Evaluation of an automated chromogenic assay for Factor VIII clotting activity measurement in patients affected by haemophilia A

Introduction: The original one-stage clotting assay is still the most widely used method to measure Factor VIII clotting activity (FVIII:C) in patients with haemophilia A (HA), although the use of chromogenic assays is increasing significantly. Aim: Evaluation of the analytical performance and diagnostic accuracy of BIOPHEN™ FVIII:C (HYPHEN BioMed, Neuville-sur-Oise, France) assay on Sysmex CS-2400 (Sysmex, Kobe, Japan) analyser. Methods: Sixty patients with haemophilia A (HA; any severity) and 120 healthy Italian subjects were included. All the assays were performed on citrate platelet-poor plasmas stored at −80°C. Chromogenic BIOPHEN™ FVIII:C was compared with the one-stage assay using Actin FS and Factor VIII deficient plasma (Siemens Healthcare Diagnostics, Marburg, Germany) on Sysmex CS-2400 and with another chromogenic automated assay (COAMATIC™ Factor VIII, CHROMOGENIX on ACL TOP analyzer; Instrumentation Laboratory, Milan, Italy). Results: Intra-assay and inter-assay coefficient of variation were <6%. Linearity was good up to 1/128 dilution (r = 0.99); mean recovery was 91.7% and limit of detection was 0.2%. BIOPHEN™ FVIII:C assay showed a good correlation and diagnostic agreement with the chromogenic COAMATIC™ assay: the Spearmen's Rank correlation coefficient was 0.98 and the inter-rate agreement K Cohen coefficient was 0.61. The K coefficient was 0.91 when BIOPHEN™ FVIII:C was compared with the historical classification of the patients, demonstrating an optimal diagnostic accuracy in HA. Conclusions: BIOPHEN™ FVIII:C showed good analytical performance and diagnostic accuracy and could be considered suitable for the introduction in routine analytical panel of coagulation for the diagnosis of HA patients.