Chapter 6

Specimen collection, processing, &

preservation
 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.
 Outline of Specimen collection, processing, &
preservation

• Specimen for biochemical analysis

• Specimen types and collection
• Coding of stopper colors
• Skin puncture
• Arterial puncture
• Venipuncture
• Common specimen factors affecting test
results
 Outline of Specimen collection, processing, &
preservation, continued..

• Stability of chemical substances in blood

specimen
• Preservation methods
• Anticoagulants and preservatives
• Common anticoagulnts
• Physiological factors affecting composition of
body fluids
• Specimen transportation
 Specimen for Biochemical analysis
• 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.

• 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.
 Specimen types and collection
• Majority of clinical chemistry tests are done
on blood : whole blood, serum or plasma.
• Blood for analysis may be obtained from
veins, arteries or capillaries.
• Specimen Types
-Random specimen
-Fasting specimen
-Timed specimen
 Coding of stopper color to indicate
additive in evacuated blood tubes
COLOR Use Additive
Gray plasma or whole oxalate(Na,K)
blood with glycolysis fluoride(Na),
inhibition 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 (Na2, K2)
Colors of Tubes
 Specimen Types
• Skin Puncture
– Capillary blood
• Arterial Blood
• Venous Blood
 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.
 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
 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.
 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.
 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
 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
 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.
 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.
 Lipemic specimen

• Milky white color – results from collecting blood

from the patient too soon after a meal
 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.
 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
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)
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.

 BEST OPTION : Test FRESH Specimen as soon as
possible after collection and processing.

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
 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.
 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.
 Common Anticoagulants

• Heparin
• Sodium fluoride
• EDTA
• Oxalates,
• Iodoacetate
• Citrate, Na
• Review question: What color coding corresponds with each of
these?
 Difference in composition of plasma and serum
Plasma value greater than serum value (%):
• Calcium………. 0.9
• Chloride……… 0.2
• LDH……………. 4.0

Plasma value less than serum value(%):

• Albumin……… 1.3 ALP………… 1.6
• AST…………….. 0.9 Bicarbonate… 1.8
• CK ……………… 2.1 Glucose………. 5.1
• Phosphorus.. 7.0 Potassium….. 8.4
• Sodium……… 0.1 Urea………….. 0.6
• Uric acid……. 0.2

No difference in serum and plasma value:

Bilirubin Cholesterol Creatinine
 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:
Uncontrollable variables- have long term
effects
 Age
 Gender
 Race
 Effect of environmental factors- altitude,
temperature, geographical location
 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
 Precautions

• When necessary, remove stoppers carefully to

prevent aerosols

• Wear gloves

• Discard properly in a biohazard container

 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.
• Common factors affecting test results are Hemolysis and
lipemia.
• If the blood sample is not diagnosed immediately,anlytes such
as potassium, billiribun, and glucose concentartion may be
affected
• Specimen preservation techniques in clinical chemistry
includes, adding antglycolysis chemicals, refrigeration and
prevent the specimen from direct sun light.
 Summary, continued

• 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.
 Reference
1. Burtis, Carl A., and Ashwood, Edward R.. Tietz:
Fundamentals of Clinical Chemistry. Philadelphia, 2001.
2. Arneson, W and J Brickell: Clinical Chemistry: A
Laboratory Perspective 1st ed. 2007 FA Davis

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

textbook of Clinical Chemistry. Philadelphia, 1999.
 Next Chapter

Chapter 7

Carbohydrate (Glucose analysis)