Clinical Engineering Merge into Medical Laboratory Science

Urinalysis Automation in Clinical Biochemistry

Continuous Professional Development Program

for Medical Laboratory Professionals

Published by
College of Medical Laboratory Science, Sri Lanka
No 25/2, Norris Avenue, Colombo 08
www.cmls.org / cmlssrilanka@gmail.com
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 Academic Panel

Dr. An Le
Clinical Application Specialist

Dr. Panduka Karunanayake

Senior Lecturer, Department of Clinical Medicine
Faculty of Medicine, University of Colombo

Dr. Gaya Katulanda

Consultant Chemical Pathologist
National Hospital of Sri Lanka

Dr. Manjula Dissanayake

Consultant Chemical Pathologist
Teaching Hospital, Karapitiya

Mr. Chaminda Karunarathne

Lecturer, Department of Medical Laboratory Science
Faulty of Allied Health Sciences, Kothalawala Defense University

Ms. EK Prabhani Pushpamalie

Medical Laboratory Technologist
National Hospital of Sri Lanka

Mr. VG Dhanushka Lalinda

Medical Laboratory Technologist
Colombo North Teaching Hospital, Ragama

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 “Healthcare professionals with update knowledge is
an asset for the country

Dr. Rajitha Senarathne

Hon. Minister of Health and Indigenous Medicine

Medical Laboratory Service is one of fastest developing area in healthcare service. Updating
medical Laboratory Professionals with novel clinical engineering innovation is vital in
developing medical laboratory service in Sri Lanka. Ministry of health always encourages
professional development programs as it is important to upgrade the knowledge among
medical laboratory professionals. It will be reflected through better healthcare delivery to the
nation.

Healthcare professionals with updated knowledge are an asset for the country and it is the key
responsibility of professional organizations. I appreciate the effort of the College of Medical
Laboratory Science holding the responsibility of upgrading the standards medical laboratory
professional.

I congratulate them for the newly implemented Continuous Professional Development

Program of the College of Medical Laboratory Science, Sri Lanka and their inaugural
program with providing better opportunity to serve the country.

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 Why does Continuous Professional Development (CPD)
Program need for a Professional Service ?

Continuous Professional Development (CPD) Program is developed

with objectives of developing profession, maintain, improve and
proaden the knowledge and skills of professionals and develop the
personal qualities required to uplift overall professional standards. It
provides a platform to provide continuous development of
professional knowledge, in the form of skills, values, ethics and
attitudes, and the competence necessary to improve the knowledge
required in career development.CPD program enables professionals
to ensure that their knowledge, skills and practices are current,
complete and adequate for the roles being performed. CPD is also a
way forward to ensure personal career development through more
advanced learning activities. CPD is a lifelong process that should be
planned by considering both current and future skills required in
those respective professions.

CPD program helps to enhance knowledge and skills and be up-to-

date in your career.CPD program improves and broadens knowledge
and skills and supports future professional development.CPD
program develops personal qualities and skills necessary to execute
professional and technical duties.Correct professional values, ethics
and attitudes that are characteristic of a professional act is the
foundation for professional competence. Professional knowledge
relates to those topics that make up the subject of accountancy as well
as other business disciplines, all of these, constitute the essential body
of knowledge for professional accountants.Professional skills are the
ability required to apply professional knowledge, professional values,
ethics and attitudes appropriately and effectively within a
professional context.

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 “Ministry of Health is expected to give more
opportunities for the professionals who perform
competently through CPD”

Dr. Anil Jasinghe

Director General of Health Services
Ministry of Health and Indigenous Medicine

It is vital to conduct a Continuous Professional Development Programs in order to maintain

professional standards of a professional service. It has become a compulsory requirement
especially for the Profession of Medical Laboratory Science which is bound with daily
upgraded innovative science and technology. Even though it was neglected in past years, the
College of Medical Laboratory Science, Sri Lanka has taken step to recover that drawback
holding the responsibility of uplifting professional standards of medical laboratory
profession. It is appreciative and I wish to see a successful Continuous Professional
Development Program (CPD). My best wishes for this inaugural Continuous Professional
Development program for being first step of revolutionary journey in medical laboratory
service.

Performance and competency evaluation is important in maintaining key professional

indicators (KPI) in a professional service. Implementation of performance based promotion
system is important in improving the quality of professional services in healthcare system and
this is high time to convert it towards more efficient system. Accordingly, Ministry of Health
is expected to give more opportunities for the professionals who perform competently
through Continuous Professional Development Programs in health care service.

Therefore I would emphasize the value of active participation at CPD programs in order to
identify revolutionary stages of the profession and formulate the future profession. It will be
the motive of developing healthcare services toward betterment of the nation. It is a key
responsibility of respective healthcare professionals. Professional college is the most relevant
organization to work collectively toward this objective and all medical laboratory
professionals are requested to join these Continuous Professional Development Programs
uplifting professional competency and performances toward betterment of medical laboratory
service.

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 “It is widely known that 50 - 70% of clinical decisions are taken
on results of medical laboratory tests

Dr. Gaya Katulanda

President
College of Chemical Pathologists, Sri Lanka

Continuing professional development (CPD) is a vital need of any profession. CPD allows
members of a particular profession to develop, maintain, increase and expand both
knowledge and skills while developing values, ethics and attitudes.

It is widely known that 50 - 70% of clinical decisions are taken on results of medical
laboratory tests. These test results help in diagnosis, treatment monitoring, screening and
prognostication of diseases. Accuracy, appropriateness, timeliness and crucial factors are
contributed in decision making. Moreover, the field of medical laboratory science is
developing constantly. Emerging tests, emerging technologies, information technology, and
augmentation of prevailing tests have rocked management strategies.

Embracement of new technologies in parallel with gaining new skills is paramount to good
health care service. In addition to analytical skills and knowledge on methods, the medical
laboratory technologists shall have understanding on test interpretation and Pathophysiology
of disease processes to provide the optimum service. A professional organization, such as the
College of Medical Laboratory Science should come forward in designing and implementing
CPD programs for the members of the profession.

I congratulate the College of Medical Laboratory Science for initiation of the CPD program
for medical laboratory technologists who play unseen but crucial role in the health care
system of Sri Lanka. This is certainly a timely requisite to improve and maintain standards of
medical laboratories in government hospitals and will be an inspiration to all those who
attend to develop their carriers. -

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 “Least possible health professionals to be replaced with science
and technology are the Medical Laboratory Professionals”
Ravi Kumudesh
President
College of Medical Laboratory Science, Sri Lanka

Someone may believe that medical laboratory professionals would be replaced by science and
technology. Some are being afraid and some are waiting for that day.

According to the prediction of the people who unscientifically interpret the close relationship
between innovative science and technology in medical laboratory science as a misfortunate,
indicates medical laboratory tests may become a simple process to be done by any person.
But scientific truth is the foremost health professionals who are modified rapidly through the
revolutionary changes of science and technology or least possible health professionals to be
replaced with science and technology are the medical laboratory professionals. It is a
privilege of medical laboratory profession. The reason for being privileged is, though the
changes of science and technology will significantly affect every profession, medical
laboratory profession is able to communicate these changes more quickly as because the
medical laboratory science has originated with evolution of science and technology.

Presumption based western medicine has already been replaced by evidence based medicine.
Therefore medical laboratory report is not a simple proof for clinical interpretation anymore
and it has become the primary factor which converts the decision making process towards a
scientific process beyond clinical assumptions.

Even though someone can superficially see the technological revolution as a simple
replacement of a machine for a laboratory test, the real conversion is the development of a
simple test towards generating more comprehensive scientific evidence which can be used
more precisely in decision making.
Evolution of science and technology will simplify most of laboratory processes and we
should be ready to perform laboratory investigations simply in a lab giving strong scientific
evidences which can interpret our horoscope scientifically. Hence it becomes a compulsory
requirement of medical laboratory professionals than any other profession to update with
relevant knowledge and skill in order to be confident in laboratory revolution.

The knowledge and skill of medical laboratory professionals who are generated by science
and technology cannot be stagnated. Previous knowledge automatically becomes outdated
and we should continuously update with new knowledge in order to perform competently and
confidently in the profession. It is mandatory for being established in the profession
especially for our tomorrow.

It is strongly believed that Continuous Professional development Program of the College of

Medical Laboratory Science will fulfill the existing vacuum of professional development
process.

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 Preface
Clinical Engineering represents the part of Biomedical Engineering focused on the
applications of theories and methodologies of the broad biomedical engineering field to
improve the quality of health services. Clinical engineering combines with the medicine
knowledge for conducing of healthcare activities by providing expertise in a wide spectrum
of topics, from human physiology and biomechanics to electronics and computer science.

Health Technology includes drugs, biologics, medical devices, equipment and supplies,
medical and surgical procedures, and support systems, organizational and managerial
systems. Medical Laboratory is major section which applies health technology and clinical
engineering. Laboratory automation is combination of clinical engineering and health
technology. Laboratory automation comprises many different automated instruments,
devices, software algorithms, and methodologies used to enable, expedite and increase the
efficiency and effectiveness of the laboratory operations. It is primarily a means to improve
efficiency and reduce human errors in all steps of testing. Automation adds value to the test
result with faster turnaround time, improved accuracy, precision and safety. The electronic
verification and authentication of results enhance the information value which ensures more
accuracy and also better storage and faster retrieval of results.

All these help in delivering quality service to the patients and their treating physicians. The
automation of laboratory testing does not mean there is no need for human intervention or
expertise. Results are still need to be evaluated by qualified medical laboratory professionals.
What automation does is ease the concerns about error reduction, staffing, report timings, and
safety.

This session is focused on urinalysis automation which merge clinical engineering, health
technology with medical laboratory science towards high quality analysis on a urine as a
body excretion fluid to provide scientific evidence for clinical decision making.
Automated urine sediment analysis is sufficiently precise and improves the workflow in a
routine laboratory. In addition, it allows sediment analysis of all urine samples and thereby
helps to detect pathological samples that would have been missed in the conventional two-
step procedure.

Urinalysis is a major diagnostic screening test in the clinical laboratory, with an important
role in diagnosing and monitoring nephrological and urological conditions.

Urinalysis has been evolved with many milestone namely image transmission for expert
observation through tele medicine, strip technology with advances of electronic detection and
automated microscopy. It consists of two methods namely imaging system and flow
cytometry.

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History of Urinalysis
From 6000 years ago Laboratory medicine was began with the analysis of human urine which
was called as uroscopy and today is termed urinalysis. Today physicians use urine to
diagnose selective conditions but from ancient times until the Victorian era, urine was used as
only a primary diagnostic tool. Physicians spoke of urine as a „divine fluid‟, or a window to
the body. Babylonian and Egyptian physicians began the art of uroscopy. Uroscopy, from the
word „uroscopia,‟ means „scientific examination of urine.‟ The word is derived from the
Greek „ouron‟ meaning „urine‟ and „skopeo‟, meaning to „behold, contemplate, examine,
inspect‟.

Sumerian and Babylonian physicians of 4000 BC recorded their assessment of urine with use
of clay tablets. Ancient Sumer, one of the earliest civilizations, recognized that urine
characteristics were altered with different types of diseases. Hindu cultures were aware that
some people's urine tasted sweet, and that black ants were attracted to this sweet urine, a
characteristic of the disease now known as diabetes mellitus.
The important theory of disease causing describe in 16th century, was that of the four humors

01. Blood
02. Phlegm
03. Yellow bile
04. Black bile

And describes these are originate from the different region of the body. To keep these four
humors in balance was the responsibility of physicians.

In the fourth century BC, Hippocrates (460–355 BC) hypothesized humors which filtrate in to
the urine came from the blood which filtered through the kidneys. Hippocrates describes
presence of bubbles on the surface of fresh urine as an indication of long-term kidney disease.
Hippocrates also noted that urine sediment increased as the fever worsened. The observed
sediment may have increased white blood cells and bacteria from a urinary tract infection.
The presence of increased blood cells in the urine was due to the kidney or bladder
ulceration.

Galen has described the urine output volume difference between kidney patient and healthy
person. This is termed now polyuria, oliguria and anuria.nThen it was invented causes
courses for cloudiness. Heat and acid would precipitate proteins, causing proteinuria to
manifest through cloudiness.

Later royal physician to King Philippe-Auguste of France, described 20 different types of

medical conditions of the body and how the changes occured in sediment and color. A
physician viewed urine, assessing color, consistency, and clarity. Rounded at the bottom and
shaped like a bladder.

Later collectively manual urinalysis was developed to detect changes of urine.

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Manual Method
In manual urine estimation freshly voided urine sample is centrifuged at low speed and
resulted supernatant and the sediment use for analysis. The manual urine analysis composed
of three parts namely macroscopical analysis, chemical analysis and microscopical analysis.

Macroscopical analysis includes number of parameters as appearance, volume, colour,

specific gravity, odour and Acidity.

Chemical Analysis includes urinary protein, glucose, ketone bodies, urine bile, bile salt and
urobilnogen as chemical parameters. Protein can be detected with use of the heat and acid
precipitation methods such as Sulphosalicylic acid. Glucose is commonly detected by the use
of Benedict‟s test or copper reduction test. Ketone bodies consist of three types in human
urine (Aceto acetic acid, Beta hydroxy butyric acid and Ammonia). Rothera‟s nitroprusside
test is commonly used for detection of ketone bodies in urine. Fouchet‟s test detects urine
bile. Urobilnogen is detected by Ehrlich‟s test. Hay‟s test is commonly used for the detection
of bile salts in urine.

Microscopic Analysis includes microscopic observation for cellular components (Pus cells,
Red cells and Epithelial Cells), Crystals (Calcium Oxalate, Triple phosphates , uric acids ,
etc.), Casts (Pus cell casts/granular t casts, Red cell cast, Epithelia casts and Hyaline Casts)
and others deposits (Yeast , Bacteria ,Spermatazoa etc).

Semi-automated method
Urine strips first made on industrial scale and offered commercially in 1950s. Introducing
dipsticks to the urinalysis become much convenient than the manual methods. The dipstick to
measure several analytes and the manual microscopic examination (MME) of the urine
sediment .Generally urine strips are consist of two parameters, four parameters and most
commonly in ten parameters. The test results can be read within 60 to 120 seconds after
dipping in freshly voided urine specimen. The color reactions are approximately proportional
to the concentration of the substance which is being tested. The results are interpreted by the
comparing the pad colors with scale provided by the manufacture. Time beyond 2 minutes,
color changes have no diagnostic value.

The dipsticks can be analyzed completely by manual method (through necked eye), semi-
automated method ( Dip in the urine specimen by hand and results are interpreted with use of
strip reader) or by fully automated method (The test strips completely analyzed by an
instrument).

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Urine dipsticks principles
The Diagnostic Strips for Urinalysis are firm plastic strips to which are affixed several
separate reagent areas. These tests may provide information regarding the status of
carbohydrate metabolism, kidney and liver function, acid-base balance, and urinary tract
infection.
Glucose
This test is based on a double sequential enzyme reaction. One enzyme, glucose oxidase,
catalyzes the formation of gluconic acid and hydrogen peroxide from the oxidation of
glucose. A second enzyme, peroxidase, catalyzes the reaction of hydrogen peroxide with a
potassium iodide chromogen to oxidize the chromogen to colors ranging from green to
brown.

Bilirubin
This test is based on the coupling of bilirubin with diazotized dichloraniline in a strongly acid
medium. The color ranges through various shades of tan.

Ketone bodies
This test is based on the development of colors ranging from buff-pink, for a negative
reading, to purple when acetoacetic acid reacts with nitroprusside.

Specific Gravity
This test is based on the apparent pKa change of certain pretreated polyelectrolytes in relation
to ionic concentration. In the presence of an indicator, colors range from deep blue-green in
urine of low ionic concentration through green and yellow-green in urines of increasing ionic
concentrations.

Blood
This test is based on the peroxidase-like activity of hemoglobin, which catalyzes the reaction
of diisopropylbenzene dihydroperoxide and 3,3',5,5'-tetra methylbenzidine. The resulting
color ranges from orange through green; very high levels of blood may cause the color
development to continue to blue.

pH
The test is based on the double indicator principle that gives a broad range of colors covering
the entire urinary pH range. Colors range from orange through yellow and green to blue.

Protein
This test is based on the protein-error-of-indicators principle. At a constant pH, the
development of any green color is due to the presence of protein. Colors range from yellow
for "Negative" through yellow-green and green to green-blue for "Positive" reactions.

Urobilinogen
This test is based on a modified Ehrlich reaction, in which p-diethylaminobenzaldehyde in
conjunction with a color enhancer reacts with urobilinogen in a strongly acid medium to
produce a pink-red color.

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Nitrite
This test depends upon the conversion of nitrate (derived from the diet) to nitrite by the action
of Gram negative bacteria in the urine. At the acid pH of the reagent area, nitrite in the urine
reacts with p-arsanilic acid to form a diazonium compound in turn couples with 1,2,3,4-
tetrahydrobenzo(h)quinolin-3-ol to produce a pink color.

Leukocytes
Granulocytic leukocytes contain esterases that catalyze the hydrolysis of the derivatized
pyrrole amino acid ester to liberate 3-hydroxy-5-phenyl pyrrole. This pyrrole then reacts
with a diazonium salt to produce a purple product.
In semi-automated urine analysis after dipstick put in to the urine specimens insert in to the
machine and gathered the results. The color reactions are approximately proportional to the
concentration of the substances which is being tested. Instead of scale provided by the
manufacturer, result interpretation can be carried out using reflectance photometry and in
which light reflection from the test pads decreases in proportion to the intensity of color
produced by the concentration of the test substance. A monochromatic light source directed
toward the reagent pads. The light is reflected to a photo detector and an analog/digital
converter.

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Fully Automated Urinalysis
With the development of science and technology innovations of clinical engineering offered
fully automation in urinalysis with use of different technologies. There are three main
technologies used in fully automated urine analysis.
 Flurorence flow cytometry
 Digital imaging technology
 Flat flow cell with digital imaging

Digital imaging technology

Identification software of imaging system classifies and quantifies the cells and particles in
uncentrifuged urine using a single, laminar flow of the specimen through the lens of a
charged coupling device video camera. The hundreds of digital camera captures are evaluated
by identification software, and each particle is classified based on characteristics, such as
shape, contrast, and texture. After classification by the instrument, the operator has the ability
to reclassify or correct the obtained images in the correct categories.

Flurorence flow cytometry

Urine particle flow cytometers (UFCs) have improved count precision and accuracy
compared with visual microscopy. Flow cytometry can be used to identify and quantify cells,
casts, bacteria, and other particles in urine sediment. Addition of fluorescent stain that binds
to microbes in the specimen adds sensitivity and specificity for detection of smaller
pathological elements. As particles pass through a flow cell, they are illuminated by a laser.
The elements in the flow cell are classified according to impedance, light scatter, and
fluorescence. Results are viewed as scattergrams with the numeric counts of each
sedimentary element appearing as a distinct cluster. This method has been used for detection
and quantitation of erythrocytes, leukocytes, hyaline casts, bacteria, and epithelial cells. Other
elements such as crystals, yeast, oval fat bodies, sperm, mucus, and pathological casts are
also detected but are not readily quantified. Specimens containing these elements are flagged
for review and quantitation by manual microscopy.

Flat flow cytometry with digital imaging

Flow cell with digital imaging technology is widely used in the globe. In the flow cell
analysis the sample will be organized in single thin layer by Flat flow cytometry technology
to avoid the cell overlapping & large number of images captured in dynamic fluid. After
images are collected, particles are identified and quantitated using Artificial Intelligence,
image recognition software by comparing libraries. Urine is aspirated into the instrument
and laminar flow is used to hydro dynamically orient particles as they pass through a flow
cell. Digital images are captured and particles are classified based upon shape, frequency,
statistic and texture features.

Due to the large number of formed elements that have features of various shapes and which
are easy to destroy or to change morphology, rich experience is required to do the analysis.
For quite a long time, microscopic examination is the main method. In recent years, due to
the continuous and rapid development of computer technology, digital image technology and
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intelligent identification technology, the automation and standardization of automatic formed
elements analyzers have been continuously improved and promoted. Nowadays in most
developed countries and more and more developing countries, the analysis of urine is mostly
automated. Manual microscopy is used only for determination of ambiguous or discrepant
results.

The flat flow cytometry with digital imaging technology is widely used in Sri Lanka as well.
Following are some of the institutes which successfully uses the same technology during last
3 years.
 TH Karapitiya
 TH Anuradhapura
 TH Jaffna
 GH Polonnaruwa
 Colombo North Teaching Hospital
 BH Horana

This document is intended to help the end users to get familiar with pictures captured by
Imaging technology analyzers and their differences compared with microscopic pictures. It
can be used as a dictionary to assist you to become an expert in imaging technology
analyzers and reduce the microscopic review rate. This manual could also be practical as a
reference book in clinical examination of urine formed elements.

In this document, we have described the significances of various formed elements

and sorted 4 variants of pictures for each formed element.
 Isolated pictures from automatic Imaging technology
 Microscopic findings of native sediment (10× concentrated urine sample in
400×magnification)
 Big pictures from automatic Imaging technology
 Microscopic findings of stained sediment (10×concentratedurine sample with
400×magnification, Sternheimer staining)

The urine sediment for manual microscopy is prepared as follows: native urine sample is
centrifuged in 2000 rpm and the supernatant is removed and the sediment re-suspended to
create a tenfold concentrated sample solution.

We use standardized staining (e.g. supravital staining) by Sternwheeler for better

recognition of elements. Staining reagent consists of 2 dyes (alcian blue and red
pyronin B in 1:1 ratio). The staining reagent is added to the concentrated urine sample in
1:10 ratio.

Major samples of this manual are outpatient and inpatient samples from University Hospital
Brno and a minority from internet. In order to guarantee the consistency of the pictures,
mostly each type of formed elements come from the same sample.

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Erythrocytes
Erythrocytes are red blood cells (RBC) without nucleus, with size of about 6-7µm,
biconcave shape and of light yellow color appearance in the microscopic field. The
morphology of RBCs in urine is affected by multiple factors including osmotic
pressure, pH, time in vitro, etc.

Abnormal forms Features

Big RBCs Diameter >7µm

Small RBCs Diameter<6µm with various sizes

Acanthocytes Cytoplasm often reaches out to one or more sides so that membrane
protrudes
Crenated RBCs The cells are shrinked in hypertonic urine and form many
like budding.
protrusions on the
Shadow RBCs Loss
surfaceofandhemoglobin
have similarfrom
size. cells or cytoplasm condensing to the
membrane to
Crescent RBCs Cells
formaare
ringshaped like a urine).
(hypotonic half-moon or a sickle.

Granular RBCs Granular formation in the cytoplasm and loss of hemoglobin.

RBC fragments Broken and incomplete RBCs.

RBC morphology in fresh urine signals glomerular and non-glomerular origin of

hematuria. RBCs concentration should be paid attention as well. For example, when
no red color is visible by naked eye and centrifuged urine contains more than 3/HP
RBCs under microscope, it is called microscopic hematuria. Hematuria can be divided
into three types.

Uniform RBCs hematuria

It is mostly non-glomerular hematuria. Most of the RBCs (more than 70%) are normal,
or in single form, meaning they have concave disk-shape and cell membrane with
integrity. Shadow RBCs or acanthocytes are seen occasionally, but not more than 2 kinds
of aberrant forms are present. It is mainly seen in case of bleeding in the lower
part of the glomerular and urinary tract due to capillary rupture. RBCs are not
compressed by the glomerular basement membrane, so they have normal morphology.
RBCs from kidney tubules are affected by pH and osmotic pressure changes but only for
a short time so slight changes are made. The RBCs are also in uniformity. Based on the
cause of the hemorrhage, RBC uniformity hematuria can be classified into four types.
Transient microscopic hematuria is found in healthy people, especially in the youth
with vigorous exercise, rapid march, and cold water bath or after long standing or hard
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physical labor. For female patients, menstrual blood pollution should be ruled out
by dynamic observation. Urinary system diseases like Urinary tract inflammation,
cancer, tuberculosis, calculi, trauma, kidney transplant rejection, and congenital
deformities are indicated with hematuria. Hematuria is sometimes the only clinical
manifestation of malignant tumors of the urinary system.,Some reproductive system
diseases such as prostatitis and vesiculitis, etc are indicated by hematuriaand many
hemorrhagic diseases caused by various reasons are visualized by hematuria.

Non-uniform RBCs hematuria

It is mostly glomerular hematuria (deformation RBCs hematuria). Abnormal RBCs in
urine (more than 70%) are of2 or more types. There are changes in sizes, shape and
hemoglobin distribution and content in RBCs. RBC sizes can differ 3-,4-fold, with big
RBCs, small RBCs, acanthocytes (most pathological), crenated RBCs, shadow
RBCs, half moon-shaped RBCs, granular RBCs. They have different hemoglobin
content.

Non-uniform RBC hematuria is caused by following pathological changes in glomerular

basement membrane (cause crushed RBCs), different parts of the renal tubules with
changed pH, osmotic pressure, media pressure, metabolites (such as fatty acids,
phosphatidylcholine, bile acid, etc) which are affecting RBCs. It is often accompanied
by increase in urinary protein, granular casts, RBC casts, renal tubular epithelial
cells and so on. It is seen in acute or chronic renal tubular nephritis, lupus
nephritis, chronic pyelonephritis, nephrotic syndrome, etc.

Mixed hematuria
In this condition both uniform and non-uniform RBCs are present in urine. Either type of
RBCs is 30 to 70%.It suggests that bleeding may not originate from single location;
both glomerular and non-glomerular originated RBCs can be present. Not many diseases
can cause mixed hematuria.

Leukocytes
Leukocytes (white blood cells/WBCs) in the urine are mainly neutrophils, and possible
small amount of lymphocytes, monocytes and eosinophils. Only small number of
neutrophils is considered normal. WBCs are common in urinary tract inflammation such
as pyelonephritis, cystitis, prostatitis, vesiculitis, urethritis, kidney tuberculosis, etc. and
also in tumors. But it shall be beware of contamination of secretions from female
reproductive system inflammation. Neutrophils in urine are in spherical shape with
diameter of 6-20 μm, bigger than RBCs. Without staining, the nuclei are indistinct;
particles in the cytoplasm are visible without significant degeneration and are often
dispersed in intact shape. They have phagocytosis function and athletic ability to
phagocytose bacteria, fungus, RBCs, bilirubin crystals and so on.

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Vital neutrophils are in spherical shape with diameter of 8-12 μm. In the urine,
sometimes they can become 40μm long rod, or oval shaped. There could be changes
around the cells of filamentous, wrinkles, curved or uneven shape. The nuclei are
segmented into commonly 2 to 4 lobes.

Dead neutrophils (also known as pus cells) are the neutrophils destroyed, degenerated
and necrotized in the inflammatory process. They are irregular with various shapes and
sizes (diameter 6-20 µm). The cytoplasm is full of granules inside and the nuclei are
ambiguous and often gathered into groups with obscure boundary. Pus cells and WBCs
do not differ substantially in microscopy or imaging tests. They often increase together
and their numbers are more important.

Semi-quantitative detection with diagnostic strip is based on reaction with granulocyte

esterase which exists in neutrophils only. The content of granulocyte esterase is
associated with the freshness of the neutrophils. Degenerated or necrotic pus cells may
have reduced or even no esterase. So the test results of leukocyte esterase could be
inconsistent with the formed elements results of WBC. Large clumps of WBCs are
typically observed when there are inflammation or bacterial infections of the renal and
urinary tract. The clumping is due to increased mucus in the urine. WBC clumps suggest
renal origin of WBCs and should be reported when present.

Epithelial cells
Desquamated epithelial cells in urine are mostly from urinary system such as tubules,
renal pelvis, ureter, urinary bladder, urethra, etc. Squamous epitheliums desquamated
from vagina can mix into urine. Inside of the renal tubules is covered by the renal
tubular cuboidal epitheliums; lower ureteral, bladder, urethra and vagina surfaces are
covered with stratified squamous epitheliums. Lesions of these parts would cause
increase of epithelial cells in urine.

Squamous epithelial cells originate from the surface of external orifice of urethra and
vulva. They are the largest epithelial cells in urine. They are flat as fish scales, irregular
in shape often with curling edges. They are large, about 30-50 µm in diameter. The
nucleus is small, round or ovoid. Sometimes there are 2 or more small nuclei. Complete
keratinized cells have smaller nuclei, which may even be invisible. Small amount of
squamous epithelial cells can appear in healthy people urine, rare in males‟ urine and
slightly more in adult females‟ urine (about 5 times of that in males‟ urine). If a lot of
squamous epitheliums appear together with increased number of WBCs, there could be
urinary tract inflammation. Squamous epithelial cells in samples from the female vaginal
secretions have generally no clinical significance. In cases of gardnerella vaginalis
infection, the squamous epithelial cells are sometimes entirely covered with Cocco bacilli
forming so called “clue cells”.

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Casts
Formation condition of casts are the cylindrical aggregates of proteins, cells and their
breakdown products set in the renal tubules, collection tubes.
 Existence of albumin and Tamm-Horsfall protein (T-H protein) in crude urine: this is the
base material and primary factor to form casts. T-H protein is most likely to form casts.
 Urine concentration and acidification function of renal tubules: concentration function
can achieve increased concentration of proteins and salt which are needed for the
formation of casts. Acidification function can stimulate the proteins to further denature,
set and subside.
 Alternated use of nephrons: it is helpful for the formation and excretion of casts. The
“rest” state nephrons in the lesion part would store the urine. The store time is enough to
form casts. When these nephrons recover to work and to excrete urine, the formed casts
would be excreted together with the urine.

Urine sample may contain more than one type of cast. The casts go through different stages
of development with increasing time in the kidney tubule: cell cast → granular cast → waxy
cast. Singular casts may have mixed morphology such as granular at one end and waxy at
the other.

Casts Compositions Clinical significances

Hyaline Cast T-H protein, albumin, a few Occasionally seen in healthy
chloride people Increase: parenchymal
lesion of the kidney
RBCs Cast Cast matrix + RBCs Bleeding of renal glomerulus
or tubules
WBCs Cast Cast matrix + WBCs Kidney infection disease
Epithelial cells Cast matrix+Epithelial cells Renal tubules disease
Cast
Granular Cast Cast matrix + decomposed Parenchymal lesion of the
products of degenerated cells kidney accompanied with nephron
Waxy Cast Evolved from fine granular casts Severe
stasis lesion of renal tubules
with poor prognosis
Fatty Cast Cast matrix + fat droplets Renal tubular injury, fatty
degeneration of renal
tubular epithelial cells
Renal Failure CastDerived from granular cast and Serious kidney disease with
waxy casts poor prognosis
Bacteria Cast Cast matrix + Bacteria Suppurative infection of the kidney
Yeast Cast Cast matrix + Yeast Mycotic infection
Bilirubin Cast Cast matrix + Bilirubin crystals Severe jaundice
Mixed cells Cast Cast matrix + different cells Recurrent attacks of nephritis,
and other compositions bleeding, vascular necrosis,
renal transplant rejection

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They are colorless and transparent, and soluble in alkaline urine. Hyaline casts varies in
sizes and are in cylindrical shape, often parallel in sides and rounded at ends (one end could
be slightly tapering like tail).The texture is thin. Hyaline casts can be divided into 2 types
depending on if there are cells and granules inside: Simple hyaline casts: no granules or
cells and compound hyaline casts which contains a few granules or cells (less than 1/3
of the volume).

Hyaline casts appear in urine of healthy adults occasionally (0-a few/LP). When there is
slight or temporary function impairment of the kidney, such as strenuous exercise, longtime
fever, heart failure, anesthesia or taking diuretics, a small amount of hyaline casts would
be present. Elderly people can also see increases of hyaline casts in urine. Significant
increase of hyaline casts is found in renal parenchymal disease, such as acute or chronic
glomerular nephritis, nephrotic syndrome, acute pyelonephritis, renal congestion,
congestive heart failure and malignant hypertension. When acute glomerular
nephritis happens, hyaline casts would appear together with other pathological casts.

Crystals
Features of normal crystals
Normal crystals originate mostly from food and the body's metabolism, usually without any
clinical significance. But some crystals, such as calcium oxalate crystals, appear in the
urine of healthy people due to taking vegetable food, are one of the diagnoses of urinary
tract stones when presenting in sustained large amount.

Bacteria
Bacteria found in urine include Gram-negative bacillus (rods) and Gram-positive
cocci, most commonly Escherichia coli, staphylococcus, streptococcus, proteus, etc. It
cannot be confirmed simply according to the morphological characteristics, so the final
result and its clinical significance must be based on the bacterial culture results.
From the formation to the storage in the bladder of healthy people urine, there is no bacterial
growth in the process.

Yeasts
Yeasts are oval, looking like RBCs, with strong refraction. Germs and pseudohyphae can be
visible, commonly found in female urine, diabetes patients‟ urine, or in alkaline urine.

They may occur in urine of immune deficient or immune suppressed patients. The most
common species of yeast in urine is Candida albicans.

Mucus
Mucus is secreted by glands in urinary tract and vagina. Its quantity could be increased
with inflammatory conditions. It is common constituent in urine with no diagnostic
significance.

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