MT 103 (PMLS) LEC NOTES

Lesson 1: Laboratory Glasswares

Composition of Glasswares:
Composition of Glasswares:  Glass
 Glass o Polyethylene
o Flint Glass ▪ Inexpensive, disposable
▪ Low heat and chemical ▪ Example: petri dishes
resistance
▪ Inexpensive
▪ Use to make disposable
container

 Glass
o Polypropylene
 Glass ▪ Sterilizable, expensive
o Borosilicate Glass ▪ Example: autoclavable syringe
▪ Low alkaline earth content filters, autoclavable plastics
▪ High thermal resistance (˜600°c)
▪ Ideal for use
▪ Free from chemical
contaminants (i.e. Heavy metals)
• Does not react with most
chemicals
▪ Example: kimax
• Pyrex – heat resistant;
o Can be used for
boiling solutions
▪ Disadvantage: some releases Basic Labwares:
ions 1. Beaker
2. Flask
a. Erlenmeyer flask, Florence flask,
Volumetric flask
3. Pipet
a. Serological, Mohr, Volumetric
4. Graduated cylinder
5. Test tubes
 Glass 6. Bottles
o Plastic 7. Others:
▪ Useful because its impact and a. Stirring rod
corrosive resistance b. Coplin jar
▪ Disadvantage: bind and release c. Forceps
(leach) solution. d. Tongs
e. Test tube holder
f. Test tube rack
g. Pipette bulb/pipettor/aspirator
h. Syringes
i. Vacutainer set
MT 103 (PMLS) LEC NOTES
Lesson 1: Laboratory Glasswares
Beakers:
 Wide-mouth, straight sided container with a
Pictures under “others” in Basic Labwares: pouring spout formed from the rim.
 With marking in milliliter (ml)
 For measurement and transferring fluid

Graduated Cylinder:
 An upright, straight-sided container with a flared
base that provides stability, octagon base.
 Marking: by increments
 Capacity: 5 – 2000 ml
 Uses:
o For non-critical volume measurement
o Hold 24-hour urine specimen
 For liquid measurements:
o Light colored liquid = read at lower
meniscus
o Dark colored solution = read at upper
meniscus.

Basic Uses of Beakers and Flasks: Test Tubes:

1. Non-critical measurements (estimated)  Uses:
2. Mixing solutions o Holds liquid samples
3. Holding liquid o Container for test reaction
Kinds of Top:
Bottles:  Open
 Used as containers for reagents  Closed
 Can be plastic or glass o Stoppered (vacutainer tubes)
 Supplied in various sizes and shapes o Screw-on (screw-capped tubes)
 Clear bottles Kinds of Bottom:
o For general use  Round (general use)
 Brown bottles  2) Open (ESR)
o Use for light sensitive chemical and  3) Flat (culture)
solutions.  4) Pointed (urinalysis)

Caution: When heating, use only heat-resistant tubes.

MT 103 (PMLS) LEC NOTES
Lesson 1: Laboratory Glasswares
Different Kinds of Vacutainer Tubes:

Different Kinds of Test Tubes:

Color Codes of Vacutainer Tubes and Their Flasks

Anticoagulants and Uses:  Erlenmeyer flask
Color Anticoagulant Use o Flat bottom, sloping sides that gradually
Red No anticoagulant Routine chemical narrow its diameter so that top opening
testing requiring serum; is bottle-like.
blood bank and o Plain flat opening
serological tests. o Capacity: 10 – 4,000 ml
Amber No anticoagulant Chemically clean tube;
general use.
Pink No anticoagulant Tissue culture
Yellow Sodium Blood or body fluid
polyethole- culture
Sulfonate (SPS)
Black Oxalate
Violet or EDTA Hematology testing
 Florence flask
Lavender
o Flat bottom, round sides, long cylindrical
Light Blue Sodium citrate Coagulation testing
neck
Green Heparin Chemistry testing
o Marking: total capacity (i.e. 500 ml)
Royal Blue Sodium Chemistry trace o Capacity: 50 – 2,000 ml
heparin/EDTA elements o Use for non-critical measurements
Gray Potassium Chemistry testing
oxalate/Sodium especially glucose and
fluoride alcohol levels
Red/Gray No anticoagulant Serum testing
(marbled) (contains silica
particles to
enhance clotting)
Green/Gray Lithium heparin Plasma determination
(marbled) plus gel for serum in chemistry studies
separation
MT 103 (PMLS) LEC NOTES
Lesson 1: Laboratory Glasswares
2) Graduated or measuring pipette
 Upper end: open with frosted band near the
Flasks mouth
 Volumetric flask  Tip: tapered
o Very long neck, pear-shaped  Long with total capacity marking near the top
o Marking: total capacity, found on etched  Markings: uniform increments from top towards
of neck which indicates appropriate fill end tip
level
 Use to transfer total capacity and partial volume
o Contain exact volume at specified
 For transfer and non-critical liquid
temperature (for accurate measurement)
measurements
o Use for critical measurements
 Example: most serological pipettes and some
o Meniscus must be observed
micropipettes
▪ Curve fluid surface
▪ Lower level with fill line viewed
at eye level.

3) Pasteur pipet
 Use to add small unmeasured quantities of fluid
to receptacles.
Pipettes  No markings; some has markings
 General Use:  Made of either:
o To measure and transfer liquid or o Plastic
solutions. ▪ Bulb already attached
 Basic Types: o Glass
o Volumetric or transfer pipette ▪ Rubber bulb needs to be
o Graduated or measuring pipette attached prior to pipetting.
o Pasteur pipette

1) Volumetric or transfer pipette

 Upper end: wide opening
 Center: oval
 Tip: tapered on the other end
 Calibrated to deliver (TD) a specific volume of
liquid in a certain time period.
 For critical measurements Kinds of Pipette:
 Manual
o Volume can be adjusted
 Automatic
o Preset volumes
o Must be cleaned and calibrated regularly
o Tips are disposable and replaceable
o Must be stored vertically with the tip end
down
MT 103 (PMLS) LEC NOTES 3) To Deliver (TD)
Lesson 1: Laboratory Glasswares  Allow liquid to drain by gravity
 Pipette is held vertically and the tip is place
against the side of accepting vessel.
Types of Manual Pipette:  Example:
o Mohr pipette
1) To Deliver/To blow-out (TBO) ▪ Uniform diameter with tapered
 Filled, allowed to drain and remaining tip fluid is ends
blown-out ▪ Graduations has uniform
 Deliver exact amount; not rinsed-out interval but away from the tip
 Example: ▪ Liquid delivered between
o Ostwald-folin desired marks
▪ A special pipette used in
measuring viscous fluid such as
whole blood
o Serological pipettes
▪ Long glass tube with uniform
diameter.
▪ Markings extend to the delivery
tip
▪ Has two frosted band near the
mouth piece.
Advantages of Using Automatic Pipette:
1) Easy to operate
2) Reduce technical error

Types of Automatic Pipettes:

1) Mechanical
a. Eppendorf Micropipet
b. SMI Micropipetor
2) Electric/Battery-operated

2) To contain/rinse-out pipette (TC) 1) Mechanical Pipettes

 Refilled or rinse-out after draining by appropriate a. Eppendorf micropipet
solvent  most common type with disposable tip
 Hold exact amount of liquid that must be  piston-operated device
completely transferred by rinsing for accurate  fixed or adjustable volume
measurement. b. SMI micropipetor
 Example:  Delivers cylindrical slender glass rods
o Micro-folin  For mixing manually solutions
o Dual purpose  Heat-resistant
o Sahli-hellige
o Kirk micro 2) Electric/Battery-operated
o Lang-levy  Used with volumetric and standard graduated
o Transfer micro pipettes (serological) of various sizes.
o White-black lambda  Upper end: fits into the holder which is electric or
battery operated
 Bottom: controls the aspiration and dispensing
 Can deliver different volumes
 Advantages:
o Easy to use
o Allow use of various volumes
MT 103 (PMLS) LEC NOTES
Lesson 2: Laboratory Equipment Centrifuge
 Use to spin samples at high speeds, forcing
heavier particles to the bottom of the container.
Basic Equipment Use in Clinical Laboratory:  Used for separation of the cellular components of
1) Centrifuge the blood from the liquid.
2) Autoclave  Types of Centrifuge:
3) Heating baths and oven o Microcentrifuges
4) Laboratory balance ▪ Microhematocrit centrifuge
5) Microscope ▪ Spins capillary tubes at high
6) Refrigerator speeds so that hematocrit can be
7) Spectrophotometer measured.
8) Others (pH meters, etc…) ▪ Speed: 11,000 – 15,000 rpm

Centrifuge
 Use to spin samples at high speeds, forcing
heavier particles to the bottom of the container.
 Used for separation of the cellular components of
▪
the blood from the liquid. o Ultracentrifuge
 Types of Centrifuge: ▪ Used to separate chylomicrons
o Clinical Centrifuge from serum, fractionate
▪ Table top models for urinalysis or lipoproteins, perform drug
serum separation. binding assays, preparation of
▪ Speed: 0 – 3,000 rpm tissue for steroid hormone
(revolutions per minute) receptor assay.
▪ Capacity: 5 – 50 ml ▪ Speed: 90,000 – 100,000 rpm

▪ ▪
o Serofuge o Refrigerated Centrifuge
▪ A small centrifuge used in blood ▪ High speed & samples keep cool
banking and serology to spin while being centrifuge (i.e.
serological tubes. Ultracentrifuge)
▪ Capacity: 2-3 ml ▪ Use in research but not required
▪ Serological tube size: 13 x 75 for routine use.
mm ▪ Speed: 0 – 20,000 rpm

▪ ▪
o Microcentrifuges
▪ Microfuge General Rules of Centrifuge Operation
• spins special microtubes 1. Always operate with lids closed.
at high speed 2. Balance contents before operating (tube must be
in opposite direction).
• Speed: 12,000 – 14,000
3. Allow rotor to stop spinning before opening the lid.
rpm
4. Spin samples with lids on to avoid creating
• Capacity: 0.5 – 1.5 ml
aerosols.
(microtubes)
5. Use appropriate test tubes.

•
MT 103 (PMLS) LEC NOTES
Lesson 2: Laboratory Equipment
Laboratory Balance
 Use: for weighing chemicals and media
Autoclave
 use to sterilize items by heating them under Types of Laboratory Balance:
pressurized steam.  Double pan (DIRECT COMPARISON)
 similar to large pressure cookers removal after o a single beam with equal arms
autoclave uses tongs or heat proof gloves. o standard weights are added manually to
 Principle: “steam under pressure” the right side to counter balance the
object weight.
Uses of Autoclave:  Single Pan (SUBSTITUTION)
1) Sterilization o arms are unequal in length
 Kill contaminants and infectious agents. o object is placed on the short arm pan and
 Sterilize glassware, pipettes, media, surgical a restoring force mechanically applied to
instruments and water. other arm until indicator is balanced.
2) Decontamination
 Biologically hazardous materials (i.e. Blood, Kinds of Laboratory Balance:
bacterial cultures) before disposal.  Manual
o Weigh up as small as 10ug (0.01 g)
Typical Autoclave Conditions: ▪ Triple beam balance
 Pressure
o 15 lbs/sq. inch (psi)
 Temperature
o 121°C
 Time •
▪ Double beam balance
o 15 – 20 minutes

Important Parts of the Autoclave:

1) Autoclave chamber
 Container for sterilize items.
2) Metal jacket •
 Digital/Electronic (Electromagnetic Force)
 Surrounds the chamber and source of steam.
o Weigh as little as 0.1 ug (0.00001 g)
3) Door or cover
o Use for most critical weighing
 Securely lock & has a seal to prevent escape of
o Electromagnetic force to balance the
steam.
weight of the object.
4) Gauges
▪ Single Pan or Top Loading
 Separate for temperature and pressure.
Balance
Precautions on Autoclave Operation:
1. Never open the door unless chamber pressure is
zero psi.
2. Items must be removed using tongs or heat-proof •
gloves to prevent burns. ▪ Cabinet Balance
3. For liquid sterilization, containers must be loosely
capped, heat-resistant and not more than half full.
4. At the end of liquid run, slowly reduce pressure to
•
prevent over boiling.
MT 103 (PMLS) LEC NOTES
Lesson 2: Laboratory Equipment

Important Reminders:
Safety Precautions in Using Laboratory Balance 1. Use only for laboratory purposes but not for
1. Keep balances clean; wipe up any spills promptly. storing /heating foods.
2. Avoid jarring instrument. 2. Should be monitored regularly to ensure proper
3. Use balance gently and appropriately (i.e. do not operating temperature using calibrated
weigh 5 ug on a balance only accurate for 1 ug). thermometers.
4. Position on draft and vibration free counter. 3. Temperatures must be checked before each use
5. Calibrate regularly and observe annual and recorded daily.
maintenance check.
6. Wear gloves and mask when weighing irritant and
strong chemicals.

Temperature Controlled Chambers

1. Above ambient or room temperature
a. Oven
b. Water Bath (Heating Bath)
c. Incubator
2. Cold temperature
a. Refrigerator
b. Freezer

Heating Baths and Oven

1. Oven
a. Used to dry chemical, extracts, media
used in electrophoresis, and glassware
b. Needs annual accuracy check of
thermometers.
2. Water Bath
a. Needed for incubation
b. Needs distilled or de-ionized water
i. Manual
1. Ex. Flowing water bath,
floating-out bath
ii. Digital
1. Ex. Electronic water
bath
2. Electronic water bath
3. Incubator
a. Use for incubating bacterial cultures and
other microbiologic procedure that needs
constant warming at 37°C.

Cold Temperature
1. Refrigerator
a. use to store sterile media, reagents,
preserve stock cultures, temperature: 4 –
8 °C
2. Freezer
a. use to store dry reagents and antibiotic
discs, lyophilized cultures, temperature: -
10 to - 80°C
MT 103 (PMLS) LEC NOTES Stage
Lesson 3: Microscope  Stage is part where the slide rests.
 Mirror (or light source) directs light upwards onto
the slide.
Microscope History
 In 1665, English physicist, Robert Hooke looked Diaphragm
at a silver of cork through a microscope lens and  Diaphragm allows light in.
noticed some “pores” or “cells” in it.
Nosepiece
Microscope Care  Nosepiece is the rotating device that holds the
1. Always carry with 2 hands. objectives (lenses).
2. Only use lens paper for cleaning.
3. Do not force knobs. Arm
4. Always store covered.
 Arm is the part where you carry the microscope.
5. Keep objects clear of desk and cords.

Compound Microscope
Types of Microscopes
 A microscope is a very powerful magnifying
glass. Light Microscope
 A microscope helps you see things like cells up  For small objects - similar to the way binoculars
close. magnify objects far away.
 The Compound Light Microscope
Parts of a Microscope:

 May have many lenses that magnify the

object
 Eyepiece = 10x magnification
 Objective = 10x (low) or 40X (high)
Eyepiece  Total magnification = eyepiece X objective
 View the specimen through the eyepiece. o Ex: 10x X 40x = 400x total
magnification
Stage Clips & Objectives
 Stage clips Stereomicroscope
o It hold the slide in place.  To look at large things that light cannot pass
 Low power objective through (one eyepiece for each eye)
o It is used to focus the microscope (short
& fat).
 High power objective
o It is used to view details of a specimen

Coarse Adjustment, Fine Adjustment, & Base

 Coarse adjustment
o It focuses adjustment  3 - D image can only magnify 10x to 600x
 Fine adjustment
Electron Microscope
o Fine tunes & gives detailed focus(usually
smaller than coarse adjustment knob)  Can magnify more than 500,00x
 Base
o It is where the microscope rests
MT 103 (PMLS) LEC NOTES
Lesson 3: Microscope

Using the Microscope

1. Place the Slide on the Microscope
2. Use Stage Clips
3. Click Nosepiece to the lowest (shortest) setting
4. Look into the Eyepiece
5. Use the Coarse Focus
6. Follow steps to focus using low power
7. Click the nosepiece to the longest objective
8. Do NOT use the Coarse Focusing Knob
9. Use the Fine Focus Knob to bring the slide into
focus
MT 103 (PMLS) LEC NOTES
Lesson 4: Occupational Hazards

Hierarchy of Controls
Key Principles of Health and Safety
 Employer is responsible for maintaining a safe
and healthy workplace.
 Employees should be involved in developing
policies and programs.
 There should be no sanctions for Health & Safety
related activities.
 Employer should implement best and most
effective practices/policies to protect workers
from hazards.

Controls: Engineering
Overview of Hazards  Control at the source!
o Limits the hazard but does not entirely
Chemical & Dust Hazards remove it
 Cleaning products, pesticides, asbestos, etc.  Proper equipment, re-designed tools, and local
exhaust
Biological Hazards  Other examples:
 Mold, insects/pests, communicable diseases, etc. o Mechanical guards
o Wet methods for dust
Ergonomic Hazards o Enclosures/isolation
 Repetition, lifting, awkward postures, etc. o Dilution ventilation

Work Organization Hazards Controls: Administrative

 Things that cause STRESS!  Aimed at reducing employee exposure to hazards
but not removing them!
Safety Hazards  Changes in work procedures such as:
 Slips, trips and falls, faulty equipment, etc. o Safety policies/rules
 Schedule changes, such as:
Physical Hazards o Lengthened or Additional Rest Breaks
 Noise, temperature extremes, radiation, etc. o Job Rotation
o Adjusting the Work Pace
 Training with the goal of reducing the duration,
Bureau of Labor Statistics Data, 2009 frequency and severity of exposure to hazards
 Injury and illness rate for public workers
significantly higher than among private industry Controls: PPE (Personal Protective Equipment)
workers  Control of LAST RESORT!
o 5.8 vs. 3.6 cases per 100 workers  Special Clothing
 Custodians fall into the top 5 in these categories:  Eye Protection
o Highest number of days away from work  Hearing Protection
o Musculoskeletal injury incidents  Respiratory Protection
 CONTROL IS AT THE WORKER!
MT 103 (PMLS) LEC NOTES
Lesson 4: Occupational Hazards
Major Elements of an Effective Safety and Health
Program
1. Management Commitment and Employee Involvement
2. Worksite Analysis
3. Hazard Prevention and Control
4. Safety and Health Training
Management Commitment and Employee
Involvement Go Together!
 Top management involvement should be visible
and have authority and resources to implement
program
 Employee involvement in the program and in
decisions that affect their safety and health
should be encouraged
 A clearly stated worksite policy should be
established and communicated with specific
goals and objectives.
 All aspects of the program should have assigned
responsibility and accountability.
 Program operations need to be reviewed at least
annually, to evaluate and make revisions as
needed.
Worksite Analysis
 Identify all existing hazards and conditions that
might create new hazards
 An efficient program includes actively analyzing
the work and the worksite to anticipate and
prevent harmful occurrences
Hazard Prevention and Control
 Triggered by a determination that a hazard or
potential hazard exists.
o Where feasible, prevent hazards by
effective design of job or job site.
o Where elimination is not feasible, control
hazards to prevent unsafe and
unhealthful exposure.
 Elimination or control must be accomplished in a
timely manner.
Safety and Health Training
 Address the safety and health responsibilities of
all personnel.
 Ensure that all employees understand the
hazards to which they may be exposed and how
to prevent harm to themselves and others.
 Ensure that managers understand their safety
and health responsibilities.
Occupational Safety and Health Administration
(OSHA)
What standards or regulations exist for the hazard you identified?
Record Keeping
 Requires most employers with more than 10
workers to keep a log of injuries and illnesses.
o Workers have the right to review the
current log, as well as the logs stored for
the past 5 years.
o Workers also have the right to view the
annually posted summary of the injuries
and illnesses (OSHA 300A).
OSHA 300 Log
 Used to document and classify work-
related injuries and illnesses and severity
of each case.
 Annual summary shows totals of injuries
and illnesses for the year in each
category.
 Summary must be posted in a visible
location February 1 through April 30 each
year.
Access to Exposure and Medical Records: 1910.1020
Exposure Records Medical Records
Environmental and Questionnaires,
biological monitoring Results of examinations,
 Personal Laboratory tests,
 Workplace Medical opinions,
Material Safety Data diagnoses, etc.
Sheets First aid records,
Description of treatments
MT 103 (PMLS) LEC NOTES
Lesson 4: Occupational Hazards
Hazard Communication Standard 1910.1200
 To ensure that employers and employees know
about work hazards and how to protect
themselves so that the incidence of illnesses and
injuries due to hazardous chemicals is reduced.
Occupational Exposure to Hazardous Chemicals in
The Laboratory 1910.1450
 Requires a Chemical Hygiene Plan
o Capable of protecting employees from
health hazards associated with
hazardous chemicals in that laboratory
 Capable of keeping exposures below the limits
 Requires:
o Standard Operating Procedure
o Employee information and training
o Designation of Chemical
o Hygiene Officer
 Plan shall be readily available to employees and
employee representatives
Blood-Borne Pathogen Standard 1910.1030
 Purpose:
o To Prevent Needle sticks and Other
Exposures at Work to Blood and Body
Fluids that Contain Blood
 Employer Responsibilities:
o Identify Workers at Risk
o Provide Safe Needles
o Ensure Universal Precautions are
Practiced
o Provide Personal Protective Equipment
o Provide Prompt Evaluation and
Treatment
o Provide Hepatitis B Vaccinations
o Recordkeeping
o Train Workers Annually
Asbestos Hazard Emergency Response Act – Ahera
 Requires Employers to:
o Establish Asbestos Management Plan for
every school site
o Conduct regular inspections and assess
conditions
o Provide training for
custodial/maintenance staff in general
awareness and more extensive training
for workers doing small jobs and/or
emergency cleanups
o Covers state and local workers who
perform asbestos work who aren’t
covered by OSHA
Finding Information
1. What reports, logs and documents should be
collected and reviewed by the committee?
2. How often should they be reviewed?
3. How do you think you should ask for these
programs and records?
Committee Resources
1. What training do you feel you need to be a better
committee member?
2. Legal Assistance, contacts?
3. Union resources?
4. Other allies/expertise? Who?
5. TIME, TIME, TIME……..
The National Institute for Occupational Safety and
Health (NIOSH)
 Valuable resource for information on all types of
hazard exposures
 Can conduct Health Hazard Evaluations (HHE) if
requested by union or members
MT 103 (PMLS) LEC NOTES
Lesson 4: Blood Collection (Phlebotomy)  Specimen Collection and Handling
o Physician orders indicate type of
specimen and time of collection
What is Phlebotomy? o Most commonly used methods:
 The term phlebotomy refers to the ancient ▪ Venipuncture
practice of bloodletting. • Insertion of a needle into
 Now the term phlebotomy is used for the a vein to remove blood.
withdrawal of blood from a vein, artery, or the ▪ Dermal puncture or skin
capillary bed for laboratory analysis or blood puncture
transfusion. • Use of puncture device
to obtain capillary blood
History of Phlebotomy by pricking the skin.
 Removing blood from veins dates back to about  Professionalism
1400 B.C when leeches were applied to skin of o Be professional
sick people. o Apply good interpersonal skills
 In the early 1800’s, medicinal leeches were used o Dress professionally
for the procedure known as bloodletting. o Many institutes require that phlebotomist
o Bloodletting was typically performed by wear a lab jacket and specified shoes to
barbers or anyone claiming medical meet OSHA guidelines
training.  Confidentiality
o All employees are responsible for
Bloodletting maintaining confidentiality of medical
Then Now information.
Performed by a cut into a Profession emerged as a  Attitude
vein with a sharp result of technology and o Tone of voice and facial expression will
instrument to drain blood. expansions in laboratory determine how patients respond to you.
function. o Be polite, friendly, calm, and considerate
The lancet was the most Performed by trained always.
popularly used professionals.  Appearance
instruments. o Your personal appearance will also affect
Aseptic practices were not Standards of practice set the impression you make.
known, so the lancet was by the Clinical and o Comply with your facility’s dress code
reused for several Laboratory Standards and personal appearance policies.
patients. Institute (CLSI)  Verbal and nonverbal communication should be
appropriate, such as:
Who is a Phlebotomist? o Avoiding the use of slang
 Collects blood and other specimens o Speaking in a calm, clear voice
 Prepares specimens for testing o Avoiding inappropriate terms
 Interacts with patients and health care o Maintaining eye contact
professionals o Neat, well-groomed appearance
 Plays a vital role in any health care system o Respecting personal space
 Other medical professionals, including doctors, o The phlebotomist must be able to
nurses, technologists, and medical assistants communicate using nonmedical terms
must also be trained to collect blood specimens.  When providing customer service:
o Be empathetic
Roles and Responsibilities of the Phlebotomist o Observe the patient’s behavior
 Patient identification o Listen to the patient’s concern
o Check armband or ID label in acute care o Address any situation promptly
settings o Be flexible
o Check driver’s license or picture ID in
outpatient settings
o 3 steps: Ask, Compare, Validate.
MT 103 (PMLS) LEC NOTES
Roles and Responsibilities of the Phlebotomist Lesson 4: Blood Collection (Phlebotomy)

Where Do Phlebotomists Work (Inpatient)?
 Inpatient Facilities
o Hospitals
o Nursing homes
o Rehabilitation centers
 Phlebotomists employed at impatient facilities
work directly with several members of the health
care team.
Where Do Phlebotomists Work (Outpatient)?
 Outpatient Facilities
o Physician’s offices
o Home health care agencies
o Ambulatory care centers
▪ Ambulatory care centers are the
fastest-growing outpatient
facilities.
o Reference laboratories
o Blood banks
Safety and Infection Control
Contact transmission of infectious agents can be caused
by either direct or indirect contact.
 Direct contact
o Requires transfer of pathogens from
reservoir to a susceptible host (person to
person).
 Indirect contact
o Contaminated item is handled prior to
contact with a susceptible host (person to
contaminated item to person).
Prevention of nosocomial infections
 Breaking the chain of infection
o The six links
o Portal of entry → susceptible host → with needlestick prevention features.
infectious agent → reservoir → portal of
exit → mode of transmission → portal of
entry
▪ Breaking the chain of infection of
any of these links will prevent an
infection from developing.
Safety and Infection Control
Centers for Disease Control (CDC) levels of protection
for the prevention of nosocomial infections
 Standard precautions
o Combines the good hand hygiene and
the use of gloves when workers are
exposed to contaminated products
 Isolation precautions
o Based on how the infectious agent is
transmitted:
▪ Airborne
▪ Droplet
▪ Contact
 Always follow standard precautions when
performing phlebotomy.
Occupational Safety and Health Administration
(OSHA)
 OSHA is the federal organization responsible for
preventing or minimizing employee exposure to
bloodborne pathogens such as hepatitis and HIV.
 OSHA mandates that health care facilities
provide annual training on preventing exposure to
bloodborne pathogens.
 OSHA requires that health care facilities provide
the necessary Personal Protective Equipment to
prevent exposure.
Needlestick Safety and Prevention Act of 2001
 Established through the recommendation of the
National Institute for Occupational Safety and
Health (NIOSH) and the Occupational Safety and
Health Administration (OSHA)
 Mandated the use of safety devices on needles
for the prevention of exposure to bloodborne
pathogens
 All devices for phlebotomy should be equipped
MT 103 (PMLS) LEC NOTES
Lesson 4: Blood Collection (Phlebotomy)

HIPAA, Ethics, and Law
Health Insurance Portability and Accountability Act
(HIPAA)
 Developed in response to medical information
transactions
 In 2003, established a national standard for
electronic health care transactions
 Protects the privacy and confidentiality of patient
information
 Patient information can be shared only for
treatment purposes
HIPAA – Protect Patient Information!
 Close patients’ room doors when discussing their
health
 Do not talk about patients in public places
 Log off computers when finished
 Turn computer screens so passerby cannot see
patient information
 Do not walk away from patient medical records;
close them when leaving
Venipuncture
 Blood is collected through a needle inserted in the
vein
Capillary puncture
 Also called dermal puncture and skin puncture
 Blood is collected from a skin puncture made with
a lancet or a similar device
Laboratory Workflow Cycle
Lab tests ordered → Order received in lab → Work lists
and labels generated → Phlebotomist dispatched.
Phlebotomist identifies patient → phlebotomist draws,
labels specimen → Specimen transported to lab →
Specimen accessioned and processed.
Lab performs tests → Lab results are reported to doctor
→ Doctor treats patient → Treatment based on lab
results.

Code of Ethics Blood-Borne Pathogens

 Ethics consist of a set of written rules,
procedures, or guidelines that aid in determining Definition
right from wrong  Infectious micro-organisms which live in the
 Golden rule: treat others the way you would want bloodstream.
to be treated  You can be exposed to bloodborne pathogens if
 All information concerning a patient’s care must you are injured with a contaminated needle.
remain confidential  You can also be exposed if your mucous
membranes, including eyes, mouth, or the inside
Patient’s Bill of Rights of your nose comes into contact with
 A phlebotomist must have a clear consent from contaminated body fluids.
the patient before performing any blood collection
procedure
 Patients have the right to:
o Refuse care
o Be treated with respect
o Have all records and information kept
confidential
o Be informed about the purpose and
expected results of treatments
o Have access to their medical records

MT 103 (PMLS) LEC NOTES

2 Main Phlebotomy Procedures
Lesson 4: Blood Collection (Phlebotomy)

Standard Precautions
Treat all body fluids as if they were infectious.
Potentially infectious body fluids include:
 Blood
 Semen
 Vaginal secretion
 Peritoneal fluid
 Pericardial fluid
 Pleural fluid
 Saliva
 NOTE: sweat and tears are not generally
considered infectious.
Personal Protective Equipment
 Laboratory coat/gown
 Gloves
 Face masks (certain types of isolation)
Hand Washing
 It is the single most important infection control
measure
 Wash hands thoroughly before, after, and
between all patient contacts.
 Be sure to turn off faucets using a paper towel to
avoid contamination
 Procedure:
o Remove rings
o Stand by the sink but do not touch it
o Apply soap and rub hands together
o Both sides of the hand, between fingers,
around knuckles, under fingernails
o Rinse hands in a downward motion
o Dry hands with a clean paper towel
o Turn off water with another paper towel
Hazardous Waste Disposal
 All needles and other sharps must be disposed of
in approved sharps disposal containers
 Other contaminated waste must be discarded in
an appropriate biohazard bag or waste receptacle
Needlestick
Needlesticks and Prevention Act
 Safety devices should always be encouraged
The Vascular System
Veins
 Have thinner walls because blood in them is
under less pressure
 Collapse more easily
 Dark bluish red or oxygen-poor bloods
Arteries
 Have thick walls to withstand the pressure of
ventricular contraction, which creates a pulse
 Normal systemic arterial blood is bright red,
hence it is oxygen-rich bloods.
Capillary
 Only one cell
 Can easily be punctured to provide blood
specimen
2 Basic Patterns of the Veins
 H and M Pattern
Other Veins
 Veins on the back of the hand or at the ankle may
be used, although these are less desirable and
should be avoided in diabetics and other
individuals with poor circulation
 Leg, ankle, and foot veins are sometimes used
but not without permission of the patient’s
physician due to potential medical complications
MT 103 (PMLS) LEC NOTES
Lesson 4: Blood Collection (Phlebotomy)
 Anatomy and Physiology Anatomy and Physiology

What is Blood? Types of Blood Specimens

 Thick red fluid flowing through circulatory system  Serum
consisting of liquid components and cellular  Plasma
components.  Whole Blood
o Plasma
▪ Liquid part of the blood Whole Blood
o Serum
▪ Liquid part of the blood after
clotting
▪ Minus fibrin
o Cells Plasma
▪ RBC, WBC, platelets
o Volume
▪ Approximately 10 pints in
average adult
Blood’s Function Serum
 Artery
o Carries O2 and nutrients to tissue
 Vein
o Carries COS and waste from tissue
 Capillary
Blood Clot
o Tiny blood vessels connecting arterioles
and venules  When a blood sample is left standing without
anticoagulant, it forms a coagulum or blood clot
 RBC
o Composed of hemoglobin, O2/CO2  The clot contains coagulation proteins, platelets,
carries and entrapped red and white blood cells.
 WBC
o Fights infection
 Platelets
o Hemostasis and/or coagulation

Source and Composition of Blood Specimens

 Arterial blood
o Primarily reserved for blood gas
evaluation and certain emergency
situations
Serum
 Venous blood
 It contains all the same substances as plasma,
o Affected by metabolic activity of the
except for the coagulation proteins, which are left
tissue it drains and varies by collection
behind in the blood clot.
site chloride, glucose, pH, CO2, lactic
acid, and ammonia levels differ may from
arterial blood
 Capillary blood
o Contains arterial and venous blood plus
tissue fluid
o Capillary glucose is normally higher
o Calcium, potassium, and total protein are
normally lower
MT 103 (PMLS) LEC NOTES
Lesson 4: Blood Collection (Phlebotomy)

Venipuncture
Venipuncture can be performed by 3 basic methods:
Evacuated Tube System (ETS)
 Most preferred because blood is collected directly
from the vein in tube, minimizing the risk of
specimen contamination and exposure to blood.
Needle and Syringe
 Used on small, fragile, and damaged veins.
Winged Infusion Set (Butterfly)
 Can be used with the ETS and syringe
 Used to draw blood from infants and children,
hand veins and other difficult to draw situation.
Sites to be avoided when performing venipuncture:
1. Intravenous lines in both arms
2. Burned or scarred areas
3. Areas with hematoma
4. Thrombosed veins
5. Edematous arms
6. Mastectomy on one or both arms
7. Arms with arteriovenous shunt or fistula
Complications of Venipuncture
 Immediate Local Complication
o Hemoconcentration
▪ It is an increase in number of
formed elements in blood
resulting either from a decrease
or increase in plasma volume
o Failure of blood to enter the
syringe/vacutainer tube
▪ Excessive pull of the plunger
▪ Piercing the other pole of the
vein
▪ Transfixation of vein
▪ Incorrect bevel position (bevel
down)
▪ Absence of vacuum
Complications of Venipuncture
 Immediate Local Complication
o Syncope (fainting)
▪ It is the transient loss of
consciousness due to lack of
oxygen in the brain and results
inability to stay in an upright
position.
▪ If a seated patient feels pain, the
needle should be removed
immediately, the patient’s head
should be lowered between the
legs and the patient should be
instructed to breathe deeply.
 Late Local Complication
o Thrombosis
▪ It is an abnormal vascular
condition which thrombus
develops within a blood vessel of
the body.
o Thrombophlebitis
▪ It is an inflammation of a vein
often accompanied by a clot
which occurs as a result of
trauma to the vessel wall.
 Late General Complications
o Serum hepatitis
o AIDS
Equipment
Trays
 It should be sanitized daily using appropriate
disinfectant
 Kept organized and well-stocked
Blood Collection Tubes
 Glass or plastic tube with a rubber stopper
 It has a vacuum so that blood will flow into the
tube
 Anticoagulants and/or other chemical additives
 Rubber stoppers of blood collection tubes are
color coded
 Each type of stopper indicated a different additive
or a different tube type.
MT 103 (PMLS) LEC NOTES
Lesson 4: Blood Collection (Phlebotomy)
 Blood Culture Bottles
Equipment  Different blood culture bottles are used for
aerobic, anaerobic, and pediatric collections.
STOP, LIGHT RED, STAY PUT, GREEN LIGHT, GO
- S – sterile Blood Collection Tubes: Safety
- L – light blue  The rubber stopper is positioned inside the plastic
- R – red shield which protects against splatter
- S – serum separator tube
- P – plasma separator tube Sizes
- G – green  Adult: 3 - 10 ml
- L – lavender  Pediatric: 2 - 4 ml.
- G – gray  Tubes for fingersticks or heelsticks ½ or less

Lavender
 EDTA to prevent clotting
 Hematology studies
 Should be completely filled
 Must be inverted after filling

Light blue
 Sodium citrate
 Coagulation (clotting) studies
 Must be completely filled Expiration Dates
 Must be inverted immediately after filling

Green
 Sodium or lithium heparin
 For tests requiring whole blood or plasma such as
ammonia

Red
 No additives
 Blood bank tests, toxicology, serology
 Must not be inverted after filing Holders
 A plastic holder must be used with the evacuated
Gray tube system.
 Inhibitor for glycolysis + anticoagulant
 Sodium fluoride +potassium oxalate
 Glucose levels

Yellow
 Acid citrate dextrose
 Inactivates complements Needle Holders with Built-In Protection Devices
 DNA studies and paternity testing

Royal Blue
 Heparin or na EDTA anticoagulants
 Tube is designed to contain no contaminating
metals
 Trace element and toxicology studies
MT 103 (PMLS) LEC NOTES
Equipment Lesson 4: Blood Collection (Phlebotomy)
 Needles with Built-In Safety Devices
Equipment  An internal blunt needle that is activated with
forward pressure on the final blood tube prior to
Syringes with Built-In Safety Devices withdrawal of the needle from the vein.

Butterfly Needle
 Winged Infusion Set
 Difficult Venipuncture Including Pediatric Draws.
 With A Syringe Or A Holder And Vacuum
Collection Tube System.
 21, 23, Or 25 Gauge

Syringe System
 Syringes are customarily used for patients with
veins from which it is difficult to collect blood.
 Blood gas analysis

Needles Butterflies with Built-In Safety Features

 Needles vary in length and diameter  Number-one cause of needle stick injuries, so
 Diameter is measured in gauge proper use of their safety devices is critical.
 The smaller the gauge, the larger the needle
Lancets
 The manufactures use colors to differentiate the
gauges  Lancets are used for difficult venipuncture,
including pediatric draws.
Needle Components

Tourniquets
 Vein easier to SEE, FEEL, and PUNCTURE.
 Applied to a patient’s arm during venipuncture.
 Distends the veins, making them larger and
Single Draw Needle easier to find, stretches the wall so they are
 Single draw needles are of the type that fit on a thinner and easier to find.
syringe, and can be used only to fill the syringe to  Must not be left on longer than 1 minute because
which they are connected. specimen quality may be affected.

Multiple Draw Needle

 Used with vacuum collection tubes.
 They have a retractable sheath over the portion
of the needle that penetrates the blood tube.

MT 103 (PMLS) LEC NOTES

Equipment Lesson 4: Blood Collection (Phlebotomy)
 Patient Identification
 Make sure the name, medical record number,
Equipment and date of birth on your order/requisition match
those on the patient’s armband.
Sterilization  Verify the patient’s identity by politely asking them
to state their full name.
 Properly identifying patients and specimens is
probably the single most critical part of your job.
 The consequences of misidentifying a specimen
can be life threatening.
 Never rely on the patient name on the door or
Bandaging Material above the bed.
o Patients are frequently moved from room
to room.
 A hospitalized patient must always be correctly
identified by an ID band that is attached to the
patient.
 It is the most critical step in phlebotomy
 Inpatients – must have correct wristband prior to
Gloves collection
 Gloves must be worn for all procedures requiring  Ask patient to state their full name and birth date
vascular access.  Match wristband information with test req.
 Non-powdered latex gloves are most commonly
used. Patients are often reassured that proper safety measures
are being followed when gloves are put on in their
Sharp Disposal Container presence.

Position the Patient

 Comfortable position
 Turn the arm so that the wrist and palm face
upward, and the antecubital area is accessible.
 When supporting the patient’s arm, do not
hyperextend the elbow. This may make vein
palpation difficult.
Collecting Blood
Applying the Tourniquet
 Tie the tourniquet just above the elbow.
Greeting
 The tourniquet should be tight enough to stop
 Always greet patient in a professional, friendly
venous blood flow in the superficial arm veins.
manner.
 The tourniquet should be 3 – 4 inches above the
 A good initial impression will earn the patients
punctured site.
trust, and make it easier and more pleasant to
 The tourniquet should be applied a maximum of
draw a good specimen.
2 minutes.
 Knock on the patient’s door before entering.
 After applying the tourniquet, you may ask the
 Identify yourself by name and department.
patient to make a fist to further distend the arm
 Explain the reason for your presence.
veins.
 The more relaxed and trusting your patient, the
 Patients often think they are helping by pumping
greater chance of a successful atraumatic
their fists.
venipuncture.
 This is an acceptable practice when donating
 Good verbal, listening, and nonverbal skills are
blood, but not in sample collection as this can
very important for patient reassurance.
lead to hemoconcentration.
MT 103 (PMLS) LEC NOTES
Collecting Blood
Lesson 4: Blood Collection (Phlebotomy)
 Picture by picture on how to do the procedure:

Collecting Blood 1. Attach needle to holder.

Choose a Site
 The median cubital vein is preferred.
 If not accessible: Cephalic vein, or the Basilic
vein.
 If not accessible: veins on the back of the hand.
o Use a much smaller needle for these
hand veins.
2. Place tube into holder.

 Using the non-dominant hand routinely for

palpation may be helpful when additional 3. Hold vein in place.
palpation is required immediately before
performing the puncture.
 Often, a patient has veins that are more
prominent in the dominant arm.

Sites to be avoided (Venipuncture):

1. Intravenous lines in both arms
2. Burned or scarred areas
3. Areas with hematoma 4. Insert needle.
4. Thrombosed veins
5. Edematous arms
6. Mastectomy on one or both arms
7. Arms with arteriovenous (AV) shunt or fistula

Cleansing the Site

 Isopropyl alcohol swab
 Outward expanding spiral starting with the actual 5. Push tube into holder.
venipuncture site.  Gently push the tube onto the needle holder so
 Allow the alcohol to dry: that the catheter inside the needle holder
o Disinfect the site penetrates the tube.
o Prevent a burning sensation  Blood flow should be visible at this point.
 Patients are quick to complain about a painful
venipuncture.
 The stinging sensation caused by undry alcohol
is a frequent, yet easily avoided, cause of
complaints.
 Allow tubes to fill until the vacuum is exhausted to
ensure the correct blood to anticoagulant ratio.

Collecting Blood MT 103 (PMLS) LEC NOTES

Lesson 4: Blood Collection (Phlebotomy)
 Collecting Blood
Blood Won’t Flow
 If you do not see blood flow, the tip of the needle:
o May not yet be within the vein.
o May have already passed through the
vein.
o May have missed the vein entirely.
o May be pushed up against the inside wall
of the vein.
Troubleshooting
Incomplete collection or no blood is obtained:
 Change the position of the needle. Move it
forward (it may not be in the lumen)
 or move it backward (it may have penetrated too
far).
 Adjust the angle (the bevel may be against the
vein wall).
 Loosen the tourniquet. It may be obstructing
blood flow.
 Try another tube. There may be no vacuum in the
one being used.
 Re-anchor the vein. Veins sometimes roll away
from the point of the needle and puncture site.
Other Problems
 A hematoma forms under the skin adjacent to the
puncture site - release the tourniquet immediately
and withdraw the needle. Apply firm pressure.
Collecting Blood
Hematoma Formation
 Rapid swelling near the venipuncture site due to
blood leaking into the tissues.
Other Problems
 The blood is bright red (arterial) rather than
venous. Apply firm pressure for more than 5
minutes.
Multiple Tube Collection
 If you are drawing more than one tube:
o Keep a firm grip in the needle holder
while pressing down on the patients arm.
o Use your other arm to interchange tubes.
Multiple Venipuncture Attempts
 Try again below the first site, on the other arm or
on a hand or wrist vein.
 If the second attempt is unsuccessful, ask
someone to take over.
Pediatric Venipuncture
 Interaction with a child
 Immobilizing a child
Geriatric Venipuncture
 Meaningful communication is important
o Alzheimer’s disease, arthritis,
coagulation problems, clouding of lens,
or cataracts, hearing loss, skin are less
elastic, Parkinson’s disease, and stroke.
MT 103 (PMLS) LEC NOTES
Lesson 4: Blood Collection (Phlebotomy)
 Needle Disposal
1. Remove the needle from the holder if appropriate,
Collecting Blood and properly discard it in an approved sharps
disposal container.
Order of Draw for Multiple Tube Collections 2. Discard all waste and gloves in the appropriate
Closure Collection Tube Mix by biohazardous waste container.
Color Inverting 3. Wash hands.
Yellow Blood cultures 8 to 10 times
Red Serum (glass tube) Specimen Labeling
Blue Citrate 8 to 10 times  Label specimens at the bedside according to your
Dark green Heparin 8 to 10 times institution’s standard procedures, or apply pre-
Light green PST gel separator printed labels.
tube with heparin  Proper labelling is the single most critical task you
Purple EDTA 8 to 10 times are asked to perform.
Then, other additives
Red/Gray SST Gel Separator 5 times Proper Labeling Generally Includes:
Tube  Patient’s first and last name
Dark Yellow SST Gel Separator  Hospital identification number
Tube  Date & time
Red Serum (plastic tube)  Phlebotomist initials
 Your institution may provide bar coded computer
Removing the Needle generated labels that contain this information.
 Gently release the tourniquet before the last tube
of blood is filled. Summary of Venipuncture Technique
 Remove the last tube from the needle. 1. Requisition form.
 Withdraw the needle in a single quick movement. 2. Greet the patient.
3. Identify the patient.
Apply Pressure 4. Reassure the patient and explain the procedure.
 Quickly place clean gauze over the site and apply 5. Prepare the patient.
pressure. 6. Select equipment and supplies.
 You may ask the patient to continue applying 7. Wash hands and apply gloves.
pressure until bleeding stops. 8. Apply the tourniquet.
9. Select the venipuncture site.
10. Release the tourniquet.
11. Cleanse the site.
12. Assemble equipment.
13. Reapply the tourniquet
14. Confirm the venipuncture site.
15. Examine the needle.
Apply Adhesive Bandage 16. Anchor the vein.
 The practice of quickly applying tape over the 17. Insert the needle.
gauze without checking the puncture site 18. Push the evacuated tube completely into adapter.
frequently produces a hematoma. 19. Gently invert the specimens, as they are
collected.
20. Remove the last tube from the adapter.
21. Release the tourniquet.
22. Place sterile gauze over the needle.
23. Remove the needle, and apply pressure.
24. Activate needle safety device.

Collecting Blood
MT 103 (PMLS) LEC NOTES
Lesson 4: Blood Collection (Phlebotomy)
 Collecting Blood Collecting Blood

Summary of Venipuncture Technique (cont’d) Heel Stick

25. Dispose of the needle.  Veins of small children and infants are too small
26. Label the tubes. for venipuncture;
27. Examine the patient’s arm.  Butterfly needles may be used to collect venous
28. Bandage the patient’s arm. blood in older children.
29. Dispose of used supplies.  Firmly grasp the infants foot.
30. Remove and dispose of gloves.  Do not use a tourniquet.
31. Wash hands.  The heel may be warmed with a cloth to help
32. Complete any required paperwork. increase blood flow.
33. Thank the patient.
 Wipe the collection site with an alcohol prep pad,
34. Deliver specimens to appropriate locations. and allow the alcohol to dry.
 Wipe the site with sterile cotton or gauze, to be
Syringe Specimen Collection
sure all the alcohol has been removed.
 Small or delicate veins that might be collapsed by
 Puncture the left or right side (outskirt) of the heel,
the vacuum of the evacuated tube system.
not the bottom of the foot.
 May also be used to collect blood culture
 Wipe away the first drop of blood since it may
specimens.
contain excess tissue fluid or alcohol which could
alter test results.
Finger Stick-Specimen Collection
 Collect the blood into the appropriate tube.
 A safety Lancet, which controls the depth of
 Do not: Squeeze the infant’s foot too tightly and
incision.
wipe with alcohol during the collection.
 Finger-sticks should not be performed on children
 After collection is completed, apply pressure to
under one year of age.
the puncture site with a sterile gauze pad until
bleeding has stopped.
Finger Stick
 Do not apply an adhesive bandage to an infant’s
 If possible, use the fourth (ring) finger or the
foot since it may injure its delicate skin.
middle finger.
 Many patients prefer that you use fingers on their
Heel Stick Neonatal Blood Collection
nondominant hand.
 These devices are designed to control the depth
 Choose a puncture site near the right or left edge
of incision, since going too deep into an infant’s
of the fingertip.
heel could injure the heel bone, and cause
 Clean the site as you would for routine
osteomyelitis (bone infection).
venipuncture.
 Select a safety lancet appropriate for the size of Butterfly
the patient’s finger.
 Butterfly needles (winged infusion set)
 You may warm the finger prior to puncture to
 They are available in smaller gauges, and are
increase blood flow.
used to draw venous blood from children, and
 Make the puncture perpendicular, rather than adults with difficult veins.
parallel, to the finger print.
 Butterfly needles come attached to a small tube
 Wipe away the first drop of blood using gauze to which may be connected to:
remove tissue fluid contamination. o An evacuated tube holder, or a syringe.
 Collect blood into an appropriate tube.
 Label specimens appropriately.
 Make sure bleeding has stopped. Apply an
adhesive bandage if necessary.
 Discard sharps appropriately.

MT 103 (PMLS) LEC NOTES

Lesson 4: Blood Collection (Phlebotomy)

Special Situations
Patients Refusing Blood Work
 If someone hesitates to let you collect a blood
specimen, explain to them that their blood test
results are important to their care.
 Patients have a right to refuse blood tests
 If the patient still refuses, report and document
patient refusal.
Fainting
 Rarely, patients will faint during venipuncture.
 It is therefore important that patients are properly
seated or lying in such a way during venipuncture
so that if they do faint, they won’t hurt themselves.
 Self-limited
Fainting: What to do?
 Gently remove the tourniquet and needle from the
patients arm, apply gauze and pressure to the
skin puncture site.
 Call for help.
 If the patient is seated, place his head between
his knees.
 A cold compress on the back of the neck may
help to revive the patient more quickly.
Unsatisfactory Specimens
 They can cause misleading laboratory results
 Must be rejected by the laboratory.
 The patient must then undergo another
venipuncture to get a better specimen.
 It costs time & money to redraw the specimen.
 The credibility of the laboratory is reduced if too
many unsatisfactory specimens are drawn.
Causes of Unsatisfactory Specimens
 Hemolyzed
 Clotted
 Insufficient
 Mislabeled
Special Situations
Hemolysis
 Hemolysis means the breakup of fragile red blood
cells within the specimen, and the release of their
hemoglobin and other substances, into the
plasma.
 A hemolyzed specimen can be recognized after it
is centrifuged by the red color of the plasma.
 Hemolysis can cause falsely increased
potassium, magnesium, iron, and ammonia
levels, and other aberrant laboratory results.
Causes of Hemolysis
 Using a too small needle for a relatively bigger
vein.
 Pulling a syringe plunger too rapidly.
 Expelling blood vigorously into a tube.
 Shaking a tube of blood too hard.
Clots
 Blood clots when the coagulation factors within
the plasma are activated.
 Blood starts to clot almost immediately after it is
drawn unless it is exposed to an anticoagulant.
 Clots within the blood specimen, even if not
visible to the naked eye, will yield inaccurate
results.
Causes of Clots
 Inadequate mixing of blood and anticoagulant.
 Delay in expelling blood within a syringe into a
collection tube.
Insufficient volume
 Short draws will result in an incorrect ratio of
blood to anticoagulant, and yield incorrect test
results.
 Short draws can be caused by:
o A vein collapsing during phlebotomy.
o The needle coming out of the vein before
the collection tube is full.
o Loss of collection tube vacuum before
the tube is full.
Labeling Errors
 Labeling errors are the most common cause of
incorrect laboratory results.
 If detected, the incorrectly labeled specimen will
be rejected.
 If undetected, it will produce incorrect results
which might adversely affect your patient’s care.
MT 103 (PMLS) LEC NOTES
Lesson 4: Blood Collection (Phlebotomy)
 Special Situations

Causes
 Failure to follow proper patient identification
procedure.
 Failure to label the specimen completely and
immediately after collection.

Ten Commandments
1. Thou shalt protect thyself from injury.
2. Thou shalt identify thy patients.
3. Thou shalt stretch the skin at the puncture site.
4. Thou shalt puncture the skin at about a 15 degree
angle.
5. Thou shalt glorify the median vein.
6. Thou shalt invert tubes containing anticoagulants
immediately after collection.
7. Thou shalt attempt to collect specimens only from
an acceptable site.
8. Thou shalt label specimens at the bedside.
9. Thou shalt know when to quit.
10. Thou shalt treat patient’s like they are family.

MT 103 (PMLS) LEC NOTES

Lesson 5: Venipuncture

Venipuncture
 The term venipuncture describes the procedure
of inserting a needle into a vein, usually for the
purpose of withdrawing blood for hematological,
biochemical or bacteriological analysis.
 It is one of the most commonly performed
procedures which, carried out skillfully, carefully
and accurately, will provide high quality blood
samples without causing discomfort to the
patient.
 To perfect the technique of venipuncture, you
should have a good understanding of the arteries,
veins and associated nerves within the arm.
The superficial veins of the arm are usually chosen for
venipuncture, namely:
1. Basilic
2. Cephalic
3. Median cubital veins in the antecubital fossa
These veins are recommended as they are well supported
by muscle and connective tissue, visible and easy to
palpate.
The walls (outer structure) of veins consist of three layers
of tissues that are thinner and less elastic than the
corresponding layers of arteries.
Veins include valves that aid the return of blood to the
heart by preventing blood from flowing in the reverse
direction.
There are two stages to locate a vein:
1. Visual inspection
 The scrutiny of the veins in both arms is
essential prior to choosing a vein.
2. Palpation
 Palpation is also an important
assessment technique as it:
o Determines the location and
condition of the veins
o Distinguishes veins from arteries
and tendons
o Identifies the presence of valves
Veins to Avoid through Visual Inspection:
1. Veins close to an infection.
2. Veins close to bruising and phlebitis.
3. Edematous limbs as there is danger of stasis of
lymph, predisposing to such complications as
phlebitis and cellulites.
4. Areas of previous venipuncture should be
avoided as a build-up of scar tissue can cause
difficulty in accessing the vein and can result in
pain.
5. Avoid veins that are thrombosed (blood clot forms
inside a hemorrhoid).
6. Do not use the affected arm in mastectomy
patients.
7. A vein sited in the region of a drip site should
never be used as it may result in the collection of
a diluted samples.
Locating a Vein through Palpitation:
 Healthy veins feel soft and bouncy and will refill
when depressed.
 Palpate to locate vein.
o Tapping with two fingers will help
sometimes.
Improving Venous Access
1. There are a number of methods to improve
venous access, for example:
o Application of a tourniquet
o Promotes venous distension
o The tourniquet should be tight enough to
impede venous return but not restrict
arterial flow.
o The tourniquet should be placed about 3-
4 inches above the venipuncture site.
o The tourniquet should not be left on for
longer than 1 minute as it may result in
hemoconcentration or pooling of the
blood, leading to inaccurate blood
results.
MT 103 (PMLS) LEC NOTES
Lesson 5: Venipuncture

Proper Way of Tourniquet Application
Improving Venous Access (cont’d)
2. Opening and closing of the fist
 The muscles will force blood into the
veins and encourages distension.
However, this action may affect certain
blood results, e.g. potassium
3. Light tapping of the vein
 May be useful but can be painful and may
result in the formation of a hematoma in
patients with fragile veins.
4. The use of heat
 In the form of warm pack to encourage
vein dilatation and venous filling.
5. Lowering the arm below the level of the heart.
Skin Preparation
 Asepsis is vital when performing venipuncture as
the skin is breached and a foreign device is
introduced into a sterile circulatory system.
 Skin cleaning is a controversial subject and it is
acknowledged that a cursory wipe with an alcohol
swab does more harm than good as it disturbs the
skin flora.
o Clean the venipuncture site:
▪ Use fresh alcohol wipe
▪ Start at site and use circular
motion.
 However, where time permits and always for
blood culture sampling or if the patient is at
increased risk of infection the skin should be
cleaned with an alcohol swab BUT you must allow
at least 2 minutes for the area to dry thoroughly
before proceeding with venipuncture.
Hand Hygiene
 Is the single most important activity for reducing
the spread of disease, yet evidence suggests that
many health care professionals do not
decontaminate their hands as often as they need
to, or use the correct technique which means that
areas of the hands can be missed.
Things to remember when doing hand washing
preparation for venipuncture:
 Improper drying can contaminate again the hands
that have been washed.
 Wet surfaces transfer organisms more effectively
than dry ones and inadequately dried hands are
prone to skin damage.
 Disposable paper hand towels of good quality
should be used to ensure hands are dried
thoroughly.
Safety of the Practitioner
 It is recommended that well-fitting gloves are
worn during any procedure that involves handling
of blood and body fluids, particularly with
venipuncture.
o This is to prevent contamination of the
practitioner from potential blood spills.
 While it is recognized that gloves will not prevent
a needle stick injury, the wiping effect of the glove
on a needle may reduce the volume of blood to
which the hand is exposed, thereby reducing the
volume inoculated and the risk of infection.
o Used needles should always be
discarded directly into an appropriate
sharp’s container, without being re-
sheathed.
 Specimens from patients with known suspected
infections such as hepatitis or HIV should be
double bagged in clear polythene bags with a
biohazard label attached.
 The accompanying request forms should be kept
separate from the specimen to avoid
contamination (two pockets in bag).
Complications during Venipuncture
I. Inability to obtain specimen due to:
 Inappropriate choice of vein
 Thrombosed vein (due to previous or repeated
attempts)
 Inexperience of operator
 Patient shocked, cold or dehydrated causing
vasoconstriction
II. Formation of hematoma due to:
 Poor technique
 Pressure not being applied to puncture site
following removal of needle
MT 103 (PMLS) LEC NOTES
Lesson 5: Venipuncture
 5. Clean site for 30 seconds (DO NOT TOUCH
AGAIN)
Complications during Venipuncture 6. Leave the punctured site to dry
7. Release Tourniquet
III. Puncture of an artery:
 If an artery is punctured: Procedure:
o Release the tourniquet 1. Put on gloves and reapply tourniquet.
o Remove the needle and apply firm 2. With your non-dominant hand; grasp the patient’s
pressure for a minimum of 5 minutes. arm firmly using your thumb to draw the skin taut
o Cover the site with a dressing once and anchor the vein beneath proposed puncture
bleeding has stopped. site.
o Re-check for signs of bleeding in 20 3. Insert the needle at approximately 30 angle with
minutes. bevel up.
4. Aim to insert the needle swiftly through the skin
Approach and Communication and into the lumen of the vein, avoiding trauma
 Introduce yourself & check patient’s ID and excessive probing.
 Explain what you would like to do & why. 5. Connect each specimen tube in turn, in order of
 Gain consent for the procedure and check for draw, onto the assembly (blood culture, no
allergies, shunts, etc. additive, gel, liquid).
 Review the treatment plan & individuals past 6. As each tube is filled, remove it from the
history as necessary vacutainer assembly by gentle rotation and
 Talk the patient through it in a considerate and traction, whilst supporting the vacutainer
courteous manner assembly in the vein.
7. Invert each tube as required.
Sterility 8. Continue until all tubes have been used
 Wash hands 9. When the final tube is drawn, release the
 Clean and prepare procedure tray tourniquet, remove the tube from the needle and
then remove the needle from the vein using a
 Collect equipment into the tray taking care not to
swift backward motion.
contaminate key parts
10. Dispose of any sharps, connector or other
assembly promptly into a sharps container.
Equipment
11. Apply gauze and ask patient to apply firm
 Skin cleansing wipes
pressure for 2 minutes if required.
 Tourniquet
12. Apply sterile plaster to puncture site if required.
 Gloves
 Gauze swabs
Venipuncture using the Evacuated Tube Method
 Adhesive dressing (ETS).
 Needles
 Blood collection tube(s)
 Sharps container

Assemble the vacutainer & needle (or other system)

as per manufacturer’s instructions. Prepare gauze swab
and others for ease of use during the procedure.

Completion and Organization

Preparation:
1. Clear up, dispose of rubbish and clean tray
1. The patient should sit in a suitable chair or be
2. Remove gloves and wash hands
lying down.
3. Label the tubes correctly at the bedside (Tell
2. Support arm on pillow or in other suitable manner
examiner)
& position the patient’s arm as needed.
4. Ensures that the samples are sent to the lab
3. Check for any contra-indications e.g. infection or
5. Thank the patient and ensure they are well
trauma
6. Document in the patients notes
Preparation: (cont’d)
4. Select suitable vein MT 103 (PMLS) LEC NOTES
Lesson 5: Skin Puncture
 Skin and other tissue and richly supplied with
capillaries, so when few ml of blood is required, capillary
puncture is done.

Skin Puncture
 It is the method of choice in pediatric patients.
 It can be used in adults with:
o Extreme obesity
o Severe burn
o Thrombotic tendencies

Sites of Obtaining Blood:

1. Fingertip
2. Heel
3. Great toe
4. Ear lobe

Requirements:
1. Lancet
2. Spirit swab
a. Sterile cotton
3. Dry cotton

Technique:
1. Identification of patient.
2. Select an appropriate puncture site.
3. Clean the puncture site with antiseptic and allow
the area to dry.
4. Make the puncture with a sterile lancet.
5. Discard the first drop of blood by wiping it away
with a sterile pad.
6. Collect the specimen in a suitable container.
7. Place gauze on the site and ask the patient to
apply pressure until the bleeding stops.
8. Dispose of used supplies in biohazard container.
9. Thank the patient.
10. Remove gloves and wash hands.
11. Complete paperwork.
12. Deliver sample to the laboratory.

Disadvantages:
 Can be used for only few tests.
 Dilution of blood with tissue fluid will give false low
count.