SPECIAL EVALUATION OF

COAGULATION DISORDERS
Thrombin Clotting Time
Measures the conversion of fibrinogen to fibrin
May be used to monitor heparin therapy.

Principle:
Commercially prepared thrombin reagent is
added to citrated plasma, and the time required
for clot formation is measured.
Normal Values: 10-20 seconds
Thrombin Time
• Normally prolonged in newborn and in multiple
myoma.

Prolonged Thrombin time :

Fibrinogen level is below 75 to 100 mg/dl
Function of fibrinogen is impaired
In the presence of heparin, fibrin/ogen ,
degradation products , thrombolytic agent
Thrombin Clotting Time
Things to remember :
TT should not be performed on heparinized
neutralizing substance that have been frozen .
Thrombin is highly unstable , to preserve its
activity it is diluted to 100 NIH units/ml with equal
parts 0.15 M NaCl and glycerol.
Plastic or siliconized glass should be used .
Reptilase Time
• Reptilase is an enzyme found in the venom of the
Bothrops atrox snake.
• Capable of converting fibrinogen to fibrin and its
unaffected by heparin.
• Helpful in testing for functional fibrinogen when
thrombin time is prolonged because of heparin.
Reptilase Time
• Principle :
• Reptilase is added to plasma, it acts by releasing
fibrinopeptide A from the fibrinogen molecule. The
resultant monomers polymerize end to end, forming
clot .

• Reference range : 18-20 seconds

 Comparison of Thrombin Time and Reptilase Time in Various
Condition
Thrombin Time Reptilase Time

Heparin Therapy Prolonged Normal

Fibrin Split Products Greatly Prolonged Prolonged

Hypofibrinogenemia Prolonged Prolonged

Dysfibrogenemia Prolonged Greatly Prolonged

Immunologic Prolonged Normal

Antithrombin
Ristocetin Cofactor Assay

• Ristocetin cofactor (VWF:RCo) has been the most

widely used method for measuring the activity of
VWF.
• Ristocetin cofactor activity a property of the
plasma von Willebrand factor, which is responsible
for vitro platelet agglutination in the presence of
ristocetin
Ristocetin Cofactor Assay
Decreased amounts or abnormalities
associated with von Willebrand syndrome

Reference range : 60-180 %

Specimen : Citrated plasma , plasma maybe
refrigerated for up to 8 hours
Ristocetin Cofactor Assay
Principle:
measures the ability of a patient’s plasma to
agglutinate formalin-fixed platelets in the
presence of ristocetin.
The rate of ristocetin induced agglutination is
related to the concentration of von Willebrand
factor and the percent normal activity can be
obtained from the aggregometer tracing.
Mixing Study
Mixing Study / Subtitution Tests
- this can be adopted if primary tests like PT or APTT are
abnormally prolonged and the indicate a factor
deficiency. The patient’s deficient plasma is diluted 1:1
with a plasma or serum substitute and the APTT or PT is
repeated.A correction if the original prolonged APTT or
PT indicates that the deficient factor has been added to
the patient’s plasma by substitution solutions as follow:
1. Aged plasma- lacks labile factors V & VIII but
retains normal activity of all coagulation factors.
2. Fresh adsorbed plasma- lacks vitamin K
dependent factors (II, VII, IX, X) but retains activity of al
other coagulation factors.
3. Aged serum- lack factors I, II, V, VII but retains
normal activity of all coagulation factors.
 FACTOR DEFICIENT PLASMA OR SERUM

Extrinsic Intrinsic Pathway Normal Plasma Adsorbed Aged Serum

Pathway (PT) (APTT) Plasma

I I + + (-)

II II + (-) (-)

V V + + (-)

VII + (-) +

VIII + + (-)

IX + (-) +

X X + (-) +

XI + + +

XII + + +