CLINICAL HEMATOLOGY 2 (HEMA312) LABORATORY

SCREENING TESTS: BLEEDING TIME AND CLOTTING TIME

GENERAL OVERVIEW
• Bleeding time and clotting time are procedures performed used to assess both quantitative and
qualitative properties of thrombocytes to help in the screening of possibly underlying conditions
• It is often requested for patients undergoing surgery, with bleeding tendencies, etc.

BLEEDING TIME

PRINCIPLE OF THE TEST

• Bleeding time is a test performed to evaluate platelet function by allowing the incision or bleeding to
stop within a given time reference. Involves making a puncture wound in a superficial skin.
o Advantages: used for pre-operative assessment of patients taking aspirin or NSAID’s
o Disadvantages: incentive and lacks reproducibility

METHODS OF BLEEDING TIME

1. Duke’s Method
▪ Most used procedure, easy to perform
▪ Materials: lancet, filter paper
▪ Procedure:
A. Make a single incision on either earlobes or finger
B. Timer starts as soon as bleeding begins and blot to filter paper immediately
C. Check evidence of bleeding every 30 seconds until no blood is seen
D. Report the bleeding time that is closest minute
▪ Reference:
2. Ivy’s method
▪ Most recommended procedure
▪ Materials: lancet, filter paper, sphygmomanometer
▪ Requires standardized pressure of 40mmHg and incision of 1mm deep and 3mm wide
▪ Procedure:
A. Three incisions are made on the volar surface of the arm with pressure
B. Timer starts as soon as bleeding begins and blot to filter paper immediately
C. Check evidence of bleeding every 30 seconds until no blood is seen
D. Remove the blood pressure cuff
E. Report the average bleeding time
▪ Reference:

HEMA312 LABORATORY DISCUSSION | OLFU-CMLS 1

 3. Copley and Lalitch method
▪ Materials: lancet, sterile saline
▪ Requires warming the puncture site using sterile saline at 37C
▪ Procedure:
A. Make a single incision with a depth of 6mm on finger and start the timer
B. Immerse the puncture site in a sterile physiologic saline solution at 37C
C. Wait until bleeding stops and no blood flow is observed
▪ Reference:
4. Standard template method
▪ Similar procedure with Ivy’s method
▪ Materials: filter paper, glass, or plastic template (lancet), sphygmomanometer
▪ Requires single incision of 11mm deep and 11mm wide

VARIABLES AFFECTING BLEEDING TIME

• Prolonged bleeding time
o Physiologic errors: aspirin intake, cephalothin intake
o Pathologic cause: anemia, thrombocytopenia, Bernard Soulier syndrome, fibrinogen
deficiencies, vascular disorders, von Willebrand disease, disseminated intravascular
coagulation, Glanzmann’s thrombasthenia
• Shortened bleeding time
o Physiologic errors: smaller incision, low pressure

HEMA312 LABORATORY DISCUSSION | OLFU-CMLS 2

 CLOTTING TIME

PRINCIPLE OF THE TEST

• Clotting time is a test performed to determine the time required for clot to form. Involves the activity
of coagulation factor and presence of a negative charge surface for its activation.
• The formation of clot or fibrin demands the presence of thrombin who is responsible for the
conversion fibrinogen to become fibrin then stabilized by transglutaminase.

METHODS OF CLOTTING TIME

1. Slide Method
▪ Most used procedure, easy to perform
▪ Materials: lancet, glass slides
▪ Procedure:
A. Perform capillary puncture. Make sure that the finger is warmed prior to puncture.
Incision should be 3mm deep. Start the timer as soon as the blood appears and don’t
forget to wipe to first drop of blood to prevent contamination of tissue juices.
B. Place a drop of blood on slide for the determination of clotting time.
C. Check the drop of blood every 30 seconds until the presence of clot or fibrin clot is
first observed and seen.
D. Report the time where fibrin is first seen.
▪ Reference:
2. Capillary Method
▪ Formation of fibrin is observed in non-anticoagulated capillary tubes by breaking it
▪ Materials: capillary puncture kit, whole blood sample
▪ Procedure:
A. Perform capillary puncture and fill up the capillary tube up to ¾ amount. Make sure
that the finger is warmed prior to puncture. Incision should be 3mm deep.
B. Start the timer as soon as the blood appears and don’t forget to wipe to first drop of
blood to prevent contamination of tissue juices. Wait for 2 minutes for blood to clot.
C. After 2 minutes, break off the capillary tube from 1-2cm from end.
D. In the absence of clot, wait for another 30 seconds and break the tube again. Repeat
the step until a fibrin strand appears.
E. Report the time where fibrin is first seen.
▪ Reference:

HEMA312 LABORATORY DISCUSSION | OLFU-CMLS 3

 3. Tube Method
▪ Also known as the whole blood clotting time; measures all stages of intrinsic coagulation
▪ Materials: venipuncture kit, tube, water bath, whole blood sample
▪ Procedure:
A. Perform venipuncture and extract 4ml of blood and label 3 test tubes from #1 to #3.
B. Dispense 1ml of blood in tube #3, then in #2 and lastly in tube #1. Discard the
remaining blood in the syringe. Do not discard the blood in sink.
C. Timer should start as soon the blood interacts with glass tube and place them in a
water bath and incubate at 37C for 5 minutes.
D. After incubation, check any presence of clot in the tube #1 by tilting at 45° angle. If no
presence of clot, return the tube the water bath.
E. Check the presence of clot every 30 seconds. Start checking from tube #1, if clot is
present, move to tube #2 and until to tube #3. You cannot move to another test tube
in the absence of clot of the preliminary tube.
F. Once all tubes have clotted, report the time where clot is seen in tube #3.
▪ Reference:

VARIABLES AFFECTIVE CLOTTING TIME

• Prolonged clotting time
o Physiologic errors: insufficient blood, intake of heparin, intake of thrombin inhibitors
o Pathologic cause: coagulation factor deficiencies, presence of circulating anticoagulants
• Shortened clotting time
o Physiologic errors: hemolysis, vigorous agitation, pre-exposure to clot activators

HEMA312 LABORATORY DISCUSSION | OLFU-CMLS 4