Haematology Watch, Vol.4 , Issue 1.
STUDENTS' CORNER
Activated Partial Thromboplastin Time
Mehmood
INTRODUCTION:
One of the important functions of human body is Haemostasis i.e. maintaining a fluid state of blood in normal conditions and securing bleeding points by localized plug made of platelets and fibrin.
This mechanism is composed of five components: coagulation factors, fibrinolysis, antifibrinolysis, platelets, and vessel wall. Coagulation factors help in making fibrin threads after activated by exposed collagen (Intrinsic pathway) and /or by expression of tissue thromboplastin (Extrinsic pathway). Either of the pathways culminate in conversion of Factor X, V, II, and I into activated forms leading to formation of Fibrin (Common pathway).
In a laboratory, assessment of function of coagulation factors is routinely done by Prothrombin Time test (assessing Extrinsic and Common pathway), and Activated Partial Thromboplastin Time test (assessing Intrinsic and Common pathway). These tests are helpful in investigating status of haemostatic system in an acutely bleeding patient, a person with suspected bleeding tendency or as a precaution before an invasive procedure.
PRINCIPLE:
Activated Partial Thromboplastin Time (APTT) is the time taken by citrated plasma to clot after adding cephalin and calcium chloride, and correlates with an overall function of Intrinsic and Common pathway of coagulation cascade. When cephalin is added to plasma, intrinsic pathway of coagulation is stimulated which, in the presence of added Calcium chloride, culminates in the formation of Fibrin, visible as an off-white clot in test tube. The time elapsed between adding the reagent and just appearance of the clot is noted, and termed APTT.
APTT test measures activity of intrinsic pathway and common pathway of coagulation cascade.
Compared to normal control plasma, the APTT test is prolonged in deficiency of coagulation factors VIII, IX, XI, XII, X, V, II, I. If a coagulation factor of Common Pathway is deficient, PT test is also prolonged.
REQUIREMENTS:
EQUIPMENTS:
Waterbath, Glass Test tubes, Stop watch, Micropipette with tips, Tube rack, Gloves.
REAGENTS:
Platelet-poor plasma of patient and of control, Cephalin, Calcium chloride.
PROCEDURE:
1. Deliver 100 µL plasma of patient into a glass test tube placed in waterbath at 37° C.
2. Start stop watch.
3. After 2 minutes, add 100 µL Cephalin.
4. Start stop watch.
5. After 3 - 5 minutes, add 100 µL Calcium chloride.
6. Make a layer of plasma by tilting the tube near horizontal, avoiding losing any sample, and dip the tube in water until stop watch reads ~5 seconds below control value (e.g. if control is 30 sec, start noting end-point at 25 seconds).
7. By holding the top of the test tube, bring it out of the rack and dip its bottom in the water gently in a to-and-fro motion at a rate of ~1 dip/sec.
8. Carefully detect the end-point i.e. appearance of off-white jelly-like clot; the later it forms, the thinner it appears. Do not look at the stop watch while observing the end-point detection.
9. When clot is formed (i.e. End-point detected), without looking at the watch, press the stop button immediately, and record the time.
10. Run the test in duplicate, and take a mean to report the APTt test result.
RESULT:
The result of the mean of the duplicate tests is expressed in seconds compared to a normal value. Normal range is a test value falling in ±5 seconds compared to the normal control value. For example, if normal value is 30 seconds, 25 – 35 is normal range; an APTT test of 26, or 31, or 34 is normal in this case, while a value of 24 or 36 is abnormal.
INTERPRETATION:
Normal APTT means that the value of a test plasma is declared normal when it will be equal to or within ± 5 seconds the value of APTT performed on Control plasma.
Compared to normal
control plasma, the APTT test is prolonged in deficiency of coagulation factors
VIII, IX, XI, X, V, II, I, which can be due to:
- Unfractionated heparin
- Liver disease
- DIC
- Haemophilias
- Dilutional coagulopathy
- Oral anticoagulant.
USES:
1. As a screening test of coagulopathy
2. As a monitor for Unfractionated heparin therapy
3. As a tool for detecting inhibitor of coagulation factors e.g. F-VIII and F-IX
4. As a tool for factor assays e.g. F-VIII and F-IX
Note: Compared to Prothrombin time, there is no INR in APTT test.