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ADVANCE PATHOLOGY
1.2A BASIC PRINCIPLES OF PHLEBOTOMY
PHLEBOTOMY: HISTORICAL PRACTICE

1.
2.
3.
4.

MODERN PHLEBOTOMY
Diagnosis and management of disease
Remove blood for transfusions
Therapeutic reasons:

Polycythemia

Hemochromatosis
BLOOD FUNCTION
Supplies nutrients to tissues:
O2, hormones, glucose
Removes end-products of metabolism:
CO2, urea, creatinine
Provides defense mechanism: WBC, antibodies
Prevents blood loss:
platelets, coagulation proteins

COAGULATION REACTION
Clotting factors + calcium thrombin
Fibrinogen + thrombin
fibrin strands

ANTI-COAGULANTS
Remove calcium
Neutralize thrombin
Whole blood
Plasma
Serum

BLOOD WITH ANTI COAGULANT

Clotting is prevented and irreversible
Mix: completely invert 8-10x
Whole blood
Centrifuge plasma
Plasma contains fibrinogen

BLOOD WITHOUT ANTI COAGULANT

Spontaneous clotting occurs and is irreversible
Fibrinogen fibrin strands
Fibrin strands entrap cells
Centrifuge serum
Serum lacks fibrinogen

APPEARANCE
Normal: clear and yellow
Abnormal:

Hemolyzed = pink to red (ruptured RBC)

Icteric = dark orange-yellow (bilirubin)

Lipemic = cloudy (fat, triglycerides)

BLOOD COMPOSITION

Formed elements (~45%)

RBC
WBC
Platelets
Fluid component (~55%)
Water (~92%)
Protein (~7%)
etc

BLOOD COLLECTION TUBES:

Contain a vacuum
Used with
Vacutainer and Syringe systems
Stoppers universal
color coded: indicates contents

Have an expiration
date

COAGULATION
In vivo

Blood is fluid

Clot is formed to
protect injured vessel
In vitro

Spontaneous reaction

Triggered by glass or poor drawing technique

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1.2A BASIC PRINCIPLES OF PHLEBOTOMY

TUBES CONTAINING NO ANTI-COAGULANT

TUBES CONTAINING ANTI-COAGULANT

RED-TOP TUBE

Glass

Plastic

No additive
Glass surface
activates clotting
sequence
Do not mix
SERUM: use for serum chemistry, Therapeutic Drug
Monitoring
Contain additive to activate clotting sequence
Contain inert gel serum separating tube (SST)
Do invert to mix additive and initiate clotting
sequence
SERUM: use for serum chemistry

GOLD OR MOTTLED RED-GRAY TOP TUBE

Contain clot activator and gel
(SST)
Invert to mix and initiate
clotting sequence

PURPLE-TOP TUBE
Anticoagulant = EDTA
Binds calcium
PLASMA whole blood

Hematology studies: CBC, immunohematology

SERUM: serum chemistry

ROYAL BLUE-TOP BLUE

Trace metal-free
Iron, copper, zinc
Label color indicates contents:

Red: no additive = serum

Purple: EDTA = whole blood or

plasma

Green: heparin = whole blood or plasma

BLUE-TOP TUBE
Anticoagulant = sodium citrate
Binds calcium
PLASMA whole blood
Must be full
Blood:anticoagulant ratio critical
Must be on ice if not analyzed within 30 minutes
Coagulation studies (e.g. Prothrombin, aPTT)
GREEN-TOP TUBE
Anticoagulant = heparin
Three formulations:
Lithium heparin
Ammonium heparin
Sodium heparin
Inhibits thrombin formation
Must be full and on ice if need pH, ionized Ca
Most plasma chemistry tests, STAT lab (plasma separator
tube/PST)
Decreases time needed for blood to clot,
Makes turnaround time better
GREY-TOP TUBE
Anticoagulant = potassium oxalate

Binds calcium

PLASMA, Whole blood

Antiglycolytic agent = sodium fluoride

Maintains plasma glucose levels

Limited use: plasma glucose, plasma lactic acid

YELLOW-TOP TUBE
ACD = acid citrate dextrose

Paternity testing

DNA
SPS = sodium polyanethol sulfonate

Used for special blood culture studies

Inhibits certain antibiotics

Both bind calcium

PLASMA, Whole blood

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1.2A BASIC PRINCIPLES OF PHLEBOTOMY

MODES OF TRANSMISSION
Parenteral: any route other than the digestive tract

Intramuscular
Non-intact skin: chapped hands,

Intravenous
cuts, cuticles

Subcutaneous
Percutaneous: needles, sharps

Mucosal
Permucosal: mouth, nose, eyes
Ingestion

INFECTION CONTROL
Stop the spread of infection
Hand washing

Primary means of preventing spread of infection

(especially nosocomial)

Minimum 15 seconds, soap, friction

Wash hands before and after each blood draw

PPE

Lab coat

Gloves

Mask
Standard precautions at all times
ENGINEERING CONTROL

1.
2.

3.
4.

6.

VALID TEST RESULTS REQUIRE

Trained personnel

Causes of pre-analytical error

Invalid test results

Quality control
Quality assurance
Sophisticated
instruments

SAFETY PRACTICES
For infection to spread:
1. Infectious substance: HBV, HCV, HIV
2. Mode of transmission
3. Susceptible host

5.

TYPE AND AMOUNT OF SPECIMEN

Dependent upon

Test
Whole blood: EDTA or heparin?
Plasma: EDTA or heparin?
Serum: trace free? Separator gel interference?

Amount of sample needed to perform test

Multiple labs needing the same specimen at the

same time

7.

Sharps container:

Discard needles, lancets

Biohazard marking

Puncture resistant

NEVER recap, bendbreak needles

Tourniquets:

Slows venous blood flow down

Causes veins to become more prominent

NEVER leave on for >1 minute

AVOID rigorous fist clenching or hand pumping

(potassium, lactic acid)

Latex allergy

Needles

NEVER reuse a needle

NEVER use if shield is broken
NEVER recap, cut, bend or break
Drop immediately into sharps container after
venipuncture
Size of needle is indicated by gauge:

Larger gauge number indicates smaller

needle diameter

21, 23 gauge needles routinely used for

phlebotomy

Used with syringe system

Used with vacutainer system

MULTI-SAMPLE NEEDLE

PPE
Sharps containers
Safer medical devices
EQUIPMENT
PPE: gloves, lab coat, mask
Cleaning agent

Alcohol pads: routine

Povidone iodine: blood culture collection and blood

gases

Soap and water: alcohol testing, allergies

Cotton balls, gauze
Bandage, tape (use caution with children)
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1.2A BASIC PRINCIPLES OF PHLEBOTOMY

BUTTERFLY NEEDLE

Most often used with syringe

Expensive, thus not used for routine draws
Used for small, fragile veins
Increased risk of needle stick injury

8.

Tube holder/
vacutainer adapter

Threaded

Flanges

Syringe

Quick
Least risk of accidental needle stick
More control
Reposition easily
Will see flash of blood in syringe hub when vein
successfully entered

THE PATIENT
Approach
Communication
Empathy
Handling special situations
Patient identification

Arm band

Legal document
Prepare patient for blood draw

Latex allergy?
SELECTING THE SITE
Antecubital area most
often accessed
Hand or wrist
Remember: 2 arms
Use tip of index finger on
non-dominant hand to
palpate area to feel for
the vein

9. Syringe
10. Black
water proof
pen

LABELING BLOOD COLLECTION TUBES

Black indelible marker (water proof)

Never pencil

Legal document

Print legibly
Required information: 5 items

Patient name

Identification number

Date of draw (mm,dd,yyyy)

Time of draw (military time)

Phlebotomist signature: first initial, last name

COLLECTION SITE PROBLEMS

Veins that lack resiliency
Extensive scarring
Hematomas
Edematous area
Side of mastectomy
Intravenous line
NEVER draw above an IV
Draw from other arm
Draw from hand on other arm
Draw below the IV

VACUTAINER OR SYRINGE?
Vacutainer

Most often used

Most economical
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1.2A BASIC PRINCIPLES OF PHLEBOTOMY

INSERTING THE NEEDLE

Anchor the vein

Grasp arm with your non-dominant hand

Use thumb to pull skin taut

Smoothly and confidently insert the needle bevel up

15-30 degree angle

(military time)
Your initials
FAILURE TO OBTAIN BLOOD
Check tube position and vacuum

Always have back up tubes near by

Needle position
Collapsed vein
NEEDLE POSITION

NO NEEDLE MOVEMENT
You must anchor the blooddrawing equipment on the
patients arm to minimize
chance of injury

FILL TUBES
Use correct order of draw:

Blood cultures

Red top

Blue (baby blue)

Green

Purple

Grey

BE CAREFUL NOT TO:

Push needle further into vein when engaging evacuated tube
Pull needle out of vein when disengaging tube
Pull needle out of vein as you pull back on the plunger
Pull up or press down when needle in vein
Forget to mix additive tubes 8-10 times

WITHDRAW NEEDLE
First release tourniquet
Disengage tube
Place cotton directly over needle, without pressing down
Withdraw needle in swift, smooth motion
Immediately apply pressure to wound
Do not bend arm

LATERAL TUBES IMMEDIATELY

In sight of patient
Patient name
Identification number
Date of draw
Time of draw

YOU SHOULD TRY AGAIN

Look at alternate site

Other arm

Hand
Use clean needle
Use fresh syringe if contaminated
Only try twice
POOR COLLECTION TECHNIQUES
Venous stasis

Prolonged application of tourniquet (>1 min)

Hemodilution

Drawing above IV

Short draw (blood to anticoagulant ratio)

Hemolysis

Traumatic stick

Too vigorous mixing

Alcohol still wet

Using too small of needle

Forcing blood into syringe

Clotted sample

Inadequate mixing

Traumatic stick
Partially filled tubes

Short draw

Sodium citrate tube draw volume critical

Using wrong anticoagulant
Specimen contamination

Using incorrect cleanser

Alcohol still wet

Powder from gloves

Drawing above IV
Specimen handling

Exposure to light

Pre-chilled tube

Body temperature
VENIPUNCTURE PROCEDURE
Remain calm
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Organize yourself
Organize your equipment: STICK TO ELEVEN

Gloves

Lab coat

Alcohol wipe

Cotton ball

Bandage/tape

Sharps container

Tourniquet

Needle

Syringe or vacutainer holder

Collection tubes with backup tubes

Water-proof marker
Wash hands
Put on gloves
Identify patient
Latex allergy?
Position arm
Apply tourniquet

Locate vein
Release tourniquet
Cleanse site in outward
rotation

Allow to air dry

Reapply tourniquet

Do not
contaminate
site
Anchor vein
Insert needle
Fill tubes

Quick mix
additive tubes
Release tourniquet
Withdraw needle

Engage safety device

Dispose of needle
immediately
Apply pressure to
puncture site
Label tubes
Recheck puncture site
Thank patient
Remove gloves,
wash hands

1.2A BASIC PRINCIPLES OF PHLEBOTOMY

SYRINGE DRAW

Syringe Safety Transfer Device

1.
2.

ACCIDENTAL NEEDLE STICK

Remain calm
Cleanse wound with alcohol

Wash wound thoroughly

Notify supervisor, instructor
Follow site protocol
Visit hospital infection control unit (HICU)
Complete incident report

Mark your spot

SKIN PUNCTURE
Method of choice for infants, children under 1 year
Adults

Scarred

Fragile veins

Hardened veins

Home glucose monitoring (POCT)

Patients with IV
CAPILLARY BLOOD
Mixture of arterial, venous, capillary blood and fluid from
surrounding tissues
Fluid from surrounding tissues may interfere and/or
contaminate the specimen
Warming skin puncture site increases arterial blood flow to the
area
Reference ranges often differ from venous (usually capillary
blood values are lower)
SKIN PUNCTURE EQUIPMENT
PPE
Cleaning agent

Alcohol pads: routine

Soap and water: alcohol testing, allergies

DO NOT use providone iodine

Interferes with bilirubin, uric acid, P, K

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3.
4.

Cotton balls, gauze

Bandage/tape

5.
6.

Sharps container
Warming device

Commercial warmer

Warm wet washcloth

7.

Lancet

8.

9.

1.2A BASIC PRINCIPLES OF PHLEBOTOMY

Heparin

Serum
Apply pressure to puncture site
Label specimen in sight of patient (indelible marker)

Always use
standardized equipment

NEVER use a surgical

blade
Micro-specimen containers

Capillary tubes

Microtainers

Capillary blood gas

tubes

Micropipet diluting
system
Glass slides: used to prepare
blood smears

1.
2.
3.
4.
5.
6.
7.
8.
9.

SKIN PUNCTURE PROCEDURE

Wash hands
Approaching the patient
Patient identification
Latex allergy?
Bedside manner
Site selection
Cleanse site: DO NOT use providone- iodine
Perform puncture: Wipe away first drop of blood
Label the specimen

Hold finger between your index finger and thumb

Puncture the finger using a quick, smooth motion
Wipe away the first drop of blood

SKIN PUNCTURE
Capillary blood collection is inappropriate for:
Severely dehydrated patients
Patients with poor circulation
Coagulation studies requiring plasma specimens
Tests that require large volumes of blood (i.e. Erythrocyte
Sedimentation Rate (ESR) and blood cultures)
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END OF TRANX

Collect sample

DO NOT touch collecting device to skin surface

DO NOT scrape collecting device across skin

surface

DO NOT scoop blood into collecting device

Order of draw is critical: platelets accumulate at puncture site

causing clot formation

Blood smear

EDTA
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