CLINICAL CHEMISTRY 1

[TRANS] UNIT PHLEBOTOMY AND SPECIMEN CONSIDERATIONS [LEC]

Figure No. 1 Phlebotomy Sites

OUTLINE
PHLEBOTOMY & SPECIMEN CONSIDERATION
I. The Vascular System
II. Phlebotomy Sites
III. Types of Blood Specimens
IV. Venipuncture Equipment
V. Order of Draw
VI. Venipuncture Procedure
VII. Pre-analytic Considerations
VIII. Capillary Specimen Collection
IX. Specimen Processing

INTRODUCTION
• PRE-ANALYTICAL- sample processing
• ANALYTICAL- chemical analysis
• POST ANALYTIC- data management

PHLEBOTOMY
• The process of collecting blood “to cut a vein”
• Two main phlebotomy procedures:
o Venipuncture
o Capillary puncture

THE VASCULAR SYSTEM

• Network of arteries, veins, and capillaries
• to deliver oxygen from the lungs to tissues and organs, and
nutrients from the GIT
• Arteries and veins has 3 layers of tissue: TYPES OF BLOOD SPECIMENS
o TUNICA INTIMA- innermost; smooth • WHOLE BLOOD
o TUNICA MEDIA- middle; thickest o Contains the plasma and the cellular
o TUNICA ADVENTITIA- outer components
• ARTERIES o Either arterial or venous
o Arterial blood • PLASMA
o bright red o Liquid portion of an anticoagulated blood
o oxygenated blood o Contains fibrinogen
• VEINS- has valves to promote the continuous flow of blood, • SERUM
and prevent back flow o Liquid portion of clotted blood
o Venous blood o Plasma minus the fibrinogen
o dark red o No additive tube, pro-coagulation tube, gel & clot
o deoxygenated activator tube
• CAPILLARIES- only 1 layer of tissue
o Smallest blood vessels Figure No. 2 Serum vs. Plasma
o One cell thick to allow for gas and nutrient exchange
• ANTECUBITAL FOSSA
o where most commonly used veins for venipuncture are
located
• Vein of choice:
o MEDIAN CUBITAL VEIN – most secure, stable, and least
painful to puncture
o CEPHALIC – usually most prominent in obese patients
o BASILIC VEIN – beneath it are nerves which may lead to
paralysis
• Other sites:
o Hand veins
VENIPUNCTURE EQUIPMENT
▪ Especially the dorsal venous network
▪ When the antecubital veins are not accessible • TOURNIQUET
▪ Smaller, less anchored, and can be very painful o Pliable rubber or a strip with Velcro used to locate the
o Base of the Foot patients’ veins
o Must not be left on longer that 1 minute to prevent

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hemoconcentration

• WINGED INFUSION SET (BUTTERFLY)

• NEEDLE o Luer fitting for syringe
o Size – gauge and bore are inversely related o Luer adapter for ETS
o 21 GAUGE- standard for routine venipuncture o Used to collect blood from people with small, fragile veins
▪ The lower the gauge, the longer the needle, the bigger (elderly and children)
the bore o Provides greater control with non-stable patients
▪ The higher the gauge, the shorter the needle, the • COMMON STOPPER COLORS AND ADDITIVES
smaller the bore o ANTICOAGULANT
▪ Light blue (citrate) – sodium citrate
Figure No. 3 Parts of Needle ▪ Green (heparin)
▪ Lavender (EDTA)
o ANTIGLYCOLYTIC AGENTS
▪ Gray (Sodium Fluoride)
▪ To inhibit blood cells from further metabolizing the
blood sugar in serum
o CLOT ACTIVATORS
▪ Red (Silica)
▪ Orange (Thrombin)
o THIXOTROPIC GEL
▪ Gold (serum)
▪ Light green (plasma)
▪ Pearl or white (plasma)

Figure No. 4 Needle Gauges ORDER OF DRAW

• To reduce the risk of specimen contamination by
microorganisms and additive carry-over
TUBE STOPPER ADDITIVE DEPARTMEN
COLOR T
1 Blood Culture Yellow SPS Sodium Microbio
Tubes Polyanethol
Sulfonate (SPS)
2 Coagulation Light Blue Sodium Citrate Hematology
Tubes
3 Glass Non Red / Red None Chem, BB,
Additive Tube Rubber Serology

4 Plastic Clot Red Silica Particles Chem

Activator Tube
Serum Gold plastic/ Silica Particles, Chem
separator Read and Thixotropic gel
Tubes Gray rubber
5 Plasma Light Lithium heparin, Chem
separator green/green Thixotropic gel
Tubes and gray
rubber
Heparin Green Lithium heparin
Tubes
• THREE BASIC METHODS 6 EDTA Tubes Lavender, EDTA Hema, BB
o EVACUATED TUBE SYSTEM Pink
▪ Most common way of collecting, except PH Plasma Pearl top EDTA, Molecular
(public hospitals) Separator Thixotropic gel diagnostics
• Multisample needle Tubes
7 Fluoride tubes Gray Sodium Chem
• Tube holder (barrel/adapter)
Fluoride/Potassi
• Evacuated Tubes um Oxalate
• NEEDLE & SYRINGE ESR Tubes Black Sodium Citrate Hema
o Syringe needles Table No. 1 Order of Draw
o Syringe
o TRANSFER DEVICE – transfer blood from syringe to • Blood Culture Tubes
tube; to prevent accidental puncture or needlestick o First because of the need to maintain sterility
• Coagulation Tubes
o 2nd because of the PT and PTT – most affected by other

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additive - Needle too deep

• Thixotropic gel • retract the needle (if ETS, you need to remove first the tube
o separation of serum and cellular components before retracting)
• Silica particles - Needle not deep enough
o promote clotting
- Needle beside the vein
• Gold
• slightly retract the needle, then redirect to the vein
o advantage: prevents the blood cells from further
metabolizing the nutrients - Collapsed vein
- Undetermined needle position
• PLASTIC CLOT ACTIVATOR TUBE & SERUM SEPARATOR
TUBES
o 4th; the effect of clot activators can be over leaded by the
effects of anticoagulant
• PLASMA SEPARATOR TUBES/HEPARIN TUBES
o 5th
o mostly used for electrolytes, protects the plasma from
further being contaminated
• EDTA
o one of the last
o can cremate RBCs, and may lead to the leaking of
electrolytes PREANALYTIC CONSIDERATIONS
• Fluoride Tubes
• Burns, Scars, and Tattoos
o mostly used for determination of blood sugar and blood
➔ prone to infection, and tattoos may contaminate the blood
alcohol levels
• Damaged Veins
➔ Some may appear hard (sclerosed) and cordlike
• Hematoma – painful
• Edema
• Mastectomy
SPECIAL EVACUATED TUBES
• IV Therapy
1. ACD Tubes ➔ look for vein in the other arm
o Yellow • Heparin and Saline Lock
o Na3 Citrate ➔ look for vein in the other arm, or collect blood below the
o Citric Acid, and Dextrose insertion point; ask the nurse to turn off IV for two minutes
o Cellular Immunology • Fistulas
o Mostly used for tissue typing to determine if tissues and ➔ surgical fission of the artery and vein of a dialysis patient
organs are compatible for transplantation • Cannulas
2. Trace Elements Tube ➔ external tubing that connects the artery and the vein = look
a. Royal blue for other sites
▪ Plain
▪ Na Heparin, or Na2 EDTA PROCEDURAL ERROR RISKS
▪ Chemistry (Toxicology)
• Hematoma - to avoid, do not probe the vein
▪ Mostly used for measurement of trace elements
b. Tan or Light Brown • Iatrogenic Anemia - anemia caused by excessive blood
▪ K2 EDTA, Na Heparin collection, especially to the children and the elderly
▪ Lead Testing • Infection of the site - disinfect, and do not touch the site
• Nerve Injury - shooting pain, electric-like tingling, or numbness
3. Blood Donor Bags
• Inadvertent Arterial Puncture - hit the artery below the needle
(if this happens, prolong the pressure for around 5 mins to avoid
VENIPUNCTURE PROCEDURE hematoma)
Must know: • Reflux - blood may backflow and may lead to embolism
- Tourniquet • Vein Damage (Scar)
• 3-4 inches above the antecubital are
- After removal from the holder, additive tubes must be inverted PATIENTCONDITIONS AND COMPLICATIONS
gently 3 – 8 times • Allergies to supplies and equipment - look for alternatives
• For silica: 3-5 times, for anticoagulant: 5-8 times • Excessive bleeding - prolong pressure for 5 mins
- The needle to be inserted in performing a venipuncture is • Fainting (syncope)
usually at 15-30° angle - stop collection
- Needle (0.5 cm below the target site) - lie down flat, or bend their head in level with the heart, give
something sweet
TROUBLE SHOOTING • Nausea or vomiting - breathe slowly or in a paper bag
- Tube Position • Obese patients
• make sure that the tube is seated correctly to the tube • Pain - withdraw immediately
holder • Petechiae - poor platelet function
- Bevel against the vein upper/lower wall– • Seizures and convulsion
• Remove the evacuated tube, slightly withdraw the needle

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CAPILLARY SPECIMEN COLLECTION SPECIMEN SUITABILITY

- Blood collected from a skin puncture (lancet) • Hemolyzed
COLLECTION SITE • Collection in the wrong tube
• Outer area of the bottom of the foot (heel stick) • Failure to follow timing and handling requirements
• Fleshy part of the last phalanx of the 3rd or 4th finger
• Fleshy portion of the earlobe - prone to infection and
• Quality not sufficient (QNS)
contamination of tissue juices which may alter the concentration • Clotting in whole blood or plasma specimen
of analyte
CENTRIFUGATION
• Specimens must be completely clotted (30-60 minutes at room
temp.)
• Visually check
a. Lipemic
- Cloudy turbid appearance, indicates presence of lipid and
a non-fasting specimen
- Interferes with colorimetric analysis
b. Hemolyzed
MATERIALS - Destruction of RBC results in plasma/serum appearing red
o Alcohol, Gauze, Bandages to pink
o Lancets - Affects K+ and enzyme testing
o Warming Devices c. Icteric
o Microcollection Tubes - Specimen with a yellowish appearance due to increased
o Microhematocrit Tubes – Sealants bilirubin content

CAPILLARY ORDER OF DRAW STOPPER REMOVAL

1. Blood Gases (Heparin; Green) • Stopper removal devices
2. EDTA (lavender) • Face shield
3. Sodium Heparin (Green)
• Splash shield
4. Other additive (gray antiglycolytic agent, gold clot activator)
5. Nonadditive ALIQUOT PREPARATION
• For multiple tests in a single specimen
• Stored at 4 or -20°C for 8 hours

INDICATIONS FOR CAPILLARY PUNCTURE

• Inaccessible veins
• Fragile veins
• Thrombotic tendency
• POCT (glucose monitoring)
• Small blood volume
• New born screening - collected by heel puncture and placed
within printed circles on filter paper

SPECIMEN PROCESSING
ROUTINE HANDLING
Mixing tubes
➔ Invert 3-8x (3-5 for clot activators, 5-8 for anticoagulants)
Transporting specimens
➔ Plastic bag with a biohazard logo, closure and slip pocket
Delivery Time Limits
➔ 45 minutes after collection
➔ Centrifuged within 1-2 hours

SPECIAL HANDLING
• Maintained at 37°C (heat block)- cold agglutinins
• Chilled (using crushed ice)- analytes
• Wrapped on foil – bilirubin

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