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Several highly purified products, suitable for haemophilia A and von Willebrand disease management, can be obtained, through the same chromatographic process, on an industrial scale.

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The mean value of FVIII:C (0.99 IU/mL) in whole-blood plasma produced by the 10 Italian regions is higher than that reported in other studies. A total of 83.1 percent of units have a FVIII:C level of at least 0.70 IU per mL. The mean level of FVIII:Ag is lower than that of FVIII:C. FVIII:Ag is higher in those units with a FVIII:C level of less than 0.70 IU per mL, while it gradually decreases as FVIII:C exceeds 0.70 IU per mL, thus showing a greater resistance to handling of plasma in the production steps mostly affecting FVIII:C stability.

The von Willebrand ristocetin cofactor assay is still the main cleared measurement test used to evaluate von Willebrand factor (vWF) activity in concentrate samples containing vWF. Although the assay's performance has been improved over the years, the test reliability is still affected by a high interassay and interlaboratory variability; moreover, it requires skilled technicians and significant time. An automated HemosIL vWF:activity test, already set up on plasma samples, was then applied to factor VIII/vWF concentrates to verify its suitability in routine analysis of concentrate samples. As first step precision, linearity and accuracy were assessed. Then, 40 commercial batches of a high purity factor VIII/vWF concentrate were examined with this new method. The Spearman's correlation between the results obtained with the HemosIL vWF:activity assay and vWF activities determined by established procedures was evaluated. Comparisons of the means between HemosIL vWF:activity and the other tests were calculated with the Wilcoxon and Bland-Altman tests. Validation study showed satisfying within-run and between-run precision values. Activities determined with HemosIL vWF:activity had good correlation with those determined as ristocetin cofactor, Imubind (vWF:Imubind) and collagen-binding. Wilcoxon test, used to compare the means between the HemosIL vWF:activity and the other activity assessments, proved no significant variation with vWF:Imubind, but displayed a slight variation with von Willebrand ristocetin cofactor and von Willebrand factor:collagen-binding. This automated HemosIL vWF:activity test could be included in routine determination of vWF activity in concentrate samples and supports traditional von Willebrand ristocetin cofactor because it is reliable, reproducible and sensitive.

The identification of hemoglobin (Hb) variants is usually performed by means of different analytical steps and methodologies. Phenotypic methods, such as gel electrophoresis and high performance liquid chromatography, are used to detect the different electrophoretic or chromatographic behaviors of hemoglobin variants in comparison to HbA0 used as a control. These data often need to be combined with mass spectrometry analyses of intact globins and their tryptic peptide mixtures. As an alternative to a 'step-by-step' procedure, we have developed a 'single step' approach for the identification of Hb variants present in biological samples. This is based on the microHPLC-ESI-MS/MS analysis of the peptide mixture generated by a tryptic digestion of diluted Hb samples and an in-house new database containing solely the variant tryptic peptide of known human Hb variants. The experimental results (full MS and MS/MS spectra) are correlated with theoretical mass spectra generated from our in-house-built variant peptide database (Hbp) using the SEQUEST algorithm. Simple preparation of samples and an automated identification of the variant peptide are the main characteristics of this approach, making it an attractive method for the detection of Hb variants at the routine clinical level. We have analyzed 16 different samples, each containing a different known variant of hemoglobin.

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