|
Test ID: FVIII CH
|
|
Factor VIII Chromogenic assay
|
|
|
|
Useful For
|
• This assay will be performed in situations where there is a clinical suspicion of hemophilia A diagnosis, but the 1-stage Factor VIII (FVIII) assay is normal
• For determination of FVIII activity in patients with hemophilia A receiving Emicizumab.
• For determination of FVIII inhibitor levels in patients receiving Emicizumab.
• Measurement of FVIII in patient with Lupus anticoagulant if LA insensitive APTT reagent is not available.
• FVIII assay in patient on direct thrombin inhibitor oral anticoagulants.
• For post infusion monitoring of modified, extended half-life recombinant concentrates, if chromogenic assay is recommended by the drug manufacturer.
|
|
Method name and description
|
Chromogenic factor VIII is a two-stage assay, the main rate-limiting step is the factor VIII level present in the test plasma. In stage one, factor VIII in the sample is activated by thrombin. Activated Factor VIII (FVIIIa) then accelerates the conversion of Factor X (FX) into Factor Xa (FXa) in the presence of activated Factor IX (IXa), Phospholipids (PL) and calcium ions. In the second stage of this assay, the activated factor X cleaves a chromogenic substrate and the FXa activity is assessed by hydrolysis of a p-nitroanilide substrate specific to FXa. The initial rate of release of p-nitroaniline (pNA) measured at 405nm is proportional to the FXa activity, thus to the FVIII activity in the sample.
|
|
Clinical information
|
Endothelial cells of the liver are the major site of FVIII synthesis.
Factor deficiency is often discovered when activated partial thromboplastin time is found to be prolonged. The deficiency causes bleeding disorder.
|
|
|
Specimen type / Specimen volume / Specimen container
|
Plasma Na Cit. Platelet-poor plasma.
Specimen Volume:2.7 ml.
Container/Tube: Light-blue top (citrate)
|
|
Collection instructions / Special Precautions / Timing of collection
|
1- Sample collected in HMC facilities/others and transported to the lab within 1hour :
• Collect sample into light blue-top (sodium citrate).
• Tubes must be filled to the mark on the tube side to provide a 9:1 ratio of blood to citrate.
2- Samples collected in HMC facilities / others and which are expected not to reach the lab within 1hour:
• Centrifuge the whole blood at 1500-2000g (approximately 3000-4000rpm) for 10 minutes at room temperature and carefully remove the plasma by pipette not by decanting and send the plasma in a temperature-controlled environment (18-25ºC ) within one hour of centrifugation.
3- If the transportation of the plasma can’t be within a maximum of 2hour , prepare platelet-poor plasma as follow:
A: Re-centrifuge plasma again.
B: Remove the top portion of plasma leaving approximately 250 mcL in the bottom to discard.
C: The double-centrifuged plasma should be aliquoted (0.5 to 1 mL per aliquot) into labeled plastic tubes. The number of tests ordered will determine the aliquots needed.
D: Freeze the separated plasma immediately at -20 ºC or below .
E: Specimens must arrive frozen.
4- Consider patient HCT %. , if HCT >55%, the volume of anticoagulant in the tube must be adjusted by the referring lab, as per the below table shows the volume of citrate (µL) to be removed from common sizes of coagulation specimen tubes prior to specimen collection. After removal of the specified volume of citrate, enough blood is added to fill the tube to the ideal volume.
| Hematocrit % |
Volume to be removed, µL |
| 2.7 ml Tube |
1.0 ml Tube |
| 56 |
50 |
20 |
| 57 |
50 |
20 |
| 58 |
60 |
20 |
| 59 |
60 |
20 |
| 60 |
70 |
30 |
| 61 |
70 |
30 |
| 62 |
80 |
30 |
| 63 |
80 |
30 |
| 64 |
90 |
30 |
| 65 |
90 |
30 |
| 66 |
100 |
40 |
| 67 |
100 |
40 |
| 68 |
110 |
40 |
| 69 |
110 |
40 |
| 70 |
120 |
40 |
| 71 |
120 |
40 |
| 72 |
130 |
50 |
| 73 |
130 |
50 |
| 74 |
140 |
50 |
| 75 |
140 |
50 |
| 76 |
150 |
50 |
| 77 |
150 |
60 |
| 78 |
160 |
60 |
| 79 |
160 |
60 |
| 80 |
170 |
60 |
|
|
Relevant clinical information to be provided
|
Patient/family history of bleeding disorder. History of anticoagulant therapy.
|
|
Storage and transport instructions
|
|
|
|
Specimen Rejection Criteria
|
Specimens with no label or missing required identification
Broken, leaking or contaminated specimen
Clotted samples
Under-filled or overfilled sample tubes.
Wrong sample container sample received
Improper specimen transport temperature (e.g. like specimens which are needed to be sent on ice)
Old specimen (test-dependent)
Grossly Hemolyzed sample (test-dependent)
|
|
|
Biological reference intervals and clinical decision values
|
Reference range for age group > 18 years:70-150%.
For age group up to 18 years: reference range for FVIII clotting is used.
|
0 Minutes-4 Days
|
0 Minutes-4 Days
|
4 Days-1 Months
|
1 Months-6 Months
|
6 Months-1 Year
|
1 Year-5 Years
|
5 Years-10 Years
|
10 Years-18 Years
|
|
50-178
|
50-178
|
50-154
|
67-141
|
70-213
|
83-170
|
75-163
|
80-166
|
- FVIII activity is done by chromogenic assay.
- Chromogenic Factor VIII activity generally correlates with the one-stage FVIII activity. However, no specific data are available for full term/premature neonates, infants, children, and adolescents.
- Reference Interval: Factor levels are expressed as percent of blood plasma concentration. By definition, pooled plasma contains 100 %(1unit/ml) of factor VIII (clotting assay).
- The reference range for age group up to 18 years is quoted from literature. The reference range values have not been verified /established by lab.
|
|
Factors affecting test performance and result interpretation
|
No interference up to:
Triglycerides 6.78 mmol/L
Hemoglobin 1.0 g/dL
Bilirubin 615.7 µmol/L
|
|
Turnaround time / Days and times test performed / Specimen retention time
|
For routine ordered test : 3 working days
For stat ordered test :
- During the Laboratory working hours from 0700 hours to 1500 hours (Sunday to Thursday): TAT is Within 8hrs.
- After duty hours, weekends and on holidays: TAT is within 12 hrs. with the following requirements:
- Laboratory should be notified by telephone about the sending of a STAT sample.
- The sample should be hand-delivered to the central processing area and requesting them to deliver it immediately to the hematology coagulation lab.
Test is done : Daily.
Specimen retention time: NA
|
|
|
