Sesmed Medikal

Biosystems Biyokimya Kitleri

Biosystems prothrombin time (pt)

Biosystems prothrombin time (pt)

PRINCIPLE OF THE METHOD
The addition of calcium thromboplastin to plasma samples induces the formation of the fibrin
clot1
. The method measures the clot formation time.
CONTENTS AND COMPOSITION
A. Reagent: 4 x 5 mL. Tissue thromboplastin from rabbit brain with stabilizers. Lyophilized.
B. Reagent: 4 x 5 mL. Buffer, with calcium ions and sodium azide as preservative.
STORAGE
Store at 2-8ºC.
Reagents are stable until the expiry date shown on the label when stored tightly closed and if
contaminations are prevented during their use.
Indications of deterioration:
 Reagent B: Presence of particulate material, turbidity.
AUXILIARY REAGENTS
Coagulation Control I (BioSystems Cod. 61007), Coagulation Control II (BioSystems Cod.
61008).
Coagulation Calibrator (BioSystems Cod. 61006).
Fibrinogen Clauss B (BioSystems Cod. 61003).
REAGENT PREPARATION
Reagent B is ready to use.
Working Reagent: Pour the contents of Reagent B into the Reagent A bottle. Mix gently
(Note 1) and keep at 18-25ºC for 30 minutes. Stability after reconstitution in the original vial:
8 hours at 37ºC, one day at 22ºC, five days at 16ºC and twelve days at 2-8ºC.
Do not freeze.
ADDITIONAL EQUIPMENT
 Coagulometer
SAMPLES
Venous blood collected by clean venipuncture2
. Mix nine parts of blood with one part of
0,109 mol/L trisodium citrate dihydrate. Mix the blood carefully and centrifuge at 1500 x g
during 15 minutes to obtain the plasma3
Uncentrifuged or centrifuged samples without separating plasma from cellular components
may be stored for up to 24 hours at room temperature3
. It is not recommended to store the
samples at 2-8ºC. Separated plasma (without cells) may be stored for up to fifteen days
at -20ºC or for up to six months at -70ºC. Thaw frozen plasma at 37ºC just prior to use3

PROCEDURE
1. Bring the Working Reagent to 37ºC.
2. Pipette 50 µL of sample (patient plasma or control) to the test tube.
3. Incubate sample at 37ºC for 2 minutes.
4. Pipette 100 µL of Working Reagent and simultaneously start stopwatch.
5. Determine the coagulation time.
This procedure refers to manual or semi-automatic coagulometers. In case of determination
by an automatic coagulometer, please refer to the manual for specific instructions.
CALIBRATION
Calibration is only needed when results must be reported in percentage (see Calculations).
Use Coagulation Calibrator (BioSystems Cod. 61006) to prepare a calibration curve starting
from 1/1 and followed by 1/2, 1/3 and 1/4 dilutions with Fibrinogen Clauss B (BioSystems
Cod. 61003).
Alternatively, a calibration curve is provided with each kit. The same calibration curve can be
used when using the same lot of reagent and performing a daily quality control.
CALCULATIONS
PT is generally expressed in seconds, but it can also be calculated as a INR (International
Normalized Ratio, Note 2), using the following general formula:
INR = (Patient PT / Mean normal PT)ISI
ISI: International Sensitivity Index. It is lot specific and reagent/instrument depending.
Mean normal PT: Mean of the normal PT range, using 20 or more individual samples. It
should be determined in each laboratory.
Alternatively, PT can be expressed in percentage by plotting the different % values of the
Coagulation Calibrator dilutions versus the PT in seconds. The % of PT present in the
sample is calculated by interpolating its PT in the calibration curve.

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