Variability in factor VIII concentrate measurement: results from SSC field collaborative studies.
ABSTRACT Seven 'field' collaborative studies on factor (F)VIII concentrate potency measurements were carried out, using local routine methodology, standards and calculation of results. Data from five of the 12 different concentrates studied are described in detail. These studies revealed that, for the intermediate-purity and the recombinant FVIII concentrates, one-stage potencies were significantly lower than chromogenic potencies, whilst for the two high-purity FVIII concentrates one-stage potencies were significantly greater than chromogenic potencies. On comparing predilution methods for the intermediate-purity concentrate, equivalent potencies were obtained using either buffer or FVIII-deficient plasma as prediluent. For the two high-purity and the recombinant concentrates, potencies obtained using buffer as prediluent were significantly greater and lower, respectively, than potencies obtained using FVIII-deficient plasma as prediluent. Interlaboratory variabilities were compared over all 12 concentrates studied and coefficients of variation (CVs) for one-stage assays were found to be much greater than for chromogenic assays. This was true for all concentrates except for the intermediate-purity concentrate and samples A and B from the first study, where the reverse was true. Furthermore, much better CVs were obtained when using FVIII-deficient plasma than when using buffer as prediluent, for all FVIII concentrates except for the intermediate-purity concentrate where the reverse was true, and sample B where CVs were equivalent. Overall, CVs were far worse than those obtained in controlled collaborative studies. Generally, however, CVs were better with chromogenic assays and predilution in FVIII-deficient plasma, as is recommended by the International Society on Thrombosis and Haemostasis/Scientific and Standardization Committee, particularly for higher purity and recombinant concentrates.
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ABSTRACT: The determination of factor VIII (FVIII) potency in FVIII concentrates can be performed using both manual and automated methods. This work aimed to validate the use of the chromogenic kit Coamatic(®) FVIII (Chromogenix) on the automated ACL(®) Elite PRO analyzer for evaluating the potency of FVIII in commercial preparations in pharmaceutical analytical laboratories. After setting the activation and reading times to 2 min and 3 min, respectively, the validation parameters, according to the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use guideline Q2 (R1), were as follows: linearity, expressed by the adjusted model: log (Absorbance) = 1.848 + 0.777∙log (Concentration), with r(2) = 0.998; accuracy was verified (P-value = 0.6959); and the coefficient of variation for repeatability and intermediate precision was ≤6.5%. The Coamatic(®) FVIII kit method has been adapted to the ACL Elite PRO analyzer with improved performance compared with a manual microplate method.International journal of laboratory hematology 08/2012; · 1.30 Impact Factor
- Journal of Thrombosis and Haemostasis 03/2012; 10(6):1175-6. · 6.08 Impact Factor
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ABSTRACT: The quantitation of factor (F)VIII by activity-based assays is influenced by the method, procedure, the quality and properties of reagents used and concentrations of other plasma proteins, including von Willebrand factor (VWF). To compare FVIII concentrations measured by activity-based assays with those obtained by an immunoassay and to establish the influence of plasma dilution on the FVIII clotting activity (FVIIIc). The APTT, a chromogenic assay (Coatest) and two in-house immunoassays were used. Albumin-free recombinant FVIII was used as the calibrator in all assays. For a group of 44 healthy individuals (HI), the mean value observed for FVIII antigen (FVIIIag; 1.22+/-0.56 nM; S.D.) is substantially higher than that for FVIIIc (0.65+/-0.29 nM) and the chromogenic assay (FVIIIch; 0.50+/-0.23 nM). A positive correlation between FVIIIag and VWFag with R(2)=0.20 was observed. Since plasma VWF has an inhibitory effect on FVIIIc, we evaluated the influence of plasma dilutions on FVIIIc in HI (n=105). At a 4-fold dilution, estimates of FVIIIc by clotting assay were much lower than FVIIIag (0.77+/-0.31 vs. 1.14+/-0.48 nM). At 10- and 25-fold dilutions, the estimated FVIIIc increased to 0.87+/-0.36 and 0.94+/-0.44 nM, respectively. 1) In plasma, FVIIIag is higher than FVIIIc and FVIIIch; and 2) Real FVIII concentrations in plasma can be estimated by measuring FVIIIag.Thrombosis Research 05/2010; 126(2):119-23. · 3.13 Impact Factor