1. Laboratory Evaluation
Laboratory tests play a very important role in the diagnosis, monitoring, and treatment of
coagulation disorders. When used in the appropriate manner, they can yield a great deal of
information and can often localize a disease process to a certain part of the cascade.
Furthermore, these tests are extremely helpful in the monitoring of anticoagulant therapy.
However, in order to realize their true utility and limitations, one must first understand
exactly how they are performed. With this knowledge, a clinician can then form an adequate
interpretation. Indeed, the tests by far reachtheir highest specificity and senstivity only when
the clinical picture is known. The most common laboratory evaluations used are the activated
prothrombin time (aPT) and activated partial thromboplastin time (aPTT). Other tests such as
thrombin time and bleeding time are also extremely valuable, but often underused.
Activated Prothrombin Time
Activated Partial Thromboplastic Time
Thrombin Time
Bleeding Time
Fibrinogen Concentrations
Assays for Lupus Anticoagulants
Factor VIII Inhibitor Assay
vonWillebrand Disease Assays

2. Prothrombin Time**
Purpose
Screening test to indentify acquired or inherited deficiencies of factors VII, X, V,PT , FIB .
Monitor oral anticoagulant therapy e’ warfarin, which decreases the activity of VII, IX, X , PT.
Method
Collect blood from the patient into a citrated tube
Take the collected blood to the laboratory
Centrifuge to isolate the plasma
Take small sample of the plasma ~0.1 ml.
Add calcium
Add Thromboplastins - which are preparations of TF and phospholipids from rabbit brain .
The time that elapses until clot forms is the activated Prothrombin Time (PT).
Interpretation
PT shorter than the reference range is not associated with any clinical condition and is
usually related to improper procedure.
Prolongation of thePT occurs when there is:
- Increased hematocrit, leading to a false +ve in PT
prolongation.
- Liver disease. - Vitamin K deficiency
- DIC. - Nephrotic Syndrome
- Treatment e’ antibiotics, chemotherapeutics, or AT drugs.
Note: The PT will increase shortly after a bolus dose of heparin.
The monitor of anticoagulant therapy requires standardization of the PT between different
laboratories in order to ensure that the degree of anticoagulation measured in one
laboratory can be compared to that in another laboratory. Therefore, the INR was
created. The INR takes into account variations in reagents and techniques between different
labs and provides one value that can be compared between labs.

3. Partial Thromboplastin Time
Purpose
Screening test to indentify acquired or inherited deficiencies of Factors IX, VIII, and XI.
Screening test for lupus anticoagulants
Monitor heparin anticoagulation
Screening test to assess reduction in the activity of FIB , V , X.
However, the PT is more sensitive for these.
Method
Collect blood from the patient into a citrated tube
Take the collected blood to the laboratory
Centrifuge to isolate the plasma
Take a small sample of the plasma ~0.1 ml
Add calcium (catalyst for coagulation)
Add partial thromboplastin -phospholipid source without tissue factor
Add a negatively charged surface such as kaolin, silica, or dextran So4 to activate coagulation
The time that elapses until clot forms is the activated Partial Thromboplastin Time (aPTT).
Interpretation
aPTT shorter than the reference range is usually the result of poor sample handling.
Prolongation of the aPTT occurs when there is:
- Factor Deficiency (see below) - Factor Inhibitor (see below)
- Anticoagulation with heparin - Contamination of sample with heparin
Note: Once heparin is started, the laboratory work-up of an abnormal aPTT is difficult.
Low molecular weight heparin(LMWH) and heparinoids generally do not prolong the aPTT.

4. When attempting to determine the cause of a prolonged aPTT, one must first rule out
the most common cause which is contamination of the blood specimen with heparin.
This usually occurs when small amounts of heparin are used to keep venous or arterial
catheters patent. If this is ruled out and the patient is not on intravenous heparin for
anticoagulation, then one can begin to suspect either a factor deficiency or a factor
inhibitor. A factor inhibitor is usually an antibody targeted against a specific
coagulation factor, which results in activation of that factor. Examples of this are the
lupus anticoagulant and Factor VIII inhibitors. The distinction of inhibitor versus
factor deficiency can be made by performing a 1:1 mix of the patient's plasma with
plasma pooled from normal individuals. (Pooled plasma is plasma taken from a large
number of individuals with no known coagulation defects.) In theory, it should
contain normal levels of all coagulation factors. In a patient with factor deficiency,
aPTT measured after mixing the patient's and pooled plasma should be normal. In
patients with factor inhibitor, mixing the pooled plasma and the patient's plasma will
still yield a prolonged aPTT because the inhibitor in the patient's plasma will also
inactivate the specific factor in the pooled plasma.
Thrombin Time
Purpose
Screening test for hypofibrinogenemia, hyperfibrinogenemia, Dysfibrinogen , and inhibitors
against thrombin or fibrin.
Tests the conversion of fibrinogen to fibrin to cross-linked fibrin.
Monitor anticoagulant therapy with fibrinolytic agents and hirudin..
The test can be used to measure fibringoen levels.
Method
Collect blood from the patient into a citrated tube
Take the collected blood to the laboratory
Centrifuge to isolate the plasma
Take small sample of the plasma ~0.1 ml.

5. Add bovine thrombin to plasma
The time that elapses until clot forms is the Thrombin Time (TT).
Interpretation
TT shorter than the reference range is not associated with any clinical condition is usually
related to improper procedure.
Prolongation of the TT occurs when there is:
1. Contamination with heparin.
2. TT can be repeated after addition of protamine sulfate and will be normal..
3. Afibrinogenemia/Hypofibrinogenemia –
4. Acquired (DIC, liver disease) and familial
5. Interference with fibrinopeptide cleavage

6. Bleeding Time
Purpose
Screening test for congenital and acquired disorders of platelet function.
Screening test for vonWillebrand's disease.
Method
A spring loaded device with a blade is used to make a small incision in the skin.
The timer is started.
Filter paper discs are used to adsorb droplets of blood at regular intervals.
The time required for bleeding to cease is the Bleeding Time.
Interpretation
Prolonged Bleeding Times are usually due to:
- Aspirin and other drugs that interfere with platelet function.
- Congenital or acquired disorders of platelet function.
- vonWillebrand Disease - can further work-up by measuring PTT, Factor VIII activity, Factor
VIII antigen, and Ristocetin cofactor activity.
Fibrinogen Concentration
Purpose
The Thrombin Time is used to screen for reduction in fibrinogen concentration.
The Fibrinogen Titer Test can also be used. (Discussed below.)
Methods
Thrombin Time (Click to go to Thrombin Time section)
Fibrinogen Titer Test
1. Thrombin is added to serial dilutions of plasma in buffered saline.
2. The dilutions in which clot formation is visible are noted.
Interpretation
Clot should normally be visible at a dilution of 1:64, and is usually seen at 1:128.
Visible clot in these dilution rules out a clinically significant quantitative abnormality in
fibrinogen and estimates the concentration to be greater than 100 mg/dL.