Anticoagulation

Whole blood

A venous, arterial or capillary blood sample in which the concentrations and properties of cellular and extra-cellular constituents remain relatively unaltered when compared with their in-vivo state.

 Anticoagulation in-vitro stabilizes the constituents in a whole blood sample for a certain period of time.

Plasma

 The virtually cell-free supernatant of blood containing anticoagulant obtained after centrifugation.

Serum

 The undiluted, extracellular portion of blood after adequate coagulation is complete.

CLOTTING MECHANISM
ANTICOAGULANTS

Anticoagulants prevent blood from clotting. They are added to the blood sample, especially 

when blood is collected by venipuncture, and sent to the laboratories for investigation.

They are additives that inhibit blood and/or plasma from clotting (ensuring that the constituent to be measured is non-significantly changed prior to the analytical process). 

Anticoagulation occurs by binding calcium ions (EDTA, citrate) or by inhibiting thrombin activity (heparinates, hirudin).

EDTA is usually used in ‘hematology’ while citrated blood is used for ‘coagulation studies’and in blood banks.

The following solid or liquid anticoagulants are mixed with blood immediately after sample collection:

A. EDTA,

B. Trisodium citrate,

C. Double oxalate,

D. Sodium fluoride and

E. Heparin.

A. EDTA (Ethylenediamine Tetra-acetic Acid)

 This is also known as Sequestrene or Versene. The sodium and potassium salts of EDTA arepowerful anticoagulants.

 Preparation

 Prepare a 10 % solution of dipotassium salts of EDTA.

 Dissolve 10 g of salt in about 80 ml of water in a 100 ml volumetric flask and then makethe volume of the solution to 100 ml.

 Mechanism of Action

EDTA acts by its chelating effect on the calcium molecules of the blood. Calcium is one ofthe important factors required in the coagulation process.

A . Effective Concentration

To achieve the chelating effect, a concentration of 1.2 mg of the anhydrous salt per ml ofblood (1.2 mg/mL) is required (4.1 mMol/L).

 Uses

 EDTA is suitable for all routine hematological investigations except coagulationstudies.

B. Citrate

 Trisodium citrate (32 g/l, Na,C,H,O,-2H,O) is the anticoagulant of choice in coagulationstudies.

Uses

It is used for coagulation studies, including prothrombin times and partial thromboplastintests, in blood banks, in the estimation of ESR, especially by the Westergren method.

C. Double Oxalate

 This is an anticoagulant containing ammonium oxalate and potassium oxalate. Therefore, itis called double oxalate.

 Potassium oxalate alone causes shrinkage of red cells whereas ammonium oxalate increases

their volume. So, double oxalate is also called balanced oxalate as it preserves cell morphology.

 Uses

This is used for estimation of ESR, PCV or investigations in which the volume of the cells

should not be affected.

D. Heparin

Heparin is, theoretically, the best anticoagulant because it is a natural constituent of blood and

introduces no foreign contaminants into the blood specimen.

Uses

 It is used for blood gas determination and pH assays.

 It is the best anticoagulant for the osmotic fragility test.

E. Hirudin

 Hirudin is an antithrombin extracted from leeches or prepared by a genetic engineeringprocess.

 Hirudin inhibits thrombin by forming a 1:1 hirudin-thrombin complex.

 Hirudin is used at a concentration of 10 mg/L.

The colour codes of anticoagulants described in ISO/DIS 6710 are:

EDTA = lavender/red;

citrate 9 + 1 = light blue/green;

citrate 4 + 1 = black/mauve;

heparinate = green/orange;

no additives (for serum) =red/white.