Anticoagulants and Preservatives for

Blood, Plasma and Serum

Purpose of anticoagulants

These are used to prepare the whole blood or plasma during collection of blood sample.

In routine used anticoagulants are :

1. EDTA Ethylenediaminetetraacetic acid as disodium or potassium salts are used.

1. This is a chelating agent which binds the calcium which is needed for coagulation.

2. It is effective at a final concentration of 1 to 2 mg / mL of blood

3. More than 2 mg / mL causes shrinkage of the cells.

4. This can be used as powder or make the solution and then add to vials. Let it dry.

5. EDTA solution of 0.1 % can be prepared and used. Let it evaporate at room
temperature.

6. This is the best anticoagulant for peripheral blood smear and studies.

7. It has little effects on the various tests.

8. Drawbacks It inhibit the alkaline phosphatase, creatine kinase, and leucine

aminopeptidase activities.

9. EDTA is not suitable for Calcium and iron estimation.

2. Heparin This is quite common anticoagulant and cause least interference with test.

1. It is mucoitin polysulfuric acid available as sodium.potassium , lithium and

ammonium salts.

2. Heparin accelerate the action of antithrombin III > which neutralizes thrombin
> thus prevents the formation of > fibrin from fibrinogen.

3. Heparin is added 0.2 mg /mL of blood.

4. Drawback It inhibits the acid phosphatase activity.

1. It give blue background for wright;s stain smears.

 2. It also effects the binding of triiodothyronine and thyroxine to their carrier
protein and produce higher free concentration of these hormones.

3. It interfere with binding of calcium to EDTA.

3. Sodium Citrate This will chelate calcium .

1. This is used as 3.2 to 3.8 g/dL .

2. In blood its ratio is 1:9 where 9 parts are blood and 1 part is sodium citrate.

3. Drawbacks This is not good for estimation of calcium.

1. It inhibits aminotransferase and alkaline phosphatase.

2. This will stimulate acid phosphatase when phenyl phosphate is used as

substrate.

4. Oxalate This may be sodium, potassium, ammonium or lithium oxalic acid salt used as
anticoagulant.

1. This form insoluble complex with calcium ions.

2. Potassium oxalate at concentration of 1 to 2 mg/mL of blood is used. This is the most

popular oxalate salt used as anticoagulant.

3. Combination of ammonium/potassium oxalate does not leads to shrinkage of the

RBCs.

4. While other oxalate cause shrinkage.

5. Drawbacks If the concentration is >3 mg/mL, then there are chances for hemolysis.

1. There is reduction of 10 % of hematocrit.

2. Oxalates inhibits several enzymes like acid phosphatase, alkaline

phosphatase, amylase, LDH, and may cause the precipitation of calcium as
oxalate salt.

5. Sodium Fluoride This is a weak anticoagulant but used antiglycolytic agent to preserve the
glucose.

1. This inhibit the system involved in glycolysis and preserve the glucose.

2. This is effective at a concentration of 2 mg/mL of blood along with other

anticoagulant like oxalate.

3. When used alone then more concentration than 2 mg/mL is needed.

4. Most specimens are preserved at 25 C for 24 hours and at 4 C for 48 hours.

 5. Sodium fluoride is poorly soluble so mix blood thoroughly before effective anti
glycolysis occurs.

6. Drawback This is also inhibitor of many enzymes.

1. Also effect urease for the estimation of urea.

6. Sodium Iodoacetate This is also antiglycolytic agent at concentration of 2 g/L.

1. This may be substituted for sodium fluoride.

2. This has no effect on urease.

3. Drawback It inhibits creatine kinase.

Whole blood

This blood sample in obtained in the test tube containing anticoagulant.

This sample will contains cells and plasma.

Plasma

This is pale yellow liquid which contains RBcs, white cells, and platelets.

Plasma forms with the help of anticoagulant which will prevent the clotting..

There is presence of fibrinogen in the plasma.

Serum

This is clear fluid which separates from the clotted blood. There are no RBCs, white cells or
platelets. There is no need for the anticoagulants.

Clotted blood is kept at 37 C for at least 20 minutes and then centrifuged.

The upper portion is called serum.

There is no fibrinogen.

Buffy Coat

This is the middle layer between the plasma and RBCs.

This will contains white cells and platelets