correlate with clinical disorders. This is a
key section of the book because MLT students usually do
not have room in their program for a separate pathophysiol­
ogy class, unlike other health science students, for whom it
is part of the program paradigm.
Pathophysiologic mechanisms of diseases and the resul­
tant effects on clinical chemistry tests are discussed in each
of the chapters. For example, it is easier to remember test
results that are elevated after an acute myocardial infarction
(MI) if it is known that the muscle is damaged and that
the dying cells release specific chemicals into the blood. If
blood is drawn at timed intervals after the MI, the person
who understands the pathophysiologic mechanism behind
the infarct will know what types of clinical chemistry results
to expect from each specimen.
When diseases are discussed that do not use laboratory
tests for diagnoses or when laboratory tests are used to
rule out other disorders, this information is given so that
students can understand the laboratory test ordering pat­
terns of health care providers. This information also helps
students better understand reflex testing and how the algo­
rithms are developed.
Part 3, Other Aspects of Clinical Chemistry, covers
therapeutic drug monitoring, toxicology, transplantation,
and emergency preparedness. The clinical laboratory has a
critical role in these areas, providing ongoing testing and
assistance.
To complement the organization, the book is writ­
ten in the active voice to help students better understand
the material. Although this may be unconventional for a
textbook at this level, I believe it helps students to better
understand complex clinical chemistry concepts and mas­
ter the material.
Most individuals are visual learners. To that end, many
figures, photographs, tables, and flowcharts are included
xi
 Preface
to help students better understand concepts. Many figures
summarize complex and complicated processes or pathways
to provide better comprehension of the material by students.
Key Features
Cha pter Outl i n e
Each chapter starts with a chapter outline that shows the
main topics that are covered. It provides students and
instructors with a roadmap to the chapter and can be easily
referenced at any time.
Objectives
The textbook format facilitates the learning process by pro­
viding students and educators with detailed objectives that
address the knowledge required to master each chapter's
content. The learning objectives are listed at the beginning
of each chapter, giving students and instructors definitive
evaluation tools to use as the chapter's content is covered.
Objectives are provided at various cognitive mastery lev­
els: comprehension, application, analysis, synthesis, and
evaluation.
Key Terms
Key terms are identified at the beginning of each chapter
and highlighted in the chapter, putting valuable terminol­
ogy at students' fingertips. The key terms are also included
in the Glossary at the back of the book.
Case in Point
A key clinical case study is provided at the beginning of every
appropriate chapter. The Case in Point feature provides appli­
cation of the student's knowledge for correlating the clinical
side of test results. Students are asked to think about impor­
tant questions related to each scenario and to use fundamen­
tal information from the chapter to determine the answers.
Poi nts to Remember
A bulleted list of important concepts is included in the first part
of the chapter, providing an overview of the chapter content.
This list gives students a simple study tool for easy reference.
Summary
A short summary at the end of the chapter highlights
key information from the chapter. Students can revisit
the various chapter topics in short form for review and
reinforcement.
Review Questions
Multiple-choice review questions at the end of every chap­
ter provide students with a unique tool as they prepare for
classroom examinations and certification examinations. The
review questions give students a chance to quiz themselves
on the chapter content, assess their knowledge of important
chapter topics, and evaluate which topics need follow-up
review.
Critica l Th i n king Questions
The Critical Thinking Questions allow students and instruc­
tors to discuss the chapter topics in a broader way. Although
these questions have correct answers, they require more in­
depth thinking, analysis, evaluation, and reflection than
other questions in the chapter.
Case Stud ies
Additional Case Studies round out each most chapters,
giving students another opportunity to apply the knowl­
edge gained from the chapter. The scenarios are meant to
stimulate interest and critical thinking and to encourage
discussion of chapter topics with other students.
Evolve Companion Website
Clinical Chemistry comes with a companion website, found
on Evolve (evolve.elsevier.com/Larson) . This website con­
tains helpful ancillaries for instructors and additional mate­
rials for students.
For the I n structor
The Evolve website has multiple features for the instructor:
• A test bank with multiple-choice questions and ratio­
nales.
• PowerPoint presentations for every chapter that can be
used as is or as a template to prepare lectures.
• A detailed Answer Key with rationales for all in-text
questions.
• The Image Collection that provides electronic files of all
the chapter figures that can be downloaded into Power­
Point presentations.
For the Student
Additional content is available for the student:
• High-definition animations to illustrate key physiologic
and pathophysiologic processes.
• Extra Case Studies for certain chapters for more practical
application of textbook content.
Part 1 : Laboratory Principles 5 Molecular Diagnostics,97

Introduction, 98
Donna Larson
1 Laboratory Essentials, 1
Nucleic Acid Structure and Function, 98
Introduction, 3
Donna Larson
Laboratory Methods, 106
History of Clinical Laboratories, 3 Diagnostic Applications, 110
Types of Clinical Laboratories, 4

Laboratories, 8
Regulation and Accreditation of Clinical 6 Automation in the Laboratory,114

Laboratory Materials, 9 Introduction, 115

Donna Larson

Chemistry Review, 10 Goal of Automation, 115

Laboratory Mathematics, 14 History of Automated Analyzers, 115
Automating Clinical ChemistryTests, 117
2 Practical Laboratory Safety,25 Total Laboratory Automation, 119

Introduction, 26
Donna Larson

Safety Regulations, 26
7 Laboratory Quality Management Systems,124
John W. Ridley and Donna Larson
The Laboratory Safety Program, 29 Introduction, 126
Introduction to Quality, 126
3 Principles of Laboratory Instrumentation, 41 Facilities and Safety Overview, 128
Purchasing and Inventory, 130
Introduction, 43
Craig Foreback and Donna Larson
Process Control, 131
Properties of Light, 43 Assessment, 148
Spectrophotometry, 44 Personnel, 149
Fluorometry, 48 Customer Service, 149
Luminometry, 51 Occurrence Management, 149
Nephelometry andTurbidimetry, 51 Process Improvement, 149
Electrochemistry and Chemical Sensors, 52 Documents and Records, 151
Chromatography, 56 Organization, 152
Mass Spectrometry, 62
Electrophoresis, 67
Colligative Properties, 71
8 Enzymes,156

Point-of-CareTesting, 72 Introduction, 158

M. Laura Parnas and Thomas Kampfrath

Flow Cytometry, 74 The Nature of Enzymes, 158

Kinetics, 159
4 lmmunoassays,78 Enzyme Reaction Conditions, 162
lsoenzymes, 163
Introduction, 80
Donna Larson
Specific Enzymes, 163
Antibodies, Antigens, and Analytes, 80
lmmunochemical Methods, 82
Label Methods, 84
Part 2: Pathophysiology and Analytes

Particle Methods, 88
Light-Scattering Methods, 92
9 Clinical Chemistry and Disease,171

Introduction, 172
Donna Larson

Performance, 93
Factors Affecting Immunoassay Analytical
Definition of Disease, 172

xiii
 Contents

Pathology, 172
Disease Mechanisms, 175
17 Blood Vessel Diseases,289

Biochemistry of Disease, 175 Introduction, 291

Donna Larson

Lipids, 291
10 Cell Injury and Its Relationship to Lipoproteins, 293
Disease,179 Normal Lipoprotein Metabolism, 294
Abnormal Lipoprotein Metabolism, 296
Introduction, 180 Laboratory Procedures and Limitations, 299
Donna Larson

Overview of Cellular Injury, 180 Lipoproteins and Clinical Vascular Disease, 301
Causes of Cellular Injury, 181
Changes in Body Chemistry, 183
LaboratoryTests, 184
18 Heart Disease,306

Introduction, 307
Sheryl Berman

11 Inflammation,187 Heart Structure and Blood Flow, 307

Myocardial Infarction, 307
Introduction, 188 Congestive Heart Failure, 311
Donna Larson

Defense Mechanisms, 188 Congenital Heart Defects, 312

The Inflammation Process, 188 Endocarditis, Myocarditis, and Pericarditis, 314
Acute Inflammation, 195
Chronic Inflammation, 196
Laboratory Procedures and Limitations, 198
19 Respiratory Diseases,318

Introduction, 319
Donna Larson

12 Body Fluids and Electrolytes,204 Structure and Function of the Respiratory System, 320
Respiratory Diseases and Pathophysiology, 320
Introduction, 205
Donna Larson

Fluid Balance and Body Fluid Compartments, 205

Electrolytes, 206
20 Gastrointestinal Disease,334

Colligative Properties, 213 Introduction, 335

Sheryl Berman

Fluid Imbalances, 215 Gastrointestinal System, 335

Gastrointestinal FunctionTests, 342
13 Blood Gases and Acid-Base Balance,219

Introduction, 220
Donna Larson 21 Diseases of the Liver,346

Blood Gases, 220 Introduction, 347

Donna Larson

Acid-Base Theory, 224 Liver and BiliaryTract, 348

Acid-Base Disorders, 226 Liver Diseases, 350
Interpreting Blood Gas Analyses, 230
22 Pancreatic Diseases and Disorders,363
14 Blood Diseases,234
Introduction, 365
Donna Larson

Introduction, 236 Overview of the Pancreas, 365

Sheryl Berman

Hematopoiesis, 236 Pancreatitis, 366

White Blood Cells, 236 Diabetes, 368
Red Blood Cells, 240 Cystic Fibrosis, 375
Malabsorption and Maldigestion, 375
15 Proteins,251

Introduction, 252
Donna Larson 23 Endocrinology,379

Biochemistry of Proteins, 252 Introduction, 382

Jimmy L. Boyd and Donna Larson

Plasma Proteins, 255 Overview of the Endocrine System, 382

Proteins in Other Body Fluids, 264
Testing, 385
Anatomy, Pathophysiology, and Laboratory

16 Cancer and Tumor Markers,268

Introduction, 269
Donna Larson 24 Kidney and Urinary Tract Diseases,412

Cancer andTumor Markers, 269 Introduction, 413

Sheryl Berman

Clinical Correlations, 280 Kidney Anatomy, 414

 Contents

Normal Physiology, 416 32 Immune System Diseases,529

Disease States, 419 Sheryl Berman
Laboratory Procedures, 425 Introduction, 531
Normal Immune System, 531
25 Reproductive Diseases and Disorders,432 Disease States, 538
Donna Larson
Introduction, 433 Part 3: Other Aspects of Clinical Chemistry
Reproductive System Structure and Function, 433
Diseases and Disorders, 435 33 Therapeutic Drug Monitoring,552
Laboratory Procedures and Limitations, 442 Laird C. Sheldahl and Donna Larson
Introduction, 554
26 Pregnancy, 446 Drug Disposition, 554
Sheryl Berman Administration of Drugs, 558
Introduction, 448 Drug Levels, 560
Pregnancy, 448 Cardiovascular Drugs, 562
Complications of Pregnancy, 450 Antibiotics, 567
Fetal Complications, 453 Antiepileptic Drugs, 571
Laboratory Diagnosis of Fetal Abnormalities, 455 Psychoactive Drugs, 576
Bronchodilators, 579
27 Bone,Joint,and Skeletal Muscle Diseases,458 Immunosuppressant Drugs, 580
Donna Larson
Introduction, 459 34 Toxicology,584
Normal Anatomy and Physiology of Bones, 459 Laird C. Sheldahl and Donna Larson
Bone Diseases, 460 Introduction, 586
Normal Anatomy and Physiology of Joints, 466 Routes of Exposure, 587
Joint Diseases, 466 Dose-Response Relationship, 588
Normal Anatomy and Physiology of Muscles, 468 Acute and ChronicToxicity, 588
Muscle Diseases, 469 Specimen Collecting and Handling, 589
Conditions Caused by Pollutants, 590
28 Nervous System Diseases,474 Toxic Agents, 592
Donna Larson Household Products, 597
Introduction, 475 Toxic Metals, 599
Nervous System Anatomy and Physiology, 476 Drugs of Abuse, 602
Nervous System Diseases, 477
Cerebrospinal Fluid Analysis, 483 35 Transplantation,616

Douglas F. Stickle
Daniel/e Fortuna, Laura J. McCloskey, Zi-Xuan Wang,
29 Skin, Hair,and Nail Diseases,488
Donna Larson Introduction, 617
Introduction, 489 Overview ofTransplantation, 617
Skin Diseases, 489 Role of Medical Laboratories inTransplantation, 619
Effects of Systemic Disease on Skin, 491 Overview of the Immune System, 619
Hair Diseases, 492 Role of the Immune System inTransplantation, 622
Nail Diseases, 493 Immunosuppression, 624
Exceptional Cases inTransplantation, 628
30 Eye and Ear Diseases,497 Future ofTransplantation, 629
Donna Larson
Introduction, 497 36 Emergency Preparedness,634
Eyes, 498 Donna Larson
Ears, 501 Introduction, 636
Emergency Preparedness, 636
31 Nutritional and Metabolic Diseases,504 Emergency Response, 637
Donna Larson Laboratory Response Network, 654
Introduction, 507
General Concepts of Health and Disease, 507 Glossary,661
Cold Injuries, 507
Heat Illnesses, 508 Answer Key,683
Nutritional Conditions, 509
Metabolic Diseases, 518 Index,707
Inborn Errors of Metabolism, 520
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 Laboratory Principles

CHAPTER OUTLI N E
I ntrod uction Chemistry Review
H i story of C l i n ical Laboratories Atomic Theory

Types of C l i n ical Laboratories

Chemical Bonds

I n patient Laboratories Factors Affecting Chemical Reactions

Outpatient C l i nics and Physicia ns' Office La boratories Acid, Bases, and Salts
Reference Laboratories Orga nic Chemistry

State and Federal La boratories Biochemistry

M i l itary Laboratories La boratory Mathematics

Reg u l ation and Accreditation of C l i n ical Laboratories

Mola rity

Reg u lation Molal ity

Accreditation Norma l ity

Di l utions
La boratory Materials
Conversions
Glasswa re and Plasticwa re
pH
Centrifuges
Beer's Law
Bala nces
Sta ndard Cu rves
Pi pettes
S u m mary
Reagents
Water

O BJ ECTIVES
At the completion of this chapter, the reader will be able to:
1. Describe the history of the clinical laboratory. 1 0. Describe the types of ba lances and their use i n the
2. List the typica l depa rtments of a clinica l laboratory. laboratory.
3. List the personnel employed in a clinica l la boratory. 1 1 . Compa re a n d contrast serologic a n d vo l u metric
4. List the characteristics of reference, federal, and m i l itary pi pettes.
la boratories. 1 2. Describe the various methods used to ca librate pi pettes.
5. Briefly describe The Joint Com mission and the Col lege of 1 3. Defi ne mola rity and mole and perform the calcu lations
American Pathologists and their roles i n clinical laboratory needed for prepa ring and worki ng with molar solutions.
oversig ht. 1 4. Defi ne molality and perform the ca lcu lations needed for
6. Describe the types of water and the uses for each. prepa ring and worki ng with molal solutions.
7. Com pa re and contrast the types of glasswa re and 1 5. Defi ne normal ity, equivalent weig ht, and m i l l iequ iva lent
plasticwa re. weight and perform the calcu lations needed for prepa ring
8. Describe the types of centrifuges used in the and worki ng with normal solutions.
laboratory. 1 6. Defi ne g/d L and mg/d L u n its and perform ca lculations
9. Describe the operating instructions and precautions for necessa ry to prepa re solutions of a desired g/d L and mg/d L
centrifuges. concentration.
2 PA R T 1 Laboratory Principles

1 7. Solve d i l ution problems for fi n a l vol ume and 1 9. Convert metric u n its from one u n it to another, the th ree

between 51 u nits and conventiona l u nits, a bsorbance to

concentration g iven the i n itial vol u mes and tem perature sca les (i.e., Fa h renheit, Celsius, and Kelvin),
concentrations.
1 8. Describe how serial d i l utions a re prepared. transm itta nce and transmitta nce to a bsorba nce, and a bsor­
bance va l ues to concentration of the u n known.

KEY TERMS
Accred iting Bureau of Health Education C l i n ical Laboratory Mole
Schools I m p rovement Act Nalgene
Acid C l i n ical la boratory scientists Needlestick Safety and Prevention Act
Alcohols C l i n ical la boratory technicians of 2000
Aldehyde C l i n ical pathology Neutral ization reaction
American Society for C l i nical Col lege of American Pathologists Normal ity
Pathologists Com m ission on Accred itation of All ied Nucleic acids
Ami nes Health Ed ucation Prog ra ms Outpatient c l i n i c
Anatomic pathology Cova lent bond Pathologist
Anion Ester pH
Aromatic ring Governing board Phenol
Atomic theory Gram per deci l iter concentration Phlebotomists
Automated pi pettes Haza rd com m u n ication Physicians' office la boratories
Bala nces Hazardous chemica l s Pi pettes
Base Hematology Proficiency testing
Beer's law Hyd rocarbons Protein
Biochemistry I nternational u n its Pyrex
Blood bank Ionic bond Reagent-g rade water
Bloodborne pathogens Ions Reagents
Board of Reg istry Ketone Reference la boratories
Carbohyd rates La boratory manager Serial d i l ution
Cations Lipids Serologic g lass pi pette
Centers for Disease Control and Medical laboratory assistants Sta ndard cu rve
Prevention Medical staff Sterols
Centrifuge Medical technologist The Joint Com m ission
Chemical sym bols Microbiology department Vol u metric pi pette
C l i n ical chemistry Molal ity Va lence
Molarity

Points to Remember • Hospitals have an organizational structure consisting of a

• The American Society for Clinical Pathologists (ASCP)
was formed in 1 922 to meet the needs of the growing
pathology profession.
• The ASCP created the Board of Registry in 1928 to cer­
tify laboratory technicians and then the Board of Schools
to accredit laboratory training schools.
• In 1933, clinical laboratory technicians formed a pro­
fessional society, the American Society for Clinical
Laboratory Technicians, to provide autonomy and a
voice for the growing profession of clinical laboratory
science.
• Laboratories produce 80% of the objective data that health
care providers use to diagnose and rule out diseases, and
they provide blood for transfusion and determine the sus­
ceptibility of pathogenic bacteria to antibiotics.
• Clinical laboratories began as part of a hospital in the
early 20th century and remain a critical part of hospitals
today.
governing board, medical staff, and management.
• Anatomic pathology comprises surgical pathology, his­
tology, and cytology.
• Clinical pathology is the largest portion of the clinical
laboratory, and it is composed of hematology, clinical
chemistry, microbiology, immunohematology, toxicol­
ogy, immunology and serology, urinalysis, specimen col­
lection, and customer service.
• Pathologists are medical doctors who oversee laboratory
testing.
• A laboratory manager is responsible for the daily activi­
ties of the laboratory.
• Clinical laboratory scientists possess a bachelor's degree
in clinical, medical, or laboratory science; 3 years of aca­
demic course work; and 6 months to 1 year of clinical
experience.
• Clinical laboratory technicians or medical laboratory
technicians have a 2-year associate degree, and they per­
form all the routine testing in the laboratory.

• Medical laboratory assistants are trained to perform or
assist in performing routine laboratory testing allowed by
law and administrative tasks.
• Phlebotomists draw blood from patients.
• An outpatient clinic or a physician's office is a location
where patients receive medical care.
• Public health laboratories are responsible for health refer­
ence tests; disease prevention, control, and surveillance;
population-based interventions; and emergency response
efforts.
• The Department of Defense operates many clinical labo­
ratories across the world.
• Federal regulations that affect clinical laboratories include
the Clinical Laboratory Improvement Act (CLIA) of
1 967 and 1 988, the Needlestick Safety and Prevention
Act of 2000, and regulations for bloodborne pathogens,
hazardous chemicals, and hazard communication.
• The Health Insurance Portability and Accountability Act
affects the laboratory as it relates to patient privacy.
• Accreditation is a voluntary process with which laborato­
ries maintain standards of quality.
• The Joint Commission accredits hospitals and many
other health care organizations.
• The College of American Pathologists is an internation­
ally known agency that accredits clinical laboratories.
• Competency testing involves testing the ability of the
laboratory professionals that perform the diagnostic
tests.
• Characteristics of glassware include thermal durability;
alkali, zinc, or heavy metal content; chemical stability;
electrical conduction; optical qualities; and color.
• Plasticware can be made from polystyrene, polypropyl­
ene, polycarbonate, Teflon, and nylon.
• The four basic types of centrifuges are horizontal head
or swinging bucket, angle-head or fixed angle, axial, and
ultracentrifuge.
• Pipettes are classified as manual, semiautomated, and
automated.
• The volumetric pipette is a long glass tube with a bubble
in the middle.
• There are two types of serologic pipettes-those used to
deliver and to contain.
• Reagents must be monitored for reliability and repro­
ducibility.
• To ensure high-quality laboratory results, high-quality
chemicals and high-quality water must be used.
• The term gram molecular weight is often used as a defini­
tion of mole.
• Molarity = (grams of compound/gram molecular weight)/
liters of solution.
• The molal concentration of a solution is equal to the
number of moles of solute per 1 000 g of solvent.
• The definition of normality is 1 gram equivalent weight
of a compound dissolved in a liter of solution.
• The g/dL concentration is defined as the number of
grams of a com pound dissolved in 1 00 mL of water.
• A percent (o/o) solution can be written as g/dL or go/o.
CHAPTER 1 Laboratory Essentials
• volume1 x concentration1 = volume 2 x concentration 2 .
• Remember:
grams ---> milligrams, multiply by 1 000
decigrams ___, milligrams, multiply by 1 00
centigrams ___, milligrams, multiply by 1 0
mm 3 ---> m L (cc), divide by 1 000
milligrams ___, grams, divide by 1 000
• Conversion of Celsius to Fahrenheit: C = 5/9 x (F 32) -
• Conversion of Fahrenheit to Celsius: F = (9/5 x C) + 32
• Conversion of Celsius to Kelvin: K = C + 273 .
• The amount of dissociation that occurs and the number
of hydrogen ions (H+) in the solution correlate with the
strength of the acid and the pH of the solution.
• Beer's law: A = 2 log o/o T
-
• Aunknown/Asrandard = Cunknown / C srandard
• Standard curves are constructed by plotting points for at
least three standards for a test procedure.
Introduction
This chapter provides a short history of the clinical labora­
tory, various practice sites for laboratories and their organi­
zational structures, levels oflaboratory personnel, laboratory
departments, and accreditation agencies. Chemistry princi­
ples and essential laboratory mathematics are also reviewed.
History of Clinical Laboratories
The first clinical laboratory in the United States opened in
1 896 at Johns Hopkins Hospital. Laboratories were small
rooms with very little equipment where pathologists per­
formed tests on patients' specimens. After the discovery of
causative agents of tuberculosis, diphtheria, and cholera,
laboratories became more important in medicine. As the
volume of laboratory tests increased, pathologists trained
young women to perform some of the simpler laboratory
tests to free the pathologist to do more complex testing.
The American Society for Clinical Pathologists (ASCP)
was formed in 1 922 to meet the needs of the growing pathol­
ogy profession. In 1 926, the accrediting body for hospitals,
the American College of Surgeons, mandated hospitals to
have a pathologist on staff. During World War I, hospi­
tals experienced a critical shortage of laboratory assistants.
Pathologists viewed this as an opportunity to standardize
educational programs for laboratory assistants, now called
technologists or scientists. To meet this need, the ASCP cre­
ated the Board of Registry in 1 928 to certifY laboratory
workers and the Board of Schools to accredit laboratory
training schools. When an individual completed an accred­
ited program, she could take the Board of Registry exami­
nation. Successful completion of the examination conferred
the ASCP tide of medical technologist (MT) .
The ASCP played a major role in the formation of the
clinical laboratory science profession by approving edu­
cation programs and certifYing laboratory workers. The
National Credentialing Agency (NCA) was an independent
4 PA R T 1 Laboratory Principles
certification agency created by laboratory professionals in
the 1 970s to credential laboratory professionals. The ASCP
Board of Registry and the NCA merged in 2009 to create
the ASCP Board of Certification.
Another organization that certifies laboratory profession­
als and other medical professionals is the American Medical
Technologists (AMT) . The AMT was founded in 1 939 and
is a nationally and internationally recognized certification
and membership society for medical technologists, medi­
cal laboratory technicians, phlebotomy technicians, medical
laboratory assistants, clinical laboratory consultants, medi­
cal assistants, medical administrative specialists, dental assis­
tants, and allied health instructors.
In 1 933, clinical laboratory technicians formed a profes­
sional society, the American Society for Clinical Laboratory
Technicians, to provide autonomy and a voice for the grow­
ing profession of clinical laboratory science. Years later, the
organization changed its name to the American Society for
Medical Technology and then to the American Society for
Clinical Laboratory Science (ASCLS) .
In the 1 940s and 1 950s, clinical laboratory testing analyzed
specimens such as blood and urine. Laboratories also housed
and used animals in the test procedures. An example is the
pregnancy test where urine from a woman suspected of being
pregnant was injected into a rabbit. After a specific time period,
the rabbit's ovaries were examined for ovulation. If the ovaries
were swollen and ovulating, the woman was pregnant. In the
1 960s, laboratories used frogs to detect pregnancy in women.
By the 1 970s, more reliable and valid test procedures were
introduced into the clinical laboratory for pregnancy testing.
More sensitive test procedures were introduced in the 1 970s
(e.g., radioimmunoassay) and 1 980s (e.g., enzyme immunoas­
says) . Bioluminescence assays attained widespread use in the
1 990s. As more sensitive test procedures were introduced in the
clinical laboratory, more test analyses were added.
Types of Clinical Laboratories
Clinical laboratories are a dynamic area in health care. Lab­
oratories produce 80% of the objective data that health care
providers use to diagnose and rule out diseases, to provide
blood for transfusion, and to determine the susceptibility of
pathogenic bacteria to antibiotics. Clinical laboratories are
found in hospitals, outpatient clinics, and physicians' offices
and as stand-alone reference laboratories. Laboratories are
constantly integrating new technology and instruments to
better meet the needs of health care providers and patients.
The following sections describe the types of clinical labora­
tories, structures of organizations and laboratories, labora­
tory personnel, and laboratory departments.
I n patient la boratories
Clinical laboratories began as part of a hospital in the early
20th century and remain a critical part of hospitals today.
Although the clinical laboratory may be located in the hos­
pital, work from outpatient clinics, nursing homes, and
other settings may be sent to the hospital's clinical labora­
tory for analysis. Clinical laboratory workers are hospital
employees, and they are an important part of the health
care team.
Organizational Structure
Hospitals are an invention of the 20th century. Hospitals
were known as almshouses before the 20th century. Alms­
houses were places where poor people or people without fam­
ily members to care for them would go to receive care. These
facilities provided food, shelter, and rest. Before the 20th cen­
tury, the best medical care was received at home; even opera­
tions were performed in the home. As medical procedures
and equipment became more advanced, the patient went to
see the doctor instead of the doctor coming to see the patient.
Hospital
There are approximately 6500 hospitals in the United States.
They are classified as public, private, specialty, community,
federal, military, or other types.
Hospitals are organized in three distinct parts: govern­
ing board, medical staff, and management. The governing
board is the body responsible for the financial health of the
organization and for setting institutional policies and goals.
The governing board appoints the medical staff as the party
responsible for quality patient care.
The medical staff members of the hospital are not usu­
ally considered to be employees; however, more hospitals
and hospital systems are employing health care providers.
In the traditional structure, the medical staff is granted
the right to admit patients and perform procedures in the
hospital.
The management portion of the hospital consists of
the hospital administrator as the chief executive officer
who is responsible for managing all hospital departments.
Figure 1 - 1 shows the relationships among the three parts
of the hospital and shows where the laboratory fits into the
organizational structure.
Clinical Laboratory
Clinical laboratories are composed of many different depart­
ments. The laboratory services department is usually sepa­
rated into anatomic and clinical pathology. The anatomic
pathology department examines all tissues, fluids, organs,
and limbs removed from the body. This discipline com­
prises surgical pathology, histology, and cytology. Personnel
in the anatomic pathology department include pathologists,
pathologists' assistants, histology technicians, and cytology
technicians. In the anatomic pathology department, tis­
sues are described by pathologists, cut into sections, fixed
with chemicals, sliced very thin, placed on glass slides, and
stained with special chemicals. After the slides are stained
and cover slipped, the pathologist examines the tissue for
abnormalities.
Clinical pathology is the largest portion of the clinical
laboratory. This section is composed of hematology, clinical
chemistry, microbiology, immunohematology, toxicology,

MediAdmicalsRecords
sions ClRadi
i n icoalloLab
I maging Rehabil itation
• Figure 1-1 Hospital organ izational chart.
Hematology
• Figure 1-2 Clinical laboratory organizational chart .
immunology and serology, urinalysis, specimen collection,
and customer service. The individual laboratory sections are
described later (Fig. 1 -2) .
Departments and Their Functions
Clinical Chemistry
Clinical chemistry is the medical discipline that uses various
methods of analysis and instrumentation to determine val­
ues for chemical components in normal and diseased states,
types and concentrations of blood toxins, and therapeutic
drug levels. Routine tests run by the clinical chemistry section
analyze levels of glucose, blood urea nitrogen (BUN), electro­
lytes, calcium, phosphorus, magnesium, lipids, liver function
values, alkaline phosphatase, creatinine kinase, creatinine,
protein, albumin, and hemoglobin A1c. The clinical chem­
istry department also runs hepatitis panels, tests for rubella
and human immunodeficiency virus (HIV) , and determines
levels of antibodies in the blood. Hormone levels (e.g., thy­
roid-stimulating hormone, prolactin, follicle-stimulating
hormone) are tested in another section of this laboratory.
The routine tests are usually run in the main clinical
chemistry department. The antibody and hormone levels
are usually considered subspecialties. Other subspecialty
departments include the toxicology, therapeutic drug mon­
itoring, molecular diagnostics, and fecal analysis. Some
clinical chemistry laboratories have a section that analyzes
blood gases.
CHAPTER 1 Laboratory Essentials
AdmiHospi tal
n istrator
BusiOffincesse
gy Respi ratory
Pharmacy AdmiFaci
n i s ies ve Bil ing CodiAccounti
l ittrati &
ng ng
Support Human Resources
Immunol
Serologyogy/ Microbiology Toxicology II Uri nalysis
Hematology
Hematology is the study of blood cells. Blood cells include
erythrocytes (i.e., red blood cells) , leukocytes (i.e., white
blood cells) , and thrombocytes (i.e., platelets) . The most
common test performed in this department is the com­
plete blood count (CBC) , which is a summary of cell
counts (i.e., red, white, and platelet) , total hemoglobin
level, red blood cell size, and hematocrit. A CBC usually
includes a differential count, which reports the percentage
of each type of white blood cell in the blood sample. Cell
counts for body fluids are also performed in this depart­
ment. Other tests include reticulocyte counts and erythro­
cyte sedimentation rates.
In many laboratories, coagulation testing is performed
in the hematology department. Routine coagulation tests
include the prothrombin time (PT) and the activated par­
tial thromboplastin time (aPTT) . These tests assess the two
major clotting pathways in the body.
Microbiology
The microbiology department identifies microorgan­
isms that cause disease and determine the most effective
antibiotic to destroy bacterial pathogens. This department
grows cultures from major body systems such as the throat,
urine, stool, wound, blood, eyes, ears, body fluids, nasal,
abscesses, vagina, urethra, and tissues. Surgeons often per­
form a culture after they drain or debride an infected area.
6 PA R T 1 Laboratory Principles
Routine cultures include aerobic and anaerobic incubation
environments. This department also performs identifica­
tion or presumptive identification of fungi, parasites, and
bacteria.
Specim en Collection
The specimen collection department collects tissue, blood,
and urine samples from patients. In the outpatient area of
the laboratory, phlebotomists also educate patients about
collection of 24-hour urine, fecal fat, dean-catch urine, and
other specimens.
U rinalysis
The urinalysis department performs chemical tests on urine
specimens and analyzes formed elements that may be present
in specimens. Urine is tested for color, clarity, specific grav­
ity, glucose, protein, ketones, occult blood, and pH. These
tests are used to monitor metabolic diseases such as diabetes.
Blood Bank
The blood bank or immunohematology department tests
red blood cells from donors for antigens and serum from
recipients for antibodies. Testing ensures that people receive
compatible units of blood during a transfusion. The blood
bank also transfuses other blood components such as plate­
lets, fresh frozen plasma, and specific clotting factors.
I m m unology and Serology
When invaded by microorganisms or other foreign bodies,
the human body produces antibodies to protect itself from
the threat. The immunology and serology department tests
blood for antibodies produced against pathogenic microor­
ganisms. Detection of antibodies against a particular patho­
gen affects the diagnosis and treatment of the disease, such
as hepatitis B virus and HIV infections. The department
also tests for abnormal configurations of antibodies.
Much testing is performed across laboratory departments.
For example, molecular diagnostics can be performed in a
microbiology laboratory to test for specific viruses and other
microorganisms. Serology and immunology testing may be
performed in the chemistry department. To increase labora­
tory efficiency, many large laboratories have a core labora­
tory. The composition of a core laboratory varies according
to the needs of the institution and its clients. One possible
configuration uses a menu of testing services for general
chemistries, hematology, coagulation, blood gases, thera­
peutic drugs, endocrine profiles, emergency toxicology, and
drugs of abuse. It usually includes automated analytic sys­
tems and specialized information management for critical
care testing on a 24-hour basis.
Technical Personnel
Laboratory workers include pathologists, laboratory manag­
ers, clinical laboratory scientists, clinical laboratory techni­
cians, medical laboratory assistants, and phlebotomists. The
educational requirements and duties of each type of worker
are discussed in the following sections.
Pathologist
A pathologist is a medical doctor who examines tissues and
oversees the quality of laboratory test results from a clini­
cal laboratory. Pathologists must complete medical school,
an accredited student resident program, and an approved
residency.
Pathologists are responsible for analyzing tissue samples
(e.g. , looking for cancer cells) and interpreting the mean­
ing of laboratory test results. They consult with treating
physicians to determine diagnostic and follow-up tests for
patients. They are also responsible for performing autopsies.
Anatomic pathologists assist surgeons by examining biop­
sies during surgery to produce an immediate diagnosis. This
helps the surgeon to determine whether additional tissue
must be removed from the patient's body to eradicate dis­
ease. Clinical pathologists oversee testing of body fluids and
confirm cellular identification in the hematology laboratory.
The clinical pathologist also consults with physicians about
blood transfusions and antibiotic treatment of bacterial and
other infections. Forensic pathologists examine evidence to
provide information for criminal and civil law cases.
Laboratory Manag er
A laboratory manager is responsible for the daily activities
of the laboratory. He or she has at least a bachelor's degree
and is a clinical laboratory scientist. The person is respon­
sible for the laboratory workers conducting tests and report­
ing test results.
Clinical Laboratory Scientists
Clinical laboratory scientists (CLSs) are also known as
medical laboratory scientists (MLSs) or medical technolo­
gists (MTs) . They perform routine and specialized labora­
tory tests. They also troubleshoot problems with specimens,
procedures, and instruments to ensure quality test results.
They examine blood and body fluids under the microscope
for microorganisms and possibly even cancer. These workers
communicate laboratory results to physicians and patholo­
gists. Clinical laboratory scientists train new employees,
perform quality control procedures on analytic test runs,
and evaluate instruments and new procedures. These indi­
viduals may also advance to department supervisors, techni­
cal supervisors, or the laboratory manager. They can also
choose to specialize in disciplines such as clinical chemis­
try, immunology, molecular pathology, microbiology, and
blood bank services.
CLSs possess a bachelor's degree in clinical or medical
laboratory science, 3 years of academic course work, and
6 months to 1 year of clinical experience. This is the most
common route to certification. Several other routes combine
education with experience that can be used to become certi­
fied. Most employers require CLSs to obtain a certification
from the ASCP Board of Certification (BOC) or the AMT.
Clinical Laboratory Technicians
Clinical laboratory technicians (CLTs) or medical labora­
tory technicians (MLTs) possess a 2-year associate degree,
 CHAPTER 1 Laboratory Essentials

and they perform all the routine testing in the laboratory.
CLTs who graduate from accredited programs are able to sit
for the national certification examination offered through
the BOC. CLTs use microscopes and all of the instrumenta-
tion in a clinical laboratory. CLTs also specialize in the same
disciplines as the CLSs.
Medical Laboratory Assistants
Medical laboratory assistants (MLAs) are trained to per-
form or assist in performing routine laboratory testing as
allowed by law and to perform administrative tasks. Some
MLAs also have duties involving patient contact. Most of
these professionals receive on-the-job training, but some
graduate from short-term educational programs accredited
by the Commission on Accreditation of Allied Health
Education Programs (CAAHEP) or the Accrediting
Bureau of Health Education Schools (ABHES).
Phlebotomists
Phlebotomists draw blood from patients. Usually, CLSs
and CLTs are also trained to draw blood as part of their
education. It is more cost effective to hire phlebotomists to
draw blood and have the CLSs and CLTs perform laboratory
tests . Phlebotomists are high school graduates with specific
training in phlebotomy. The ASCP BOC offers a certifica-
tion examination for phlebotomy technicians (Table 1-1 ).
~ ublic ea di laboratories are operated at a state level. The
Outpatient Clinics and Physicians' Office
Laboratories
An outpatient clinic or a doctor's office is a location
where patients receive medical care. Clinics usually ,Have
small laboratories that perform routine tests as allowed
by law. Physicians' office laboratories (POLs) range
from a small laboratory (for one to five physicians) that
performs a few tests to laboratories with a large volume
(500,000 tests per year) chat serve up to 200 physicians.
The large POL is usually the exception. POLs are defined
as a laboratory that performs tests in a physician office
setting, provides results to be used during the office visit,
and performs tests to be used for screening, diagnosis, and
monitoring.
Reference Laboratories
Reference laboratories are independent, commercial, large
laboratories chat perform routine and specialty testing.
POLs, nursing homes, and hospital laboratories send labo-
ratory testing to these facilities. Reference laboratories have
specialized equipment and perform low-volume specialized
tests. Reference laboratories usually have drawing stations
located in convenient locati0ns for patients.
State and Federal Laboratories
or isease Control and Prevention (CDC)
operates o , cl two biosafety level 4 laboratories in the
United States.it is an example of a federal laboratory. Many
tW.v0rk of public health laboratories plays a vital role in
kee2ing Americans healthy. Public health laboratories are
resp~ sible for performing public health reference tests; dis-
c.s:ase prevention, control, and surveillance; population-based
interventions; and emergency responses.

■ Laboratory Professionals' Profile

Laboratory Education
Professionals Where They Work Special Skills Required

Laboratory director Hospitals, reference Attention to detail, big PhD or MD

laboratories, pharma- picture; good communicator,
physicians ceutical companies planner, leader

Clinical laboratory Performs routine and Hospitals, reference Problem solver, troubleshooting Bachelor's degree
scientist (CLS) complex tests laboratories, clinics skills, attention to Licensure or
Performs quality control detail, organized, good certification
time management

Clinical laboratory Performs routine tests Hospitals, reference Good coordination, ability to Associate degree
technician (CLT) Performs quality control laboratories, clinics manipulate small objects, Licensure or
with supervision attention to detail, computer certification
literate

Clinical laboratory Performs or assists with Hospitals, reference Good coordination, ability to On-the-job training
assistant (CLA) routine laboratory tests laboratories, clinics manipulate small objects, or completion
as allowed by law attention to detail, computer of a short-term
literate program
Phlebotomist Collects blood specimens Hospitals, reference Good coordination, ability to On-the-job training
from patients laboratories, clinics manipulate small objects, or completion
attention to detail, computer of a short-term
literate program
8 PA R T 1 Laboratory Principles
M i l ita ry Laboratories
The Department of Defense operates many clinical labo­
ratories across the world. Military hospitals perform rou­
tine laboratory testing and are accredited by the College of
American Pathologists (CAP) . The very large military hospi­
tals perform routine tests for the physicians assigned to that
hospital and specialized tests for other military hospitals
around the world.
Military hospitals operate American hospitals to treat mili­
tary members and their dependents. Military hospitals have
laboratory officers and medical laboratory technicians staffing
the clinical laboratory. Laboratory officers have at least a bach­
elor's degree and CLS certification, and the enlisted members
serve as medical laboratory technicians and are graduates of
the service's medical laboratory technician school.
Regulation and Accreditation of Clinical
Laboratories
Federal regulations and accreditation agencies govern the
operation of clinical laboratories. Federal regulations that
affect clinical laboratories include the Clinical Laboratory
Improvement Act ( CLIA) of 1 967 and the Clinical Labo­
ratory Improvement Amendments of 1 988, the Needle­
stick Safety and Prevention Act of 2000, and those for
bloodborne pathogens, hazardous chemicals, and hazard
communications. The regulations concerning safety are
discussed in Chapter 2, and CLIA is discussed in the next
section. The Health Insurance Portability and Accountabil­
ity Act affects the laboratory as it relates to patient privacy.
Reg u lation
Congress first passed the CLIA in 1 967. The purpose of
this Act was to regulate clinical laboratories involved in
interstate commerce. Hospital and reference laboratories
were the only clinical laboratories affected by the Act. In
1 988, Congress passed regulatory amendments to the Act
in response to public concern about the quality of Pap
smears. The provisions of CLIA 1 988 govern the activities
of all laboratories. It was designed to enhance the quality
of laboratory services provided to all patients by mandating
quality control, quality assurance, and proficiency testing.
Trained personnel were required to perform particular levels
or complexities of tests. The more complex tests a labora­
tory performs, the higher the standards required for the per­
sonnel working in that laboratory. If a laboratory performs
only simple tests, the laboratory can obtain a certificate of
waiver. Laboratories performing "waived" tests are exempt
from proficiency testing requirements under CLIA.
Accred itation
Accreditation is a voluntary process by which laborato­
ries maintain certain standards of quality. Two accrediting
agencies have been given "deemed status" by the federal
government's Centers for Medicare and Medicaid Services
(CMS) . If laboratories are accredited by either agency, the
laboratory does not need to be inspected by the Department
of Health and Human Services. The two accrediting agen­
cies are The Joint Commission and the College of Ameri­
can Pathologists (CAP) .
The Joint Commission
The Joint Commission (formerly known as the Joint Com­
mission for the Accreditation of Healthcare Organizations
[JCAHO]) accredits hospitals and many other health care
organizations, such as ambulatory care facilities, stand­
alone surgery centers, long-term care facilities, behavioral
health centers, and laboratories. A team of individuals
from peer institutions that are accredited by The Joint
Commission visits an institution seeking accreditation or
reaccreditation. These site visitors examine each standard
and the evidence compiled by the institution for com­
pliance with the standard. Institutions must also collect
data on core measures (ORYX) and must comply with the
National Patient Safety Goals annually issued by The Joint
Commission. The Joint Commission accepts accreditation
by the CAP as evidence of compliance with a good portion
of laboratory standards.
College of American Pathologists
The CAP is an internationally known agency that accredits
clinical laboratories. Clinical laboratory professionals per­
form inspections at clinical laboratories using accreditation
checklists developed by CAP. CAP strives for excellence
well beyond regulatory compliance to assist physicians in
providing the best patient care possible. The foundation of
CAP accreditation is rigorous accreditation standards that
are molded into specific, comprehensive checklists. The
inspection team uses the checklists to analyze laboratory
operations.
Proficiency Testing
Proficiency testing is required by CAP, The Joint Commis­
sion, and the federal government through CLIA 1 988. Pro­
ficiency testing is a process in which a laboratory is provided
samples to analyze with a regular run. These samples are
provided for every department in the laboratory that per­
forms diagnostic tests. The laboratory analyzes the samples
and then sends the results back to the agency that provided
the samples. The agency analyzes the laboratory's results and
provides the analysis to the laboratory. This process tests the
accuracy of laboratory results being produced in that labora­
tory. Excellent clinical laboratories must produce accurate
and reliable laboratory test results.
Competency Testing
Competency testing involves testing the ability of the labo­
ratory professionals who perform the diagnostic tests. This
must occur yearly to ensure that individuals performing
diagnostic tests are well trained and competent.

Laboratory M aterials
Laboratory professionals use many types of equipment and
chemicals in the laboratory. The following sections describe
common, nonautomated equipment and chemicals used in
the laboratory.
Glasswa re and Plasticwa re
All glassware is not made the same and has different char­
acteristics for different purposes. Characteristics of glass­
ware include thermal durability; alkali, zinc, or heavy metal
content; chemical stability; electrical conduction; optical
qualities; and color. Pyrex can be used in high-temperature
experiments, and it is heat shock resistant. Other qualities
of Pyrex include acid resistance and a low alkali content,
which is good for high-purity laboratory work. The name
probably looks familiar because Pyrex glassware is used for
home baking.
Many types of plasticware are sold for laboratory use.
Nalgene is a leader in providing high-quality plasticware
to laboratories. Plasticware can be made from polystyrene,
polypropylene, polycarbonate, Teflon, and nylon. Many
types of plasticware are biologically inert, chemically resis­
tant, break resistant, and durable. Because breakage is less
of an issue than when working with glassware, plasticware
makes good laboratory equipment.
Cleanliness of laboratory equipment is extremely critical
because contaminants residing in a piece of glass or plastic­
ware can severely disrupt the next analysis performed. All
glass and plasticware should be rinsed thoroughly after use
with water and a mild detergent solution. After using the
detergent, the item should be rinsed thoroughly with water.
If using a dishwasher to clean glass and plasticware, follow
manufacturer's guidelines for the best results.
Centrifuges
A centrifuge is a piece of motorized equipment that uses
centrifugal force to separate a mixture such as clotted blood.
There are four basic types of centrifuges: horizontal head
or swinging bucket, angle-head or fixed angle, axial, and
ultracentrifuge. Centrifuges can be small enough to set on a
bench top or large enough to stand alone on the floor. They
can be refrigerated or nonrefrigerated. They can have small
openings for placing test tubes or large openings for placing
a unit of blood.
Uses for Centrifuges
There are many uses for centrifuges in a clinical labora­
tory. Blood specimens are spun down in a centrifuge to
separate the red blood cells from the serum or plasma.
Urine specimens can be poured into a disposable plastic
tube and spun down in a centrifuge to concentrate the
nonliquid material that may be present in the urine speci­
men. Antibodies and antigens can be separated through
centrifugation.
CHAPTER 1 Laboratory Essentials
Periodic Maintenance
New centrifuges should be calibrated before they are put
into service in the laboratory and after repair. Centrifuges
should spin at the speed recommended by the manufacturer
because spinning too fast can lyse or break apart red blood
cells, and spinning too slowly can fail to adequately con­
centrate materials in a urine or other specimen. The speed
should be checked approximately every 3 to 6 months using
an external tachometer.
The timer should also be checked for accuracy periodi­
cally. If the centrifuge is refrigerated, the temperature should
be checked and recorded monthly. The temperature should
fall within the manufacturer's guidelines.
Bala nces
Types of Balances
Balances are devices used to accurately weigh substances.
There are two designs for balances: double pan and single
pan. The double pan balance has a single beam with two
arms of equal length. The single pan balance has arms of
unequal length. Both types of balances can be mechanical
or electronic. Balances should be placed in a vibration-free
and airflow-free area away from centrifuges.
Analytical balances are used in laboratories for preci­
sion measuring in weighing substances requiring 0 . 1 -mg to
1 0-).lg readability. Analytical balances can be electronic or
manual. Types of electronic balances are the electromagnetic
balancing or electrical resistance wire. Although they are
based on different principles, neither type of balance directly
measures mass. Instead, they measure the force that pushes
the pan downward. This force is converted to an electrical
signal, and the signal on the digital display is interpreted
as the mass of the object on the pan. The electromagnetic
balancing principle uses a magnet and a coil to generate an
electromagnetic force that is converted to an electronic sig­
nal and interpreted as mass. The electrical resistance wire
uses the change in resistance of a wire that is attached to a
piece of metal that bends when a force is applied. Balances
use reference weights to calibrate the output, which corre­
lates force to a particular number of grams.
Periodic Maintenance
Analytical standard weights are used to verifY the accuracy
of balances. The National Institute of Standards and Tech­
nology (NIST) recognizes five different classes of analytical
weights: M, S, S- 1 , P, and J. Class M weights are designated
as primary standard quality and are used to calibrate other
weights. Usually laboratories use class S weights to verifY the
accuracy of balances for weights between 1 00 g and 1 mg.
Pi pettes
Pipettes are devices used to transfer a specific amount
of a liquid to another container. Pipettes are classified as
manual, semiautomated, and automated. The two types
of manual pipettes are volumetric (i.e., transfer) and sero­
logic (i.e., measuring) . Semiautomated pipettes can have a
10 PA R T 1 Laboratory Principles
fixed volume or variable volume. These pipettes use plastic,
disposable pipette tips to draw up and dispense the liquid.
Semiautomatic pipettes are especially useful for transferring
extremely small volumes of liquids, such as 1 0 J..LL , 5 J..LL ,
1 00 J..LL , or 200 J..LL
Automated pipettes are usually electronic, computer­
ized pipettes that control the amount of liquid aspirated and
the amount of time allowed for aspirating and dispensing
liquids. All types of pipettes used in the laboratory must be
routinely calibrated to ensure accuracy. The manufacturer's
instructions provide details on calibration.
Volumetric Pipettes
The volumetric pipette is a long glass tube with a bubble in the
middle. The liquid being transferred is drawn up in the pipette
until it reaches an etched mark on the pipette. This mark indi­
cates the exact volume for the pipette. Volumetric pipettes
come in different sizes, and each pipette has only one volume.
Serologic Pipettes
The serologic glass pipette is etched with gradations so that
different amounts can be delivered with the same pipette.
There are two types of serologic pipettes: "to deliver" and "to
contain." "To deliver" pipettes retain some liquid in the tip
after the specified amount of liquid has been delivered. The
"to contain" pipettes require the liquid that remains in the
tip after delivery to be pushed out of the pipette for accurate
delivery.
Reagents
Reagents are chemical solutions that are used in diagnos­
tic tests. They are usually liquid, lyophilized, or frozen.
Reagents come in various purity states. Because there is no
agreement about the purity of a reagent, the standards put
forth by the American Chemical Society (ACS) are used to
determine reagent or analytical reagent grade. ACS chemi­
cals are considered to have very high purity and to be suit­
able for quantitative analyses.
Reagents must be monitored for reliability and reproduc­
ibility. The U.S. Food and Drug Administration Department
of Biologics enforces tough federal regulations to ensure
quality. Laboratories must be vigilant and verify the integ­
rity of purchased reagents. When changing lots of reagents,
the laboratory must perform parallel testing to ensure reli­
able results. Laboratories develop operating instructions for
performing this function.
Water
Water is a common substance with many laboratory uses.
Drinking water contains many impurities that can affect
laboratory test results. To ensure high-quality laboratory
results, high-quality chemicals and high-quality water must
be used.
Several methods are used to produce water that is free
of impurities and suitable for laboratory use. The methods
are discussed in great detail in the Clinical Laboratory Stan­
dards Institute (CLSI) guideline, Preparation and Testing of
Reagent \Vtzter in the Clinical Laboratory: Approved Guide­
line, 4th edition.
The most common purification processes used in clini­
cal laboratories include distillation, deionization, reverse
osmosis, and ultrafiltration. Distillation is a good pro­
cess for removing particulates and some dissolved con­
taminants. It is less effective at removing dissolved ions.
Deionization involves passing water through cation- and
anion-exchange resins. This is an excellent method for
removing ions, and when coupled with a carbon filter, most
dissolved organic compounds can be removed. This pro­
cess is less effective at removing particulate matter. Reverse
osmosis involves forcing water under pressure through a
semipermeable membrane. The semipermeable membrane
filters out dissolved organic, ionic, and particulate impu­
rities. This method is less effective at removing dissolved
gases. Ultrafiltration involves passing water through semi­
permeable membranes (i.e., pores less than 0.2 mm) to
remove most particulates from the water. It does not do
a good job of removing dissolved solids and gases. Most
laboratories choose water filtration systems that produce
the best water possible for its use.
There are three types of reagent-grade water. Type I
reagent-grade water is the highest quality water, and it is
used in test methods requiring minimal interference and
maximum sensitivity. Type II water is used for general lab­
oratory testing. Type III water is used for the initial rins­
ing and washing of glassware. The CLSI standard bases the
purity of reagent-grade water on microbiology content (col­
ony forming units per ml) , pH, resistivity, silicates, organ­
ics, and particulate matter. Water used for most routine
clinical laboratory testing is defined as clinical laboratory
reagent water by CLSI and has a resistivity of at least 1 0 mQ
· em at 25o C.
Chemistry Review
A clinical laboratory analyzes specimens from the human
body and other living animals. Clinical chemistry deals with
the concentrations of chemicals and ions in the body and
the changes that occur to these chemicals and ions in nor­
mal and disease states of the body. The following sections
review the chemical principles needed to understand clinical
chemistry.
Atomic Theory
Atomic theory states that all matter is made up of atoms.
Atoms have protons (i.e., positively charged particles [1 +])
and neutrons (i.e., neutral particles) in the center or nucleus
and electrons (i.e., negatively charged particles [ 1 - ] ) that
circle around the nucleus. Electrons are located in specific
areas around the nucleus called electron shells. The shells are
located a specific distance from the nucleus. Smaller shells
are located closer to the nucleus of the atom, and larger

shells are located farther away from the nucleus. Scientists
think there are up to seven electron shells surrounding the
nucleus. In most cases, electrons fill or partially fill the lower
energy level electron shells before filling the higher energy
level shells.
Various atoms have different numbers of protons, neu­
trons, and electrons. The outermost shell containing elec­
trons is called the valence shell. Electrons located in the
valence shell are usually involved in bonding with other
atoms to produce chemical compounds.
The valence of an atom is the number of electrons that
can be lost, gained, or shared by an atom when forming
a compound. If the atom gains electrons (- 1 charge), the
atom's valence is negative. If the atom loses electrons, the
atom's valence is positive. As a rule, when 2 atoms combine
to form a molecule, the sum of the valences of the atoms
is zero. The resulting molecule is considered to be neutral.
For example, hydrogen and oxygen combine to make water.
The valence of hydrogen is + 1 , and the valence of oxygen is
- 2. The result of combining 1 hydrogen atom and 1 oxygen
atom is a molecule with a valence of - 1. Another hydrogen
atom is needed to form the neutral molecule of water (H 2 0) .
When an atom loses or gains electrons, it becomes an ion.
Ions are charged atoms. If a hydrogen atom loses its elec­
tron, it becomes a positively charged ( + 1) ion, also known
as a cation. If the oxygen atom adds 2 electrons, it becomes a
negatively charged ( - 2) ion, also known as an anion. Oppo­
sitely charged atoms attract each other, and this force holds
the resulting molecule together. The force that holds atoms
together to form molecules is called a bond.
Chemica l Bonds
Atoms combine through ionic, covalent, coordinate cova­
lent, nonpolar covalent, and polar covalent bonds. In an
ionic bond, one atom transfers its electrons to another
atom. The atoms in this molecule each have their valence
shells completed. These atoms are held together with an
electrovalent bond. In a covalent bond, each atom donates
one or more electrons that are subsequently shared between
the two atoms. A coordinate covalent bond is a special case
of a covalent bond in which one atom donates all the elec­
trons to be shared. A nonpolar covalent bond occurs when
both atoms sharing electrons have similar characteristics. A
polar covalent bond occurs when one atom in a molecule is
more electronegative than the other atom. Chemical bonds
play a role in chemical reactions.
Factors Affecting Chemica l Reactions
Many factors affect chemical reactions. Some chemical reac­
tions are reversible, and others are irreversible. Some chemi­
cal reactions go much faster than other chemical reactions.
By understanding the factors that affect chemical reactions,
it is easier to predict the outcome or troubleshoot a prob­
lem. Factors affecting a chemical reaction include tempera­
ture, light, pressure, concentration, and catalysts.
CHAPTER 1 Laboratory Essentials
An increase in temperature causes an increase in the
rate of a chemical reaction. A higher temperature provides
energy for the molecules to move faster and collide more
frequently. Due to the increased collisions, the chemical
reaction rate increases. Conversely, lowering the tempera­
ture slows the chemical reaction rate and the collisions
between the molecules. Light is another form of energy
that can increase the rate of a chemical reaction. When
working with gases, increased pressure adds energy to the
chemical reaction and forces more molecular collisions,
resulting in an increased chemical reaction rate. The con­
centrations of the reactants may also influence the reac­
tion rate.
Important factors affecting a chemical reaction rate are
catalysts. Many biological reactions are extremely slow by
nature and require a catalyst to increase the reaction rate.
Some catalysts are organic and are called enzymes. Enzymes
are a clinically important group of compounds for diagnos­
ing diseases. Chemical reactions are also affected by the con­
centration of the reacting compounds.
Acid, Bases, and Sa lts
Acids, bases, and salts are important compounds in the
body. Acids and bases are produced and used in urine forma­
tion and respiration. Salt is the basis for the blood that runs
through our veins. Understanding the properties of these
substances helps to explain and troubleshoot test principles.
An acid is a substance that donates hydrogen atoms in
a water solution. Acids occur as liquids, solids, and gases.
When a strong acid is mixed with water, the acid completely
dissociates or ionizes. When a weak acid is mixed with water,
the acid partially dissociates or ionizes. Weak acids are used
as buffers to minimize large pH changes with the addition
of strong acids or bases to a system such as blood.
A base is a substance that donates hydroxide (OH - ) ions
in a water solution. Acids donate protons, and bases accept
the protons. Most bases have an -ide suffix: sodium hydrox­
ide (NaOH) , potassium hydroxide (KOH) , or lithium
hydroxide (LiOH) .
A neutralization reaction consists of combining an
acid and a base to produce a salt and water as products.
The hydrogen donated by the acid and the hydroxide ion
donated by the base combine to form water. The other
atoms in the acid and base compounds combine to form
the salt.
Orga nic Chemistry
Organic chemistry is the study of carbon-based compounds.
Carbon is a special compound that can have a valence of +4 or
- 4, meaning that it can donate all four of its electrons or take
on four electrons. Examples of other atoms that are found in
organic molecules include hydrogen, nitrogen, sulfur, chlorine,
bromine, and iodine. The versatility of the carbon atom's bond­
ing creates more than 5 million known organic compounds.
Most organic compounds are held together by covalent bonds.
Another random document with
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jeune fille un regard limpide dont elle se ressouvint plus tard, comme
si, en cet instant-là, un autre « moi » eût pris possession d’elle-
même.
La figure de Julien, longue, plutôt fine que robuste, était dominée
par un front d’une ampleur éclatante. Une force de réflexion
tranquille s’accumulait en ses yeux, des yeux d’un brun clair, devant
qui tout semblait doux et fraternel. Sa moustache n’empêchait pas
de voir au coin de sa lèvre une fossette pleine de grâce. Il avait le
teint vermeil, la main effilée, les signes d’une élégance native qu’un
fond sanguin de vigueur pondérait.
Pauline cependant tourna aussitôt son attention vers Edmée
Rude ; ravie de délier sa langue avec elle, car, depuis sa venue à
Sens, elle vivait sans aucune compagne. Edmée, rose et fluette, le
menton enfoncé dans une étole de fourrure, présentait une vivacité
de minois toute bourguignonne. Pauline se pencha pour baiser les
joues de Marthe, la cadette ; celle-ci, avec un battement de cils, la
dévisageait de son œil hardi, profond.
— La gentille petite sœur que vous avez, dit Pauline bonnement.
— Oui, gentille, même trop, repartit Edmée tandis qu’elle
caressait les cheveux déliés et blonds de Marthe. Elle a de ces idées
parfois qui nous font peur. Hier soir, elle regardait, derrière la vitre,
les étoiles : On ne peut pas les attraper avec des échelles ? nous a-
t-elle demandé. Le bon Jésus saura bien me mener là-haut. Est-ce
qu’il m’y mènera bientôt ? Tu viendras m’y trouver, Edmée, et Julien
aussi. J’aurai des ailes, n’est-ce pas, maman ?
Une surprise altéra le sourire de Pauline ; elle ne pouvait
comprendre cette curiosité du Paradis ; aux premières paroles
d’Edmée, l’obstacle chrétien se posait entre elles. Edmée ne savait
pas encore Pauline irréligieuse ; mais elle devina qu’une chose
inconnue les séparait ; et, sans s’attarder sur des intimités vaines
pour une étrangère, elle lui parla du paysage qu’elles surplombaient,
« bien vilain sous son capuchon gris ».
— C’est au printemps qu’il faudra le voir et à l’automne. D’ici,
vers la mi-octobre, la plaine est délicieuse. Je ne sais si vous êtes
comme moi ; j’aime tant l’automne, l’odeur des feuilles tombées, les
peupliers légers, tout en feu comme des tabernacles !
— Moi, répondit Pauline, toute saison me va ; mais j’adore l’été.
Quand le soleil chante, que les oiseaux chantent, je me sens plus de
cœur à chanter.
— Vous devez être musicienne…
— J’ai de la voix, répondit simplement Pauline, dédaignant de se
faire valoir ; et vous ?
Edmée lui déclara qu’elle se passerait de pain plus volontiers que
de son piano ; son père jouait du violon, son frère, du violoncelle ;
chaque dimanche, après leur promenade, et le soir, de temps à
autre, ils exécutaient des trios.
Cette découverte d’une affinité précieuse charma Pauline
davantage qu’Edmée, parce que sa solitude lui rendait une amie
plus désirable. Tout en causant, elles se dirigeaient vers une butte
d’où, jadis, suivant la tradition, les sentinelles romaines observaient
au loin la vallée.
— Si nous grimpions là-haut, insinua Marthe à sa sœur.
— Allons-y, fit Pauline. Elle entraîna Marthe par une main, Edmée
s’empara de l’autre, et toutes trois prirent leur élan jusqu’au faîte du
glacis ; puis, riant et courant, elles redescendirent.
— Vous êtes, mademoiselle, plus leste que les chèvres, dit à
Pauline M. Rude qui survint avec son fils et M. Ardel.
— J’ai eu des aïeux montagnards, répliqua-t-elle en manière de
badinage, je suis faite pour les cimes !
Julien, au son de ces derniers mots, la fixa, se tut une seconde,
et reprit la conversation qu’il avait entamée sur le livre de M. Ardel.
L’auteur jouissait de s’entendre commenter par ce jeune homme
avec une ferveur ingénue.
— Vous allez me trouver sentimental, poursuivit Julien ; mais un
des traits que j’admire en Saint-Simon, c’est d’avoir ordonné, dans
son testament, qu’on liât après sa mort son cercueil à celui de sa
« chère épouse » par des anneaux et des crochets de fer, afin que
leurs corps fussent unis jusqu’à la Résurrection. Pour ma part, si je
me marie jamais, je ne voudrais qu’un amour de cette trempe, long
et fort comme l’éternité…
Pauline n’entendit pas sans étonnement un langage si nouveau
pour elle ; mais elle s’étonna peu de voir, à mesure que Julien
s’animait, le professeur plisser sa bouche d’une moue d’ironie
sceptique.
— Voyez, dit tout à coup M. Rude, le joli rayon, derrière nous, là-
bas !
En effet, à la chute du jour, tandis que les coteaux de l’Est et la
plaine succombaient sous un brouillard de plus en plus dense où
des cheminées d’usines brandissaient leurs fumées sombres, les
nuées du couchant se fendirent, le soleil apparut, tel qu’un prêtre en
chape rutilante qui s’en va dans l’abside illuminée d’une basilique, et
au-dessus de Saint-Martin un peu de ciel flotta, fugitif et doux. La
coloration de l’air froid communiquait aux visages une sorte de
pureté diaphane. Julien, pour Pauline, en fut transfiguré.
— Voici l’heure, dit-il, que nous aimons en hiver, celle où
s’allument les lampes des boutiques, et les réverbères, un à un,
dans la brume, le long des quais…
— Et l’heure, acheva Edmée, où des étincelles pétillent sur les
fourneaux des marchands de marrons.
Tous rirent de cette saillie et ils reprirent ensemble le chemin de
la ville. Les deux jeunes filles descendaient en avant ; Julien suivait,
et Marthe, lasse de la course un peu longue pour des jambes de
cinq ans, se pendait à sa main. Plus haut, dans l’étroit sentier,
sonnait le pas martelé des deux professeurs ; la grosse voix de M.
Rude roulait comme un grondement. Il expliquait à son collègue
qu’après avoir surveillé cinq ou six heures par jour les barbouillages
de ses élèves, il reprenait, chaque soir, dans la belle saison du
moins, avec une joie d’enfant, son labeur de peintre :
— Mais, ajoutait-il, je conçois l’exécution d’un tableau comme
l’aurait conçue un disciple de Memling, et vous pensez que de temps
j’y mets.
 M. Ardel ne lui cacha point qu’à sa place il se fût hâté de produire
des toiles faciles et fructueuses ; de la sorte, il vivrait indépendant et
se donnerait tout à son art :
— Non, mon cher, répliqua M. Rude tranquillement. Je suis un
pauvre passeur qui mène d’une rive à l’autre les générations ; quand
personne ne vient me quérir sur la berge, je rentre dans ma cahute
et je songe à mes pinceaux.
Pauline, en descendant, contait à Edmée son uniforme existence
entre un père tyrannisé par ses livres et un grand-oncle célibataire,
maniaque et morose, qu’ils avaient recueilli. Elle l’entretenait de leur
peine à trouver une domestique, du logis où ils étaient encore assez
mal installés :
— Votre rue me plairait, observa Edmée, parce que l’église est à
deux pas de chez vous.
Pauline, après un court intervalle, confessa :
— Nous n’avons que faire d’une église ; mon père n’est pas
croyant, ni moi non plus…
Elle regarda Edmée, aperçut dans ses yeux affables une
désillusion subite ; et pourtant elle ne regretta point de l’avoir avertie
sans réticence ; une pointe d’orgueil exaltait sa franchise ; si Edmée
la voulait pour amie, elle l’accepterait comme elle était. Mais Julien,
à deux pas derrière, émit d’une voix paisible et pénétrante :
— Si vous saviez quel don c’est de croire !
Elle tourna la tête et riposta durement :
— Ce don-là m’est aussi étranger que les chimères d’un fumeur
d’opium.
Julien se rapprocha : bien qu’une émotion vibrât dans sa gorge, il
se maintenait calme au dehors :
— Des chimères ! Pour les aveugles-nés, le soleil aussi est une
chimère, ou le serait, s’ils ne croyaient en ceux qui voient.
— C’est possible, trancha Pauline, je suis une aveugle-née.
 Un silence d’embarras aurait succédé à ces chocs imprévus,
sans une diversion qui s’offrit.
Ils attendaient au bas de la montée M. Rude et M. Ardel. Un
prêtre, venant de la campagne, passa dans le crépuscule, sur la
route. Un paysan, venant de la ville, ivre et hors d’état d’aller plus
loin, s’était assis au bord du fossé. A la vue de l’ecclésiastique, il
montra le poing, grogna des invectives. Le prêtre s’arrêta en face de
cet homme avec une attitude compatissante :
— Mon ami, lui dit-il, d’où êtes-vous ? Voulez-vous que je vous
aide à rentrer ? Le froid de la nuit va vous prendre. On vous
ramassera mort demain.
L’ivrogne tenta de se dresser, et tomba sur les genoux ; mais il
vociférait :
— Viens ici, feignant de corbeau, que je te fasse ton affaire !
Le prêtre immobile le considéra tristement, puis il s’éloigna.
Avant de disparaître sous le pont du viaduc, il se retourna une fois
encore.
Du talus, M. Ardel avait pu l’examiner ; un étonnement anxieux
l’attachait à suivre la forme noire qui s’en allait ; dès qu’il ne la vit
plus, il dit à M. Rude :
— Tout de même, il y a des rencontres inexplicables. Zoroastre,
d’après la légende, croisa, dans une allée de son jardin, sa propre
image, son double qui déambulait. Ce qui m’arrive est autre. J’ai un
frère vicaire à Lyon ; or, ce prêtre, là-bas, lui ressemble comme son
fantôme. C’est son encolure, sa taille, son profil. Matériellement, ce
ne peut être lui, et pourtant c’est lui…
— Il serait facile de s’en assurer, offrit Julien. Si vous me
permettez, je le rattrape et lui demande son nom.
M. Ardel s’y opposa : alors même que son frère, par une
fantastique coïncidence, visitant la région, se fût promené, à cette
heure, sur cette route, il ne tenait pas à le revoir ; depuis longtemps
ils étaient brouillés.
 Cependant, on se remit en marche. Au passage à niveau, le
sifflet furieux d’un rapide arrivant de Paris les arrêta. Devant eux, les
deux lampes du chariot brûlant coururent le long des parallèles
d’acier ; le train roula, trépida, comme un ouragan, dans la fumée, et,
avant qu’ils eussent traversé la voie, le fanal du dernier wagon se
perdait au fond de la nuit tombante.
Pauline en prit occasion pour confier à Edmée son désir des
grands pays lointains que, sans doute, elle ne connaîtrait jamais ;
elle se divertissait en lisant des récits exotiques, de même qu’en
chantant : Cet asile aimable, d’Orphée, elle trouvait l’illusion d’irréels
bocages élyséens.
Leur propos revint à la musique, comme à un des points solides
où leurs enthousiasmes concordaient. Il fut décidé que, le dimanche
suivant, M. Ardel et sa fille iraient en écouter chez les Rude. Pauline
se sépara d’eux, le cœur dilaté d’une joie naïve ; avide d’affection,
elle s’élançait à cette sympathie neuve. M. Rude l’attirait par une
largeur de bonté dont son propre père semblait incapable. Elle voyait
déjà en Edmée une sœur élue, et si tendre, si délicate ! Quant à
Julien, plus distant, elle ne lui gardait nulle rancune de sa légitime
réplique à une parole vexante : Nous sommes quittes, pensait-elle.
Sans être troublée de son image, elle lui reconnaissait une
mystérieuse supériorité, une âme loyale, ardente que, malgré leurs
contradictions, elle aimerait.
Dans le soir funèbre et glacial, elle rentra tout en fête ; sa vie
prochaine s’ouvrait comme un champ de roses sous une lune de
printemps.
 II

La maison des Ardel donnait sur la rue de la Synagogue, une rue

monastique, faite de longs murs et de portails fermant des jardins.
On l’appelait dans la ville la maison à la treille, parce que c’était la
seule qui eût gardé, selon la mode d’autrefois, un tortis de vigne
contre sa façade. Pauline, de sa chambre, n’avait à contempler que
le toit rouge d’une grange ; si elle se penchait, elle découvrait à sa
gauche des acacias sans feuilles et le clocher rond de Saint-Pierre.
Mais, la plupart du temps, elle se tenait en bas, dans la salle à
manger, occupée du ménage, cousant, lisant, et le soir, au salon,
lorsqu’elle ouvrait son piano pour chanter.
Cette demeure avait au moins cent cinquante ans d’âge. Ses
fenêtres en retrait dans les murailles épaisses conservaient leurs
menus croisillons, et les plaques des cheminées montraient en relief
les trois lys de France. L’amour des anciens logis n’était pas ce qui
avait décidé M. Ardel et Pauline à louer celui-là ; ils l’avaient pris,
faute d’en rencontrer un plus commode où chacun fût
« indépendant » ; car l’oncle Hippolyte, leur payant sa pension, se
croyait en droit d’exiger « ses aises ».
La maison pourtant exerçait sur Pauline un ascendant singulier.
A Roanne, ils avaient habité une rue bruyante, un appartement
moderne où on se sentait campé, jamais chez soi. Ici, au contraire,
après un mois de séjour, elle se figurait y être fixée pour la vie. Les
meubles de famille se rangeaient chacun à une place qui paraissait
leur convenir uniquement. En accrochant des estampes aux
cloisons, elle se disait que ces boiseries fanées les avaient, depuis
un siècle, attendues. Les chambres, immenses, avec leur plafond
traversé dans sa longueur par une maîtresse-poutre, détenaient la
gravité confidentielle des vieilles gens qui savent beaucoup de
secrets. Leur silence équivalait, pour elle, à un silence d’église. Si,
de fois à autre, le colloque des passants, des galoches claquant sur
le pavé, les ressauts d’une charrette, et, tous les quarts d’heure,
l’horloge de la cathédrale n’eussent couvert les battements légers de
la pendule, elle aurait pu se croire à vingt lieues d’un pays fréquenté.
Quand son père sortait ou rentrait, elle l’entendait à peine, tant les
parois étaient sourdes. Par les nuits de tempête, les plus folles
bourrasques s’amortissaient en un vague ronflement.
Tout d’abord, elle ne s’ennuya point de ce calme absolu ; ses
pensées prenaient là une couleur d’intimité si pleine de délices
qu’elle ne songeait pas à y rien changer ; tandis qu’elle ordonnait
céans toutes choses, elle s’attachait davantage à l’intérieur qu’elle
faisait sien. Elle emplit de vaisselle et de linge les placards, aligna
sur des rayons les livres du professeur, appendit des rideaux aux
fenêtres de l’oncle. Cet emménagement ressuscitait une foule
d’objets domestiques auparavant ensevelis sous la poussière
d’autres armoires. Dans celle de sa propre chambre elle mit, non
sans l’avoir épousseté, un crucifix d’ivoire, relique probable de sa
grand’mère, et dont un bras était cassé.
Les premières semaines, ces soins l’absorbèrent. Ensuite, sa
tranquillité lui devint excessive ; elle n’en souffrait pas jusqu’à l’ennui,
trop bien portante pour subir des idées mélancoliques, apercevant
toujours une tâche précise à remplir, et capable, sans être
tourmentée de ses rêves, d’en meubler son isolement. Mais elle
souhaitait une occasion de le rompre : plus tôt qu’elle ne l’espérait,
sa rencontre avec les Rude répondit à cette attente. Ce fut, toute la
soirée du dimanche, l’aliment de ses méditations.
M. Ardel, au souper, avait dit des Rude : Ils sont très bien. Mais,
sur Julien, il ajouta une réserve immédiate :
— J’ai peur que ce garçon ne soit un dangereux mystique.
 — Pourquoi dangereux ? s’inquiéta Pauline.
— Parce qu’il doit s’évertuer à endoctriner tous ceux qu’il
approche.
Elle sourit d’une façon quelque peu méprisante :
— Je lui ait fait sentir qu’avec nous il n’y a rien à faire.
— Ah ! dit-il en se tortillant la moustache, c’est donc qu’il a
essayé ?
— Non, protesta-t-elle vivement, nous avons échangé deux ou
trois mots pointus, et c’est tout.
M. Ardel voulut savoir « quelle botte » Julien lui avait poussée et
comment elle « l’avait parée ». Pauline répéta la phrase : « Si vous
saviez quel don c’est de croire », et sa violente riposte. Mais elle tut
l’allusion aux « aveugles-nés », dans la crainte vague que son père,
froissé par le dogmatisme inflexible de Julien, ne prît en méfiance
tous les Rude, au point de briser net leur amitié naissante. Puis,
cette réflexion l’humilia :
— Est-ce moi, Pauline, qui ruse ainsi ? Faut-il que cette famille
me tienne déjà au cœur ? Qui sait si je ne me trompe pas comme
une sotte sur les sentiments d’Edmée ?
Néanmoins, la figure si franche de la jeune fille, le premier regard
de Julien, le timbre de sa parole s’imposaient à sa mémoire ; elle
entendait l’« A bientôt » ! cordial de M. Rude ; se pouvait-il que leurs
avances fussent un mensonge ?
— C’est vrai, conclut-elle, je commence à les aimer. Mais eux,
que pensent-ils de moi ? Ils ont dû me juger pédante et brutale…
Tant pis ! Ce n’est pas ma faute s’il m’insinuait ses opinions
absurdes. Il m’appelle une aveugle-née, parce que je n’admets pas
avec lui que trois dieux n’en font qu’un, qu’il y a un enfer pour les
incrédules, et que les prêtres auraient le droit de me brûler vive en
punition de mes péchés ! L’aveugle, est-ce moi ou lui ? Quelle chose
étrange ! Sur d’autres questions il raisonne admirablement. Après
tout, Kepler croyait aux astrologues, et c’était quand même un grand
génie…
 En fait, Julien, par cela seul qu’elle le connaissait, avait entamé
la sécurité de son incroyance ; mais trop d’orgueil l’empêchait de se
l’avouer ; autrement, elle se fût détournée de lui avec irritation. Il
s’offrait comme un livre dont certaines pages étaient écrites en une
langue énigmatique. La douceur dominatrice qu’émettaient ses
moindres gestes, elle l’attribuait non à une vie transcendante qui
dégageait en lui l’essence divine de la beauté d’un homme, mais à
sa noblesse native et à sa culture d’esprit.
Quoi qu’il en fût, elle se coucha en pensant aux Rude, et, le
lendemain, au réveil, elle y eût pensé encore si l’impression d’un
songe pénible ne se fût interposée : pendant son sommeil, sa mère
lui était apparue.
Mme Ardel, après la naissance d’un enfant mâle qui ne vécut
pas, avait succombé à une fièvre lente. Pauline se la rappelait
exposée sur son lit avec des fleurs contre elle, tant de fleurs qu’on
en suffoquait. Seulement, elle écartait d’habitude ce souvenir
comme tout ce qui la mettait vis-à-vis de la mort. Mais, cette nuit, la
défunte était revenue : debout devant une glace où se mirait, jaune,
desséché et affreusement triste, son visage de cadavre, elle avait
l’air de se coiffer, elle se penchait, démêlait ses cheveux gris ; une
sorte de phosphorescence dansait autour d’eux depuis leur pointe
jusqu’à leur racine ; et, du creux noir de ses orbites, se détachait par
instants une larme semblable à une goutte de cire brillante. Pauline
était là, elle se voyait telle qu’à douze ans, assise sur une chaise de
paille un peu haute, les deux pieds joints, et brodant un feston. Elle
s’était levée soudain, pour courir à sa mère, les bras étendus. Celle-
ci alors avait tourné la tête à regret ; sa face se découvrit tout
entière, tordue et consumée par une inconcevable affliction. Sa fille
allait, en la touchant, s’assurer que c’était bien elle ; mais une larme
tomba sur sa main, et il lui sembla qu’une épingle rougie au feu la
transperçait.
L’illusoire souffrance de cette brûlure resta tellement poignante
qu’à demi-réveillée elle regarda si sa peau n’en portait aucune
marque. Elle se frotta les yeux et secoua sa vision : les morts
pouvaient-ils se montrer, puisqu’ils ne sont plus rien ? Mais est-on
sûr qu’ils ne soient rien ? Le petit souffle qui enflait leurs narines de
vivants se dissout-il dans l’air où ils ont expiré ? De leur conscience,
subtile vibration d’atomes, quelque chose d’impondérable
n’échappe-t-il pas au néant ?
Ainsi raisonnait Pauline, perdue dans les cavernes de son
ignorance métaphysique. Elle avait interrogé quelquefois M. Ardel
sur ce mystère, et il s’était contenté de répondre : « Nous ne savons
pas. » Cependant, elle gardait, comme lui, de ses ancêtres italiens,
deux rudiments de l’instinct religieux : le culte des Mânes et
l’appréhension de l’Inconnu.
— Au cimetière de Roanne, pensa-t-elle, ma mère est seule ;
personne n’ira plus la voir. Je vais écrire qu’on mette des bruyères
du Cap et des roses de Noël…
Mais elle ajouta intérieurement, avec plus de curiosité que
d’angoisse :
— Que se passera-t-il pour moi dans cette maison et dans cette
ville ?
Elle sauta hors du lit, prompte à se lever, les jours où le
professeur faisait sa classe le matin ; elle-même, en effet, lui
préparait son bol de chocolat. Pieds nus, elle ouvrit les volets de ses
deux fenêtres. L’aube grelottait sur le toit d’en face, gris de givre ; le
ciel, d’acier pâle, d’un rose diaphane à l’orient, présageait un lundi
splendide. L’air aigu, des ablutions froides et l’espoir du soleil
montant la remirent en gaieté. Le soleil était son idole ; lorsqu’il se
montrait, les vitres de sa chambre flambaient comme des vitraux ; il
se prélassait, jusqu’à trois heures après midi, contre la maison ; le
mur le buvait par toutes ses pierres et la vigne par tous ses
sarments :
— Que vivre est beau ! se disait Pauline, enfilant les manches
d’un peignoir douillet. Qui donc a fait la mort ?
Elle descendit en hâte, à un bruyant coup de sonnette ; la laitière
venait de poser ses berthes sur le trottoir. L’ample Mme Naudot
entra comme un tourbillon et proféra d’un gosier criard, avec son
accent de l’Ile-de-France :
 — Je vous amène le beau temps ; c’te nuit, à une heure, quand je
me suis levée, le ciel n’était qu’une étoile.
Pauline s’amusait de son babil et admirait en elle une race qu’elle
croyait disparue, la bonne femme de jadis, simple et carrée, diligente
au labeur, toujours joviale. Elle paraissait jeune, bien qu’elle eût
quatre filles et deux fils dont l’aîné « avait fini son temps ». Un
mouchoir noué autour du chignon, une « marmotte » telle qu’en ont
les paysannes de la Brie, serrait son front court, entaillé d’une ride
horizontale ; sa rude mâchoire soutenait des joues rougeaudes, si
rebondies qu’elles renfonçaient ses yeux pétillants. Elle savait
Pauline sans cuisinière et lui en offrit une de sa connaissance, « une
fille honnête et forte, travailleuse, propre, mais aussi propre qu’un
oignon » ! Pauline la remercia : elle en attendait une autre qu’on
devait tout à l’heure lui présenter.
Aussitôt que le déjeuner fut prêt, elle agita une cloche afin
d’avertir « ses deux hommes ». L’oncle Hippolyte arriva le premier,
ponctuel à la manière d’une horloge « dont le mouvement, disait-il
lui-même, restait bon ».
Ce petit vieillard chauve, droit dans sa robe de chambre, affirmait
une solidité de charpente faite pour éprouver la patience de ses
héritiers. Son crâne bossué, pointu, semblait dur comme du silex ;
ses bajoues, fraîchement rasées, s’avivaient de colorations fermes.
Si ses pupilles de myope et de bureaucrate nageaient dans le vague
sous ses lunettes, un sourire de santé bénévole montait de ses
lèvres lippues aux ailes voluptueuses de son nez. Il élevait entre ses
doigts, d’une façon gauche et comique, un habit à queue râpé, fripé,
avec des parements crasseux et une doublure en loques :
— Tiens, fit-il à sa nièce qui riait, un cadeau que je t’apporte.
J’aurais bien pu le mettre encore un an ou deux.
— Voilà les cadeaux de mon oncle, remarqua in petto Pauline.
Il rangea dans un coin une chaise de cuir qu’il jugeait mal alignée
— car l’ordre était une de ses manies les plus despotiques — et, en
silence, il s’attabla.
 M. Hippolyte Ardel avait exercé trente ans l’emploi de caissier au
Crédit Lyonnais. Les millions des autres, en coulant par ses mains,
n’avaient su qu’empirer sa pingrerie instinctive. Il choyait l’argent
pour l’argent ; et, lorsque sa vue faiblissante le contraignit de
renoncer à la cage grillagée de son bureau, ce fut le seul crève-
cœur de sa vie. Il ne s’était point marié, professant qu’il faut, avant
tout, « penser à soi ». Victorien lui avait offert son domicile dans un
sentiment de fidélité familiale et la prévision d’un héritage qui ferait la
dot de Pauline.
L’oncle ne soufflait mot de ses affaires à personne ; on le
supposait, en sa qualité d’avare, plus riche qu’il n’était. D’ailleurs,
ses penchants sordides se révélaient peu aux étrangers ; il
conservait, en sa mise, lorsqu’il sortait, une correcte bienséance.
Dans la maison, au contraire, il usait ses hardes jusqu’à la corde ;
mais, Pauline l’ayant plaisanté sur son frac ignominieux, il le
sacrifiait, non sans mélancolie. Sa nièce obtenait de lui cette
surprenante concession.
— Au moins, dit-il tout d’un coup, après s’être gratté la gorge,
garde-toi de le donner à un pauvre qui le vendrait pour cent sous. Je
n’entends pas que ma garde-robe aille finir sur le dos d’un
chenapan.
Pauline, tout en se préparant une tartine de beurre, le rassura :
— Les mendiants savent déjà qu’il est inutile de sonner ici.
Elle excluait de toute compassion « les mendiants ». Ses père et
mère et ses maîtres de morale lui avaient tant ressassé que les
pauvres sont des exploiteurs, que l’aumône est une prime à la
fainéantise ou un outrage à la dignité humaine, et qu’on ne doit plus
parler de charité, mais de justice ! Dans le pauvre, elle apercevait
une figure de la mort exécrable.
Cependant, Victorien était survenu, pressé par l’heure, et
déjeunait quatre à quatre. Contre la croisée glissa au dehors la
silhouette d’un ecclésiastique. Cette ombre ramena dans l’esprit de
Pauline le prêtre de la route ; jamais, depuis son enfance, elle n’avait
approché d’un homme en soutane ; elle éprouvait à leur égard la
méfiance oppressive qu’infligent des êtres occultes, puissants et
dangereux :
« Que de bizarreries dans une famille ! Moi, libre-penseuse, je
suis la nièce d’un prêtre ! »
Pourquoi M. Ardel s’était-il brouillé avec l’abbé Jacques ? Le
professeur observait sur son frère un perpétuel silence de
réprobation ; il le reléguait au fond d’oubliettes dont Pauline, pas une
seule fois, n’avait osé soulever la trappe ; et même après l’allusion
brève de la veille, elle s’était abstenue de le questionner.
Préoccupée des Rude, tout le soir elle négligea le singulier épisode.
Maintenant, le fantôme du prêtre et le simulacre de sa mère se
rejoignaient en son idée par des chemins obscurs. Et, soudain, elle
voulut éclaircir ce qu’elle ignorait : l’inimitié des deux frères sortait-
elle seulement de leurs discordances religieuses ? Ce ne fut pas à
Victorien qu’elle s’adressa : la bouche encore pleine, il mettait son
manteau pour partir ; mais l’oncle Hippolyte, plus lent à manger,
demeurait :
— Jacques est un vilain monsieur, répondit-il d’un ton aigre où
perçait une implacable rancune. Il a entortillé ma belle-sœur Lætitia,
si bien qu’elle a légué cent mille francs aux Missions africaines de
Lyon, et, nous autres, nous nous sommes partagé les bribes.
L’oncle, en même temps, ramassait vers le creux de sa main les
miettes de son déjeuner et les jetait au fond de son bol, attentif à ne
rien perdre. Il plia rageusement sa serviette, l’enfila dans un coulant
dédoré, et l’envoya rouler à l’autre bout de la table, comme pour
souffleter au loin le « vilain monsieur ».
Il remontait en sa chambre, quand la jeune bonne attendue se
présenta ; son père l’accompagnait, un journalier d’assez malingre
tournure, avec les jambes arquées, le teint vineux, et qui, après avoir
touché son feutre en manière de salut, le garda sur sa tête. Sa fille
reproduisait son profil de mouton, son nez en pied de marmite, mais
plus grande et plantureuse, pourvue d’épaisses mains écarlates mal
déshabituées du travail des champs ; elle avait un air de placidité
soumise, l’œil rond et béat.
 Pauline lui posa les questions d’usage, et s’enquit pour quel motif
elle avait quitté ses précédents maîtres. Le père se lissa la
moustache et entama une explication :
— Mademoiselle, commença-t-il, je vais vous dire le fait sans
prendre des mitaines ; c’est moi qui l’a retirée, rapport à des
manières qui ne me plaisaient pas, oùsqu’elle était. Ses patrons
l’envoyaient à la messe, à confesse. Pas besoin de tant d’affaires.
Ma fille n’a pas été baptisée, elle n’a point fait de communion, et
vous voyez qu’elle a bien profité quand même. Sa mère et moi, nous
lui avons donné de bons bras et de bonnes jambes. Que veut-on de
plus ? Elle est forte, elle est honnête. Pour la fréquentation, elle sait
qu’on n’aime pas ça dans le grand monde, elle se tient bien. Mais
que voulez-vous ? On a le sang vif à dix-neuf ans. Je vous la donne
pour ce qu’elle est ; si nous nous arrangeons, je vous la loue ; si elle
ne vous convient pas, je n’ai pas l’habitude d’impatienter mes clients
et de leur casser la tête…
Ces propos, il les dégoisait d’une gorge grasse, écarquillant ses
doigts qu’il secouait par saccades, et gonflé d’une satisfaction niaise,
outrecuidante. Pauline eut grande envie de leur montrer la porte.
Cependant, une aide dans le ménage lui était nécessaire, et au plus
tôt. Elle répondit simplement qu’elle n’envoyait personne à la messe,
puisqu’elle n’appartenait à aucune confession. La fille, lorsqu’elle
l’eut fait parler, sembla moins sotte que le père ; et sur-le-champ elle
la retint.
— Comment vous appelez-vous ? lui demanda-t-elle.
— Égalité Lacroix.
— Égalité ? Ce prénom-là n’est pas dans mon calendrier. Notre
dernière bonne s’appelait Marie ; je vous nommerai comme elle,
Marie.
Elle apprit, en reconduisant Lacroix, qu’il était bûcheron, natif du
Morvan, qu’il avait quitté tout jeune ce pays de misère « où les
nobles voulaient tenir les petits ».
— Moi, déclara-t-il, j’étais majeur à sept ans ; j’étais maître à
douze ans de ce que je gagnais. Je suis un fils naturel non reconnu !
 Il articula ce titre de gloire avec une grotesque vantardise, devant
sa fille impassible, et, rejetant son feutre en arrière, il continua :
— J’ai battu bien des grosses villes, j’ai fait le maraîcher, j’ai roulé
la vie de Paris. Là où je suis, j’y resterai six ans et, après, j’irai
ailleurs. J’ai été marié deux fois, je suis veuf de ma seconde femme.
Elle avait eu d’un autre un gars avant notre mariage, je l’ai reconnu
— ici, il baissa la voix — ; j’ai essayé là une boule que je ne sais
pas si elle réussira. Le gamin n’est pas fort ; s’il meurt, c’est à ma
fille que l’argent revient, l’argent des grands-parents ; ils ne sont pas
malheureux…
Pauline le poussa presque dehors ; sans quoi il n’eût jamais fini.
Cet homme lui révélait une espèce déplaisante, le nomade sans feu
ni lieu, cynique, n’ayant pris de ses ancêtres paysans que la
tortuosité des calculs, un chétif anarchiste aigri contre tout ce qui
l’humiliait. Pour elle, un seul mérite corrigeait ces tares : affranchi
des errements superstitieux, Lacroix suivait jusqu’au bout la logique
de son incroyance. Elle aurait, dans la personne de Marie-Égalité,
une servante façonnée, par un endroit capital, à son image.
M. Ardel, rentré pour midi, ratifia le choix de Pauline ; il ne la
blâma point d’avoir baptisé d’un prénom usuel et commode la
nouvelle venue ; « Égalité » choquait ses préjugés de caste, plus
forts que son irréligion.
Après le repas, vers la fin du dessert, comme il méditait dans la
vapeur d’une tasse de café et allumait sa cigarette, quelqu’un sonna.
Égalité alla ouvrir, puis revint, la mine ahurie.
— Monsieur, c’est un Monsieur le Curé qui vous demande, vous
ou Mademoiselle.
— Un curé ! Vous ne pouviez pas dire qu’il n’y a personne ! tança
le professeur en levant les bras au ciel. Ce doit être pour une quête ;
vas-y, commanda-t-il à Pauline, expédie-le un peu sec.
Elle obtempéra sans empressement, et, pendant qu’elle gagnait
par la cour le vestibule, préparait une phrase de refus. Mais une
surprise la confondit : le prêtre qui attendait était celui de la route.
Elle n’avait pas oublié son cou maigre, les lignes anguleuses de sa
figure italienne. Pâle, maladif d’aspect, il se présentait dans une
contenance douce et modeste ; digne pourtant, point embarrassé ; il
vint au-devant d’elle avec un sourire cordial, mais douloureux :
— Pauline, dit-il d’une voix qui ressemblait à celle de Victorien, je
suis votre oncle Jacques ; voulez-vous prévenir votre père ?
Le visage de Pauline se fit dur comme un marbre. Le griefs de
l’oncle Hippolyte résonnaient encore à ses oreilles, et l’arrivée de ce
prêtre dans la maison contractait tout son corps d’un malaise
insurmontable. Elle avait beau savoir qu’il était son proche, la
violence de ses préventions suffoquait l’instinct du sang. Une parole
lui brûla les lèvres : « C’est inutile ; mon père ne veut pas vous voir. »
Mais l’abbé la pressait d’un regard humble et impérieux ; il la
dominait par la force, difficile à éluder, du faible qui s’appuie sur une
Toute-Puissance invisible. Dans la salle à manger il avait perçu un
dialogue, il se disait : « Mon frère est là », et s’avançait vers le seuil.
Pauline n’osa rien répondre que ces mots, d’une froide politesse :
— Veuillez entrer, monsieur.
Elle s’effaça devant lui et, sans pénétrer à sa suite, referma la
porte ; toutefois elle resta derrière pour écouter. Le tressaut de deux
chaises reculées brusquement signifia que Victorien et l’oncle
Hippolyte, comme à l’approche d’un spectre, s’étaient levés en émoi.
Elle entendit M. Ardel qui s’exclamait :
— Toi ! Jacques ! Est-ce possible ? Que viens-tu faire par ici ?
— Victorien, expliqua la voix du prêtre, incisive et néanmoins
tremblante, j’ai dû quitter le diocèse de Lyon, je te dirai plus tard
pourquoi, et je suis, depuis septembre, curé d’une petite paroisse,
tout près de Sens, à Druzy. Hier seulement, j’ai appris que nous
étions voisins. Tu ne peux te faire une idée de ma joie. Enfin je te
retrouve ; il y avait treize ans que je fatiguais Dieu de cette prière…
— Tu n’es pas encore exaucé, coupa M. Ardel sarcastique et
brutal ; tu sais tout ce qui nous sépare.
— Quand on s’est conduit comme toi, appuya l’oncle Hippolyte,
je m’étonne qu’on ait le front de se présenter chez les gens, après
avoir tout fait pour les mettre sur la paille !
 L’abbé devait avoir prévu cet accueil ; car la véhémence de
l’attaque ne parut qu’affermir sa riposte.
— Mon oncle, commença-t-il, je suis bien aise que vous abordiez
si nettement la question. Le legs de la tante, jamais je ne m’en suis
mêlé. C’était à moi qu’elle comptait donner les cent mille francs. Elle
m’a écrit ses intentions ; j’ai répondu que je refusais, je l’ai suppliée
de penser à vous. Cela, je te l’ai dit une fois : Victorien, tu t’es buté à
ne pas me croire, sans réfléchir que si j’avais ensorcelé, comme tu
le prétendais, la pauvre tante, j’eusse travaillé d’abord à mon profit.
Or, je n’ai hérité d’elle qu’une miniature et son secrétaire Empire à
plaques de cuivre ; et, dans son secrétaire, vendredi, par une
rencontre miraculeuse, j’ai retrouvé la lettre où j’opposais mon refus.
Elle avait glissé entre deux tiroirs. Tiens, lis-la ; l’enveloppe est
encore timbrée, datée…
Tout se tut un instant ; ce silence anxieux exaspéra la curiosité de
Pauline. Les révélations qu’elle venait d’entendre la bouleversaient :
l’oncle, dont elle se faisait un monstre, elle le sentait un homme, un
homme souffrant, bon, et envers qui on était apparemment injuste.
Chez elle, la haine de l’injustice tendait à s’exagérer, pour
compenser l’indigence d’autres notions morales. Une honte brusque
la prit d’écouter à la porte, comme une petite fille indiscrète, et elle
entra résolument.
L’abbé, debout près de la table, épiait sur le visage de son frère,
tandis qu’il lisait la lettre, l’aveu d’une immédiate conviction. Victorien
persistait en sa rigueur, et tirait des bouffées de sa cigarette ou en
appuyait le bout sur le cendrier. Lorsqu’il eut fini, il remit le pli dans
l’enveloppe, et, la tendant à Jacques :
— Ce n’est pas ce qui s’appelle un document probant. Enfin…
assieds-toi.
Ce langage et le geste dont il l’alourdissait énonçaient une
condescendance tellement blessante que Pauline songea : « Si
j’étais lui, je m’en irais. » Mais, voulant réparer l’aigreur de son père,
elle rapprocha une chaise, insista :
— Asseyez-vous, mon oncle.
 L’abbé avait rougi, s’était mordu les lèvres ; sa fierté lui
commandait de partir ; malgré tout, allait-il, dès le premier choc,
consentir à une défaite ? Il était venu chercher son frère, s’humilier
devant lui en justifiant ses actes ; maintenant, il le tenait presque, il
espérait, bientôt, pouvoir l’étreindre dans ses bras, et, plus tard, lui
rouvrir ceux du Père pitoyable aux cœurs aimants. Son affection
l’emporta ; il s’assit donc et dit à Pauline :
— Vous aviez à peine quatre ans, la dernière fois que je vous ai
vue, chez l’oncle Jérôme. Je me souviens d’une poupée habillée de
rouge, dont vous pleuriez la tête toute fendue. Vous l’avez mise sur
mes genoux, je vous ai demandé : « Que veux-tu que je lui fasse, à
ta poupée ? » Et vous m’avez répondu : « Elle est bien malade,
guéris-la. »
Nul de ces détails ne surnageait dans la mémoire de Pauline ;
mais, à mesure que l’abbé parlait, il cessait d’être pour elle un
étranger.
Ce n’était pas seulement sa voix qui sonnait le son des Ardel. Il
avait la même façon que Victorien de lever et de baisser les
paupières sur des pupilles sombres, tour à tour fulgurantes et
lasses. La moue dédaigneuse de la lèvre renflée s’atténuait d’une
compassion meurtrie. La contrainte d’une discipline ascétique
épurait sa maigreur, faisait son nez plus mince et son menton plus
ovale ; une âme qui avait beaucoup souffert modelait en son visage
quelque chose de la beauté des Saints.
Pauline se laissait subjuguer par une vénération ; cependant, elle
ne s’accoutumait pas encore au costume de son oncle : la funèbre
soutane, le chapeau singulier, les mains gantées de noir hors des
manches de la douillette la repoussaient par un vague effroi, comme
si de cet extérieur émanait une autorité inquiétante, un pouvoir de
vie et de mort sur les hommes.
L’oncle Hippolyte, dès qu’il vit l’abbé s’asseoir, sortit au fond par
la cuisine en grommelant assez haut pour être entendu :
— Tout à l’heure ils s’embrasseront. Ah ! c’est du propre !
 M. Ardel avait allumé une autre cigarette ; il allait et revenait, à
pas allongés, entre la table et le grand poêle de faïence que
décorait, en haut, un buste de Stendhal :
— Je soupçonnais, fit-il, que tu gîtais dans ces parages. Hier soir,
tu as passé devant nous au bas de Saint-Martin, tu t’es arrêté près
d’un ivrogne. Mais par quelle lubie as-tu lâché Lyon pour t’échouer
au fond d’une misérable campagne ?
— Une aventure, répondit l’abbé, comme il n’en arrive qu’aux
Ardel. J’ai souffleté publiquement un jeune faquin de journaliste qui
tenait en ma présence un propos indigne. La presse a mené quelque
vacarme autour de l’incident ; l’archevêché s’est ému. Bref, j’ai
compris qu’à Lyon j’étais flambé. Tu le sais aussi bien que moi, par
expérience : dans la vie sociale il est irréparable d’avoir trahi qu’on
est violent… Ici, je connaissais un des vicaires généraux ; les prêtres
manquent, on m’a donné de suite une paroisse.
— Et tu es heureux ?
L’abbé crut inutile d’initier Victorien à toutes ses douleurs
sacerdotales. Druzy, depuis un demi-siècle, végétait dans la plus
sinistre indifférence, sauf trois ou quatre vieilles femmes, les
villageois entraient à l’église tout juste pour les mariages et les
sépultures. Ils y pénétraient, le chapeau sur la tête et la pipe à la
bouche. Son prédécesseur avait achevé de les perdre. On le trouvait
quelquefois, au moment des offices, ivre-mort en sa cave. Il laissait
dans les burettes pourrir des cadavres de mouches noyées. Les
gens l’invitaient par dérision à des enterrements civils. Le clergeon
qui lui servait sa messe n’y consentait que s’il empochait, avant
l’Introït, ses deux sous de salaire, et, quand le curé oubliait de
fermer à clef la porte, il se sauvait pendant la Consécration.
L’archevêque avait suspendu le prêtre impuissant et méprisé.
L’église était demeurée close huit mois, quand l’abbé Ardel
accepta, pour le ressusciter, ce pays de mécréants. D’abord, il avait
pleuré amèrement, mais sans perdre confiance ; à présent, ses
espoirs se confirmaient, et ce fut de l’œuvre commencée qu’il
entretint son frère :