Chapter 6

Specimen collection, processing

& preservation

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 Objectives
At the end of the chapter, the student will be able to
• Define specimen
• Explain about different kind of specimens
• Discuss about factors affect blood composition
• List the important steps in specimen collection
• Discuss about preservation, storage, and
transportation of specimens.

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 Outline
• Specimen for biochemical analysis
• Specimen types and collection
• Coding of stopper colors
• Skin puncture, arterial puncture, venipuncture
• Common specimen factors affecting test results
• Stability of chemical substances in blood specimen
• Preservation methods
• Anticoagulants and preservatives
• Common anticoagulants
• Physiological factors affecting composition of body fluids
• Specimen transportation
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 Specimen for Biochemical analysis
 Many factors besides diseases affect the composition of
body fluids, e.g. pre-analytical factors

 The effects of theses factors on test values must be

recognized and considered in the evaluation of lab data.

 Many of theses pre-analytical factors are associated with

specimen collection and processing.

 Specimen for analysis is the most important element in

obtaining reliable test results.

 Proper collection, preparation, processing and

preservation of the specimen is essential for
obtaining best results. GIGO 4
 Specimen containers must be labeled at least
with the patient’s name, hospital or
identification number , ward, and date and
time of collection.

 A properly filled test requisition slip should

accompany each specimen.

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 Specimen types and collection
• Biological specimens analyzed in clinical labs
include
 whole blood, serum, plasma, urine, feces, sweat,
saliva, CSF, synovial, amniotic, pleural, pericardial,
peritoneal fluids and various types of solid tissues.
• Blood for analysis may be obtained from veins,
arteries or capillaries. Phlebotomy

• Specimen Types
Random specimen
Fasting specimen
Timed specimen
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 Coding of stopper color to indicate
additive in evacuated blood tubes
COLOR Use Additive

Gray plasma or whole blood with Oxalate (Na, K)

glycolysis inhibition fluoride (Na),
Iodoacetate(Na)

Yellow Sterile interior of tube None

Green plasma or whole blood Heparin (Na,

Li, NH4+ )
Red Serum None
Blue Plasma or whole blood Citrate (Na)

Lavender Plasma or whole blood EDTA (Na, K) 7

Colors of Tubes

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 Blood Specimen Types

• Skin Puncture
– Capillary blood
• Arterial Blood
• Venous Blood

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 Skin puncture

• In young children and in some adults

when only small volume of blood is needed.
• It is predominantly capillary blood.
• Obtained using a sterile blood lancet by making
a puncture on the skin. The tip of finger, or an
ear lobe is the usual site.
• In infants <1 yr, the lateral or medial plantar
surface (sole) of the foot should be used.

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Differences in composition of capillary
and venous serum
Capillary value greater than venous value (%):
 Glucose …….. 1.4
 Potassium ….. 0.9
No difference in capillary and venous:
 Phosphorous
 Urea
Capillary value less than venous value (%):
 Bilirubin …………. 5.0
 Calcium …………. 4.6
 Chloride ………… 1.8
 Sodium …………. 2.3
 Total protein …… 3.3
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 Arterial puncture

• This is used mainly for blood gas analyses.

• The procedure requires considerable skill and is
usually performed only by physicians or specially
trained technicians or nurses.
• The preferred sites of arterial puncture are, in
order:
 The radial artery at the wrist
 The brachial artery in the elbow
 The femoral artery in the groin.

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 Venipuncture technique
• Special precautions to be followed regarding the
collection are :
– Do not apply the tourniquet too tightly or for too long
a period

– Do not collect the blood from an arm into which an

intravenous (IV) infusion is being given.

– If an anticoagulated specimen is required, add the

correct amount of blood to the tube or bottle and mix
the blood with the anticoagulant by gently inverting
the container several times. Do not shake the
specimen.

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 Precaution considered during
specimen collection

During collection of blood specimens, the listed

factors can affect the correctness of test results
:
 Incorrect venipuncture technique.
 Haemolysis of red cells
 Collection of specimen into the wrong
container
 Instability of some chemical substances in
whole blood, serum, and plasma

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 Changes in composition of serum when
venous occlusion is prolonged from 1 min to
3 min
Increase % Decrease %
Total protein 4.9 Potassium 6.2
Iron 6.7
Total lipids 4.7
Cholesterol 5.1
AST 9.3
Bilirubin 8.4

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 Common specimen factors affecting
test results
Hemolysis and lipemia may be considered
as common factors affecting test result
hemolysis
 Haemolysis causes physiological and
chemical interference to the tests.

 Serum shows visual evidence of hemolysis

when hemoglobin conc exceeds 20 mg/dl.

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 Haemolysis can be avoided by

 Checking that the syringe and needle are dry and

that the barrel and plunger of the syringe fit well.
 Not using a needle with too fine a bore.
 Not withdrawing the blood too rapidly or moving
the needle once it is in the vein. Frothing of the
blood must be avoided.
 Removing the needle from the syringe before
dispensing the blood into the specimen container.
Allow the blood to run gently down the inside wall
of the container.

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 Lipemic specimen

• Milky white color – results from collecting blood

from the patient too soon after a meal

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 Specimen Collection precautions
• Adding the correct amount of blood to
anticoagulant. Do not shake the blood but gently
mix it with the anticoagulant.

• Using clean dry glass tubes or bottles

• Centrifuging blood samples for a minimum

period of time.

• Not storing whole blood samples in, or next to,

the freezing compartment of a refrigerator.

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 Stability of chemical substances in
blood specimens
• More concentrated in cells:
potassium, phosphate, AST, LDH

• More concentrated in plasma:

sodium, chloride, carbon dioxide

• Concentration about equal in cells and

plasma:
creatinine, glucose, urea, urate
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Some of the changes which may occur in
blood specimens within a few hours of being
collected include:
 Diffusion of potassium and some enzymes
 Diffusion of carbon dioxide off the surface of the
blood
 Decrease in the concentration of glucose by
glycolysis (at a rate of about 10%/hour
 Reduction or loss in activity of certain enzymes,
for example acid phosphatase
 Decomposition of bilirubin in daylight or
fluorescent light.
 Decrease in volume by evaporation (if not
stoppered)
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 Preservation methods

 Separating the plasma or serum from the red cells

as soon as possible

 Do not refrigerate whole blood before the serum

or plasma is separated.

 Use antiglycolytic agents to prevent glycolysis

 Protect specimen from light.

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If analysis is to be delayed or the
specimen has to be transported:
Separate from cells
Refrigerate or cool
Protect from light
Preserve with appropriate chemical
Stopper tubes
BEST OPTION : Test FRESH Specimen as
soon as possible after collection and
processing.
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 Anticoagulants and preservatives
for blood
 Used when whole blood or plasma is
desired for testing.
 Anticoagulants: chemical substances
added to fresh blood to prevent clotting.
 The right type of an anticoagulant in
correct amounts should be used.

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Precautions in the selection and use of
anticoagulants:
 It should neither remove nor add the substance
to be measured;

 Use neither excess nor less amount of an

anticoagulant.

 It should not be an enzyme inhibitor if sample is

to be used for enzyme assays.

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 Common Anticoagulants
• Heparin
• Sodium fluoride
• EDTA
• Oxalates,
• Iodoacetate
• Citrate, Na
• Review question: What color coding corresponds
with each of these?

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 Difference in composition of
plasma and serum
Plasma value greater than serum value (%):
• Calcium………. 0.9 LDH……………. 4.0
• Chloride……… 0.2
Plasma value less than serum value(%):
• Albumin……… 1.3 ALP………… 1.6 Uric acid……. 0.2
• AST…………….. 0.9 Bicarbonate… 1.8
• CK ……………… 2.1 Glucose………. 5.1
• Phosphorus.. 7.0 Potassium….. 8.4
• Sodium……… 0.1 Urea………….. 0.6
No difference in serum and plasma value:
Bilirubin Cholesterol Creatinine
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 Physiological factors affecting
composition of body fluids
Controllable variables: have short-lived
effects
• Posture
• Hospitalization and immobilization
• Exercise and physical training
• Circadian variation-
• Travel- across several time zones affects
normal circadian rhythm
• Food and stimulants:
• Underlying medical conditions: 28
 Specimen Variables

Uncontrollable variables- have long

term effects
Age
Gender
Race
Effect of environmental factors- altitude,
temperature, geographical location

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 Specimen transportation

• Primary containers should be capped tightly to:

• Prevent the escape of contents when the
container is exposed to extreme heat or
sunlight

Shipping materials
• Materials which is used to hold one or more
specimen tubes or bottles must be constructed
to prevent breakage

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 Precautions

• When necessary, remove stoppers carefully to

prevent aerosols

• Wear gloves

• Discard properly in a biohazard container

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 Summary

• Proper collection, preparation, processing

and preservation of the specimen is
essential for obtaining best results.

• Blood specimen depending on the required

purpose can be collected from Skin Puncture
Capillary blood), Arterial puncture, and Venous
puncture.

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 Summary, continued

• Specimen preservation techniques in

clinical chemistry includes, adding
antglycolysis chemicals, refrigeration and
prevent the specimen from direct sun
light.
• The types of anticoagulants used during
specimen collection for clinical chemistry
depends on the specific analyte required.
How ever serum sample is recommended
one, in most analytes measurements. 33
 Review Questions

• What are three biochemicals that require

the sample to be analyzed quickly to avoid
their disappearance?
• What are two common specimen errors
and their causes?

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 References

1. Burtis, Carl A., and Ashwood, Edward R..Tietz:

Fundamentals of Clinical Chemistry. WB
Saunders, Co., Philadelphia, 2001.
2. Arneson, W and J Brickell: Clinical Chemistry:
A Laboratory Perspective 1st ed. FA Davis, Co.,
2007
3. Burtis, Carl A., and Ashwood, Edward R. Tietz:
textbook of Clinical Chemistry. WB Saunders,
Co., Philadelphia, 1999.

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 Next Chapter

Chapter 7

Carbohydrate (Glucose analysis)

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