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Learning Objectives

▪ Familiarize with different materials used in phlebotomy and to know its

function.
▪ Identify the correct order of draw
▪ Familiarize and perform the correct phlebotomy procedures.

Phlebotomy
Mark Kerwin F. Sayas, RMT, MSMLS (c.)
Course Facilitator

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Evacuated Tube System Needles

▪ Most frequently used method for venipuncture. ▪ Multisample needles
▪ Blood is collected directly into the evacuated tube ▪ Hypodermic needles
▪ Winged blood collection needles
▪ Advantages
▪ Eliminating the need for transfer of specimens
▪ Minimizing the risk of biohazard exposure ▪ Should be sterile, disposable and SINGLE USE.
▪ Sizes vary by both length and gauge (diameter)
▪ ROUTINE: 1 inches to 1.5 inches

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Needles Needles
▪ Gauze – diameter of the needle bore
▪ Gauge 16 = blood collection units
▪ Gauge 23 = smaller veins
▪ Gauge 22 = commonly used
▪ The smaller the gauge number the bigger the diameter
▪ Smaller gauge may cause hemolysis
▪ Color coded for easy identification Needles should never be recapped once the
shield is removed regardless of whether
they have or have not been used.

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Needle Holders

▪ Made of plastic
▪ Act as safety shield for the used needle

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Loss of tube vacuum is a primary cause of

failure to obtain blood. The venipuncture can be
performed before placing the tube on the needle. Sharps
Practice both methods, and choose the one with Container
which you are most comfortable.

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Collection tubes
Removing the rubber stoppers from evacuated
▪ Evacuated tubes tubes can be hazardous to laboratory personnel
▪ Contain premeasures amount of vacuum
because an aerosol of blood can be produced if
the stopper is quickly “popped off.” Stopper
▪ Vacutainers should be covered with a gauze pad and slowly
▪ 1.8 to 15mL: amount of blood loosened with the opening facing away from the
▪ Sterile, many are silicon coated body.

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Anticoagulants and Additives Lavender (Purple) Top

▪ Binds to calcium ▪ Ethylenediaminetetraacetic acid
▪ Ethylenediaminetetraacetic acid (EDTA) ▪ Contains spray-coated or liquid anticoagulant
▪ Citrate ▪ Tripotassium EDTA (K3EDTA) (liquid)
▪ Oxalate
▪ Dipotassium EDTA (K2EDTA) (spray-coated)
▪ Prevent clotting by inhibiting the formation of thrombin ▪ Coagulation is prevented by the binding of calcium in the
▪ Heparin sample, thereby preventing the participation of Ca2+ in
coagulation cascade.
▪ 8 inversions

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K2EDTA Increased amounts of K2EDTA

▪ Complete blood count ▪ Shrinks RBC
▪ Maintains cellular integrity ▪ Decrease hematocrit, RBC indices and ESR results.
▪ Inhibits platelet clumping
▪ Does not interfere with routine staining procedures
▪ Can also be useful in immunohematology procedures
▪ Can’t be used for coagulation studies because EDTA
interferes with Factor V and the thrombin-fibrinogen
reaction.

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Pink Top White Top

▪ Contains spray-coated K2EDTA ▪ Contains spray-coated K2EDTA and separation gel
▪ Use specifically for blood banking procedures. ▪ Plasma preparation tubes (PPTs)
▪ 8 times inversion ▪ Used for molecular diagnostics
▪ Used for myocardial infarction panels, ammonia levels
▪ 8 times inversion

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Light Blue Top Black Top

▪ Sodium citrate ▪ Contains buffered sodium citrate
▪ Prevents coagulation by binding calcium ▪ For Westergren Sedimentation Rate
▪ 3.2 or 3.8% = preserves labile coagulation factors.
▪ Ratio: 4:1 (Blood to anticoagulant)
▪ 3-4 times inversion
▪ Blood to citrate ratio: 9:1
▪ Increased citrate = interfere the coagulation tests

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Green Top Light Green Top

▪ Heparin (Sodium Heparin, Lithium Heparin, Ammonium Heparin) ▪ Contains lithium heparin and separation gel (Plasma
▪ Prevents clotting by inhibiting thrombin in the coagulation cascade
separation tubes)
▪ For chemistry tests (plasma or whole blood)
▪ Ammonium heparin may interfere BUN ▪ PST
▪ Lithium heparin produce least interference. ▪ Potassium determination
▪ 8 times inversion ▪ Prevents contamination by red blood cell
▪ NOT FOR HEMATOLOGY ▪ 8 inversions
▪ Interferes with Wright Stain
▪ Causes blue background on blood smear

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Gray Top Gray Top

▪ PRESERVING GLUCOSE ▪ Interfere enzymes
▪ Sodium fluoride
▪ Antiglycolytic agent ▪ Not for hematology: oxalate distorts cellular
▪ Maintains glucose stability for 3 days morphology
▪ NOT ANTICOAGULANT

▪ Potassium oxalate or Na2EDTA

▪ Anticoagulant
▪ Prevents clotting by binding calcium

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Royal Blue Top Tan Top

▪ For toxicology, trace metal and nutritional analyses ▪ For lead determinations
▪ With spray-coated silica clot activator serum ▪ Certified to contain less than 0.1 ug/mL of lead
▪ K2EDTA or sodium heparin ▪ K2EDTA
▪ 8 times inversion ▪ 8 inversions

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YELLOW TOP Bactec Bottle

Acid Citrate Dextrose (ACD) Sodium Polyanethol Sulfonate (SPS)
Red cell preservatives Prevents coagulation by binding calcium
Used in blood banking Aids in the recovery of microorganisms
HLA phenotyping Inhibiting the actions of
 Complement
DNA and paternity testing
 Phagocytes and
Prevents clotting by binding calcium  Certain antibiotics

Dextrose prevents the red blood cells

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Light Blue/Black Top Red/Green Top

▪ Sodium citrate and polyester gel ▪ Sodium heparin and polyester gel and density gradient
▪ Whole blood molecular diagnostic testing liquid
▪ 8 times inversion ▪ Whole blood molecular diagnostic testing
▪ 8 times inversions

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Orange Top Red/Gray and Gold Top

▪ Containing thrombin-based medical clotting agent and ▪ Contains clot activator and separation gel
separation gel (Rapid Serum Tubes (RST)) ▪ Serum separator tubes
▪ Clot within 5 minutes
▪ Centrifuged for 10 minutes
▪ 5 times inversion

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Order of Draw for ETS and Syringe

▪ Recommended by Clinical & Laboratory Standards Institute (CLSI)
▪ Blood cultures (yellow stopper tubes, culture bottles)
▪ Light Blue Stopper tubes (Sodium Citrate)
▪ Red/Gray, Gold Stopper tubes (SST) red stopper plastic tubes
(clot activator) and red stopper glass tubes
▪ Green stopper tubes and light green (Heparin)
▪ Lavender stopper (EDTA)
▪ Gray Stopper (Potassium oxalate)
▪ Yellow/gray or orange stopper tubes (thrombin clot activator)

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Syringe Winged Blood Collection

▪ Advantage ▪ Butterfly
▪ Phlebotomist is able to control the suction pressure on the vein ▪ Used for infusions of IV fluids
▪ Used to very small or very fragile veins
▪ Pediatric patients and geriatric
▪ 21 or 23 gauge
▪ ½ to ¾ inch needle

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Tourniquet Vein Locating Device

▪ Make it easier to locate patients’ vein
▪ Flat latex or vinyl strips (MOST COMMON)

▪ Blood pressure cuff

▪ Can be used as tourniquet
▪ 40 mm Hg

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Venipuncture Steps 1. Review and Accession the Test request

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Common Test Status

▪ Stat (Immediately)
▪ ASAP (As soon as possible)
2. Approach, Identify and prepare the patient
▪ Fasting
▪ NPO

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Remember
1. Knock on door gently 3. Verify the patient’s diet restrictions and
2. Make good impression by greeting the patient latex sensitivity
3. Stay organized and have all supplies
4. Maintain calm expression
5. Introduce yourself and explain the procedure
6. Remain compassionate
7. Thank the patient

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4. Sanitize hands 5. Position the patient, apply tourniquet, as the

patient to make a fist

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Tourniquet application REMEMBER!!

▪ 3-4 inches above the site or 4-5 inches width ARTIFACTUAL HYPERALBUMINEMIA
▪ Not more than 1 minute AKA
PROLONG TOURNIQUET APPLICATION
Hemo-concentration
INCREASED in K, A, L, A
Potassium, Albumin, Lactate, Ammonia
P A L A
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6. Select the vein, release tourniquet and ask 7. Clean and air-dry the area
patient to open fist

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8. Prepare the equipment and put on gloves 9. Reapply the tourniquet, uncap and inspect
the needle

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10. Ask the patient to remake a fist anchor to 11. Establish the blood flow, release the
vein and insert needle tourniquet and the ask the patient to open fist

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12. Fill, remove and mix the tubes in correct 13. Place gauze, remove the needle, activate
order of draw the safety feature and apply pressure

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15. Label the tubes

14. Discard the collection unit, syringe needle ▪ PATIENT’S COMPLETE NAME
or transfer device ▪ Date of birth
▪ ID number (IF APPLICABLE)
▪ Date and time of collection
▪ Initials of phlebotomist
▪ Additional information

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16. Observe special handling instructions 17. Check patient’s arm and apply bandage

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18. Dispose contaminated materials 19. Thank the patient, remove gloves and
sanitize hands

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Special Populations
1. Pediatric puncture
20. Transport the specimens to the laboratory 2. Geriatric venipuncture
3. Dialysis patients

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