Anticoagulants are the chemicals which prevent clotting of blood when mixed with blood in proper proportion (Willard et al; 1999). These chemicals, apart from hepian works by binding calcium ions preventing coagulation proteins from using them. Heparin interferes with the formation and/or activity of thrombin and the activity of clothing factor 1X, X, XI, XII (Hylek et al; 2007).
Characteristics of anticoagulants;
An anticoagulant selected for use in hematological examination must have the following qualities;
1. It must not alter the size of red cells
2. It must not cause haemolysis
3. It must minimize platelet aggregation
4. It must minimize disruption of the staining and morphology of leukocytes
5. It must be readily soluble in blood. (Schalm et al; 2006).
The Frequently used anticoagulants includes;
1. Ethylenediaminetetra acetic acid (EDTA)
2. Oxalate
3. Heparin
4. Sodium citrate
5. Sodium fluoride/potassium oxalate
6. Citrate phosphate dextrose and adenine (CPD-A).
7. Fluoride oxalate (FO)
8. Acid citrate dextrose (ACD)
Ethylenediaminetetra acetic acid, heparin, fluoride oxalate and sodium citrate are the ones will be used for the study and hence they are explained below;
Ethylenediaminetetra Acetic Acid (EDTA)
EDTA is a widely used acronym for the chemical compound Ethylenediamintetra acetic acid. EDTA is a polyamino carboxylic acid with the formular (CH2N) CH2 CO2H2)2. The colorless, water soluble solid is widely used to dissolved scale. Its usefulness arises because of its role as a chelating agents, that is its ability to “sequester” metal ions such as Ca2+ Fe3+. After being bound by EDTA metal ions remain in solution but exhibit diminished reactivity. EDTA is produced as several salts; notably disodium EDTA and calciumdisodium.
EDTA (Vitalongevity 2007). (Jain 2006) said that EDTA consist or comes in 2 forms;
a. The tripotassium salt (K3 EDTA)
b. And the disodium salt (Na2 EDTA). He also added that the potassium salts (Liquid or dry powder) are used in commercial tubes because they are more soluble. (Calcium EDTA is not used as an anticoagulant, but in the treatment of lead poisoning).
Synthesis of EDTA;
The compound was first described in 1935 by Ferdinard Munz, who prepared the compound from ethylenediamine and chloroacetic acid (Vitalongevity 2007). Today, EDTA is mainly synthesized from ethylenediamine (1,2-diamino ethane), formaldehyde (Methanal), and sodium salt which can be converted in a subsequent step in the acid forms;
H2NCH2CH2NH2+4CH2O+4NaCN+4H2O®(NaO2CCH2)2NCH2 CH2N (CH2CO2Na) 2 + 4NH3
(NaO2CH2 NCH2)2 NCH2 CH2N (CH2 CO2Na)2 4HCI® (HO2CCH2) 2 NCH2 CH2N(CH2CO2H2+4NaCL.
Impurities cogenerated by this route include glycine and niterilotriacetic acid (New York Times; 2008).
Mode of Action
It acts as a powerful calcium chelating agent. The calcium in blood is bound in an unionzed and soluble complex with EDTA
Tests which are performed by using EDTA blood are:
1. Haemoglobin
2. White blood count
3. Packed cell volume determination
4. Erythrocyte sedimentation rate by wintrobe’s method
5. Platelet count
6. Differential white blood cell count
7.
Advantages
1. It gives the best preservation of cellular morphology. Good morphology of the cell is observed even after 2 to 3 hours of blood collection
2. Since platelet clumping is inhibited for platelet counts using this anticoagulant is preferred (Willard et al 1999)
Disadvantages;
1. Excess EDTA causes shrinkage of red blood cells and erroneous packed cell volume, mean cell volume and means corpuscular haemoglobin (Jain 2006).
2. Platelets swell and disintegrate due to excess of EDTA and artificially high platelet count may be obtained due to disintegrated platelets.
3. EDTA is not suitable for use in the coagulation studies mainly in the determination of prothrombin time
4. EDTA 2k (potassium salt is recommended as anticoagulant for complete blood count. (Willard et al 1999)
Mechanism of Action of Anticoagulants
The thromboplastin released by damaged tissue, or platelets converted inactive prothrombin into active thrombin in the presence of calcium ions. Thrombin coverts soluble fibrinogen into insoluble fibrin clot in the presence of calcium ions.
Thromboplastin
Prothrombin ——————-> Thrombin
Ca++
Thrombin
Fibrinogen(soluble) ——————-> Fibrin (insoluble)
Ca++
fibrin + blood cells clot (fine threads)
Some anticoagulants such as potassium oxalate prevent clotting of blood by precipitating ionic calcium in plasma while anticoagulant such as sodium citrate ions into unionized form. Since calcium ions are not available, blood is prevented from clotting. Heparin on the other hand acts as antithrombin and thus prevents the formation of thrombin and blood clotting (Willard et al; 1999
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