CHAPTER III

ADDITIVES

Learning objectives
At the end of this chapter, the student shall be able to:

 Define anticoagulants
 Describe the proportion, mechanism ofanticoagulation and advantages of EDTA,
Trisodium citrate, double oxalates and heparinanticoagulants.
 Prepare the different anticoagulants in the rightconcentration
 Site other additives used in the laboratory

INTRODUCTION
Additives are chemical preparations added to various tubes in the laboratory. Different
phlebotomy tubes are identified by the color of the stopper or shield. Specific colors indicate a
presence of certain additives. They are categorized according to its use and additive present.
Correct choice, proportion, mixture and use of tube ensure accuracy of the results
generated.

3.1 Major Additives:

a. Antiglycolytic agent- additives that inhibit the use of glucose by blood cells. Such inhibition
may be necessary if testing of blood glucose level is delayed.
Example: Sodium fluoride and Lithium iodoacetate containing
Along with antiglycolytic agents, anticoagulant is also added to the tubessuch as
potassium oxalate (K2EDTA) to yield plasma for more rapid testing

b. Anticoagulant – additives that prevent blood from clotting

Example:

 EDTA (Ethylene diaminetetraacetic acid) – prevents coagulation by chelating calcium

Usage: 1.5 mg/mL of blood; Within 2 hours cellular morphology is preserved
o K2 (DIPOTASSIUM SALT) – Sequestrene(Recommended as it is in powdered
form, thereby preventing possible hemodilution)
o K3
o Na2 (DISODIUM SALT) – Versene

Important reminders in using EDTA: WBC counts, Platelet and ESR can be
determined up to 24 hours in an EDTA containing sample if it is refrigerated at 4 oC

 Heparin – prevents coagulation by acting as anti-thromboplastin and antithrombin

Usage: 15-20 U/mL of blood

Heparin is an excellent natural anticoagulant extracted from mammalian liver or

pancreas. It is more expensive than the artificial ones and has a temporary effect of only 24
hours. Heparin prevents clotting by inactivating thrombin, thus preventing conversion of
fibrinogen to fibrin. It is the best anticoagulant when absolute minimal hemolysis is required
(e.g., osmotic fragility test and hematocrit determination).

Important reminders: Blood preserved with heparin is not ideal for blood smear
preparation using Wright’s and Leishman’s stain as it provides bluish discoloration of the
background

 Oxalates – prevents coagulation by combining with calcium to form an insoluble calcium

oxalate salts
o Dried potassium oxalates – shrinks rbc; not ideal for ESR hematocrit, blood
smear and blood potassium determination
o Dried ammonium and potassium oxalate – balanced oxalates used for ESR

 Citrates – prevents coagulation by combining with calcium in a non-ionized form

o Used for transfusion (ACD solution)
o 3.2% sodium citrate
 1:4 anticoagulant to blood ratio – used for ESR determination Westergren
method
 1:9 anticoagulant to blood ratio – used for coagulation studies
Name: __________________________ Score: ______ Instructor’s Signature: _________
Review Questions

1. Define anticoagulant

2. List the anticoagulants that are commonly used in hematology. How does each of
these anticoagulants exert their functions? What tests/procedures use such tubes?

3. Write the proportion of the volume of blood to the volume of each of these
anticoagulants as well as the recommended number of inversions.

4. List the correct order of draw in a venipuncture procedure

5. Enumerate the miscellaneous tubes used in the laboratory as well as its usage,
additives present and recommended number of inversions