Prothrombin Time (aPTT)

Mr. Khaled alzatari

 Prothrombin time is the time required for the plasma
to clot after an excess of thromboplastin and an
optimal concentration of calcium have been
added.

 it is sensitive to a quantitative or qualitative

abnormalities of any of the factors involved in the
extrinsic and common pathways of the coagulation
system (Factors II, V, VII, X, and Fibrinogen).
 The PT used to determine the clotting tendency of
blood, in the measure of warfarin dosage, liver
damage, and vitamin K status.

 Occasionally, the test may be used to screen

patients for any previously undetected bleeding
problems prior to surgical procedures.
 When reagent thromboplastin--to which calcium
has been added--is mixed with plasma (derived
from sodium citrated whole blood), the time (in
seconds) it takes for the formation of a clot is
reported as the Prothrombin time (PT).

 Calcium is necessary for the correct orientation

and binding of a number of complexes including
: tissue factor-VIIa, IXa-VIIIa, and Va-Xa.
 Citrated plasma: 1 part of sodium citrate
solution (0.11 mol/ L) to 9 part of venous
blood, avoiding the formation of foam.
 Control: normal plasma (Commercial,
Pooled Plasma)
 Each Thromboplastin Reagents must be
calibrated against standard PT reagent
established by the WHO.
 ISI = International Sensitivity Index.
◦ ISI is assigned by the manufacturer for each lot of
reagent using reference material from WHO.
◦ The lower the ISI the more sensitive the Reagent
1.ISI of 1.8 to 2.4 = Low sensitivity
2.ISI of 1.4 to 1.8 = Average sensitivity
3.ISI 1.0 to 1.4 = High Sensitivity
 Reconstitute tissue thromboplastin according
to instructions. Label the thromboplastin with
the time, date and initials. The thromboplastin
reagent is stabile for 7 days after
reconstitution.

 Allow to sit 10-15 minutes, then invert gently

several times.

 Mix well prior to pipetting any of this reagent

at any step in this procedure.
 Prewarming of the reagent by pipetting 1-2
mls, using a plastic pipette, of the tissue PT
reagent into a plastic test tube and place
in a 370 C water bath incubator.

 Pipette 100 µL of normal control, Patient

PPP into each of the test tubes.
 Allow at least one (1) minute to reach 37°C.

 Pipette 200 µL of PT reagent into the tube

containing the control. Start the stop
watch simultaneously.
 Mix the tube and leave in the water bath for
a minimum of 7-8 seconds. Then remove,
wipe the exterior, tilt back and forth gently
until a visible clot is formed. As the clot forms,
the mixture will gelatinize and may turn
cloudy.

 Stop the stop watch and record the result. If

the results from run 1 and run 2 are within +
10% second from each other, average the
two results and report with appropriate units.
 If results are not within required limits, a
third run should be performed and
average the two that match within
acceptable limits.

◦ Be sure and cross out any values you are not

using for the final calculation. Include
measurement unit of seconds on report sheet.

 Carry out 1 significant figure passed the

decimal point. For example, if your result
is 12.23 seconds, report as 12.2 seconds.
 Repeat the procedure for the samples
and Record the time.
Results are expressed as the mean of the duplicate reading in:
Seconds
Prothrombin ratio
International Normalized Ratio (INR)
 PT: 11.0 – 13.0 seconds.
 Therapeutic levels are at a P/C ratio of 2.0 – 3.0

 CRITICAL VALUES*
PT critical value changed to > 42.1 seconds;
INR changed to > 4.5
 Used to monitor oral anticoagulant therapy
(Warfarin / Coumadin).
 When a patient who is not taking anti-
coagulant drugs has signs or symptoms of a
bleeding disorder
 When a patient is to undergo an invasive
medical procedure, such as surgery, to
ensure normal clotting ability.
1. Vitamin K deficiency
2. DIC,
3. Liver disease,
5. A deficiency in one or more of the Concerning
factors: Factor I (Fibrinogen), Factor II
(Prothrombin), Factor V (Proaccelerin, Labile
Factor), Factor VII (Proconvertin, Stable Factor,
Factor X (Stuart-Prower Factor, Factor XIII (Fibrin
Stabilizing Factor) In addition, inhibitors can cause
prolonged PT’s.
 Diet: ingestion of excessive green, leafy
vegetables will increase the absorption of
vit-K, which promotes blood clots.
 Alcoholism, Prolonged PT levels
 Diarrhea and vomiting decrease PT because
of dehydration.
 Quality of Vein puncture.
 Medication : Antibiotics , Aspirin,
Cimethidine….
 Prolonged Storage of plasma at 4o C.
1. Associated with specimen (Preanalytical)
a. Inappropropriate ratio of anticoagulant to blood
b. Failure to correct citrate volume if hematocrit >
55%
c. Clotted, hemolyzed or lipemic samples
d. Lack of PPP
e. Delay in testing or processing
f. Inappropriate storage
Sources of Error

2. Associated with Reagent (Analytical)

1. Incorrect preparation of reagents
2. Use of reagents beyond reconstituted
stability time or expiration date
3. Contaminated reagent.
3. Associated with procedure (Analytical)
1. Incorrect temperature
2. Incorrect incubation times
3. Incorrect volumes of sample, reagents or
both