suggests that the patient’s blood is hypercoagulable. 3. Vigorous agitation of the tubes will significantly shorten the coagulation time. So tipping should really be very gentle just to see if the blood has clotted.

 

¾ Filter paper

¾ Glass slide

¾ Alcohol sponges.

Method

1. Clean the lobe of the ear or tip of a finger with alcohol

and let dry.

2. For ear—glass slide is placed behind the ear lobe

and held firmly in place. This provides a firm site for

incision.

3. Pierce the lobe of the ear by a firm stroke against the

glass slide (or pierce the finger-tip). Discard the glass

slide if ear lobe has been incised. Start the stop watch

when the stab was made.

4. Bleeding of the wound should be allowed to proceed

without pressure and the blood is allowed to drop on

the filter paper. The paper should be moved so that

each drop will fall on a fresh area. When bleeding

slows, the wound is touched gently with a fresh area

of the filter paper at 30 second intervals. When blood

no longer stains the filter paper, the watch is stopped

and the time recorded.

Normal Values

The normal range is up to 6 minutes. Between 6 and 10

minutes, the results are borderline. Over 10 minutes is

definitely abnormal.

Precautions

1. In children, heel should be used.

2. In suspected cases of a bleeding disorder, the bleeding

may not be controlled easily from the ear lobe hence,

fingertip puncture wounds are better.

3. The area to be punctured should not be congested.

4. The size and depth of the wound may vary if one does

not have a standardized technique.

5. If bleeding persists for more than 15 minutes it should

be stopped by placing a dry gauge sponges over the site

and applying finger pressure (the filter paper used to

collect the drops of blood can be dried and saved as a

record of the procedure).

Ivys’s Method

(Preferred because of greater ease of standardization).

Method

1. Cleanse the inner aspect of the forearm with spirit and

let dry.

2. Place a blood pressure cuff on the upper arm, inflate

at 40 mm Hg, and maintain the same throughout the

test.

3. Select an area on the forearm—Volar aspect which is

devoid of superficial veins. Stretch the skin laterally

between the thumb and forefinger and hold in a taut

position.

4. Take a cork, through which a no. 11 surgical blade has

been inserted with the tip extending 3 mm beyond the

cork surface (both cork and blade should have been

sterilized before), the blade should be withdrawn from

the cork and autoclaved before being used again.

5. Hold the cork with the thumb and forefinger of the

free hand, and with the heel of the hand resting on

the patient’s arm, quickly make two skin punctures

(actually they are small incisions) in the selected area.

It is important that the surface of the cork meet the skin

to ensure a 3 mm deep incision. Holding the skin taut

prevents the test area from being depressed when the

blade enters the skin.

6. Timing is begun as soon as the incisions are made and

bleeding starts.

7. Using the edge of a piece of a filter paper to collect the

blood, gently touch paper to the drop of blood, which

Clinical Hematology: Bleeding Disorders 275

forms over the wound every 30 seconds. Do not rub or

remove the clot. Do not touch the skin. Any disruption

of formed fibrin or clot will prolong the bleeding time.

8. The bleeding time is reported when no blood stain

is seen on the filter paper after a gentle touch. It is

reported in intervals of 30 seconds. One can measure

both wounds and average them, or take the reading of

the last one to stop bleeding.

Normal Values

Normal values are 1 to 6 minutes. More than 6 minutes

should be taken as abnormal.

Interpretation

1. Results of duplicate tests performed on the same

individual should agree within 2 to 3 minutes at most.

2. Bleeding time is prolonged:

• When platelet count < 100,000/mm3

• In patients on aspirin therapy.

• In acquired fibrinogen disorders.

(If the platelets are young even in a thrombocytopenia

patient, the bleeding time may not be raised as young

platelets have enhanced hemostatic capabilities).

When platelet counts are low, one can calculate the

expected bleeding time with the following formula:

Bleeding time =

Platelet count/cu mm

30 5

3850

. ×

A bleeding time longer than that calculated from platelet

numbers alone, suggests defective platelet function in

addition to reduced number. It is also possible to detect

above-normal hemostatic capacity in cases in which

active young platelets comprise the entire population of

circulating platelets.

Clinical Implications

1. Bleeding time is prolonged when the level of platelets

is decreased or when the platelets are qualitatively

abnormal, as in

a. Thrombocytopenia

b. Platelet dysfunction syndromes

c. Decrease or abnormality in plasma factors such as

von Willebrand’s factor and fibrinogen

d. Abnormalities in walls of the small blood vessels—

vascular defects

e. Severe liver disease

f. Leukemia

g. Aplastic anemia

h. DIC disease.

2. Bleeding time can be either normal or prolonged

in von Willebrand’s disease. It will definitely be

prolonged if aspirin is administered prior to testing.

3. A single prolonged bleeding time does not prove the

existence of hemorrhagic disease because a larger

vessel may have been punctured. The puncture should

be done twice (on the contralateral side) and the

average of the bleeding times can be taken.

Interfering Factors

1. The normal range may vary when the puncture is not

of standard depth and width.

2. Touching the incision during the test will break off any

fibrin particles and prolong the bleeding time.

3. Heavy alcohol consumption (as in alcoholics) may

cause bleeding time to be increased.

4. Prolonged bleeding time will result from the ingestion

of 10 g of aspirin up to 5 days before the test.

5. Other drugs that may cause the bleeding time to be

increased include:

• Dextran

• Streptokinase—streptodornase

• Mithramycin

• Pantothenyl alcohol.

Patient Preparation

1. Explain the purpose and procedure of the test to

patient.

2. Warn patient not to consume aspirin for 5 days prior

to test.

3. Advise patient not to consume alcohol in any form.

Coagulation Time

Capillary Tube Method of Wright

Blood is collected in about a dozen capillary tubes from

a finger prick made after aseptic precautions. The tubes

are sealed with plasticine and immersed in water bath at

37o

C. After 4 minutes, remove the first tube from the bath

and expel the blood in it with one end immersed in a dish

containing water. Repeat this every 30 seconds with the

other tubes till the blood is expelled in a worm clot and note

the time.

An alternative way of determining the end point is to

break the capillary tubes every 30 seconds until a clot is

seen between the two broken ends. By these methods, the

normal clotting time is 5 to 10 minutes at 37oC and longer

if performed at room temperature. This test should be

avoided as tissue thromboplastin contaminates the oozing

blood and hence, false reports may be obtained.

Lee and White’s Method

Principle: Whole blood, when removed from the vascular

system and exposed to a foreign surface, will form a solid

276 Concise Book of Medical Laboratory Technology: Methods and Interpretations clot. Within limits, the time required for the formation of

the solid clot is a measure of the coagulation system.

Requirements

1. Stop watch

2. Equipment for collection of blood

3. Clean, dry glass test tubes (10 × 75 mm)

4. Water bath (37°C).

Method

1. Make a clean venipuncture with as little trauma to

(or time spent passing through) the connective tissue

between skin and vein as possible. One may routinely

or in selected cases use the two-syringe technique,

whereby one rinses the needle of all interstitial tissue

fluid by drawing back 1 cc. of blood after entering the

vein. Then remove the first syringe from the needle

and quickly place on a second clean and dry syringe

and draw back blood for the test.

2. Timing is begun when the blood first enters the syringe.

The second syringe in the ‘Two-syringe’ technique.

3. Draw 3–5 mL of blood and withdraw the syringe and

needle. Disconnect the needle. Place approximately

1 mL of blood in each of three (10 × 75 mm) test tubes.

4. Place the tubes in a stand so that they remain upright

and undisturbed, at room temperature for 10 minutes.

If a 37oC water bath is available one may do the entire

test at 37oC, and shorter clotting times will be found

(if the test has been done at 37oC, do not wait for more

than 5 minutes).

5. After 10 minutes (or 5 minutes) take the first of the

tubes and gently tip it every 30 seconds to test for

clotting. Do not tip it further than necessary to get the

information.

6. When the first tube is clotted (can be inverted without

blood running down the edge of the tube), record the

time and start the tipping of the second tube every 30

seconds until it is also found to be clotted. Then do

the same with the third tube (tipping is intended to

allow one to ascertain when blood is clotted—not as

a means of hastening clotting or of assuring mixing of

the blood).

7. The time recorded for the clotting of the third tube is

taken as the clotting time (the purpose of the first two

tubes is to tell one when to start looking in the third

tube, since the agitation of tipping does hasten the

clotting).

 Some choose to tip the tubes in rotation (at 37oC) every

15 seconds, or tip all tubes at once, and average the

results of the three tubes.

Normal Values

Normal times depend on method used. Normal range

at 37oC is usually 5 to 10 minutes. Normal times at room

temperature will vary with the degree of temperature

present and the method used. If one uses the method

which waits 10 minutes before starting to tip, then normal

values may go as high as 22 to 25 minutes, especially in

the cool season. Values shorter than 10 minutes should

be suspected and the test repeated using the two-syringe

technique to rule out contamination by tissue fluid (in the

heat of April, May or June warm tropical climate blood will

clot before 10 minutes without having been contaminated

by tissue fluids). If one uses the method which waits 5

minutes before tipping begins, normal results are between

8 to 18 minutes. Longer than 20 minutes is abnormal. If

clotting occurs in less than 7 minutes, the test should be

repeated using two-syringe technique.

Precautions and Errors

1. The venipuncture must be without trauma to avoid

contamination with tissue thromboplastin.

2. If all three tubes are clotted at 10 minutes (or 5

minutes) when one starts to tip the first tube, the test

is unsatisfactory and should be repeated. If blood was

drawn by single syringe technique, the most likely

explanation is contamination, of the blood by tissue

thromboplastin. If a two-syringe technique is used, it

suggests that the patient’s blood is hypercoagulable.

3. Vigorous agitation of the tubes will significantly

shorten the coagulation time. So tipping should really

be very gentle just to see if the blood has clotted.

Clinical Implications

1. Severe deficiencies of any of the coagulation factors

must be present before the coagulation time will

be prolonged. Fibrinogen for example, needs to

be decreased to 50 mg/100 mL or less before the

coagulation time is affected, the normal range of

fibrinogen is 200 to 400 mg/100 mL.

2. When prothrombin is diminished to a level of 30% of

normal, there will be a small change in coagulation

time.