Optimization of pre‐analytical heat treatment for inhibitor detection in haemophilia A. (18th May 2018)

• Record Type: Journal Article
• Authors: Batty, P.; Hart, D. P.; Platton, S.
• Is Part Of: International journal of laboratory hematology. Volume 40:Number 5(2018:Oct.)
• Page Start: 561
• Page End: 568
• Publication Date: 2018-05-18
• Subjects: Bethesda -- haemophilia -- heat treatment -- inhibitors -- pre‐analytical; Hematology -- Periodicals; Blood -- Diseases -- Periodicals; Hematology -- Periodicals; 616.15005
• DOI: 10.1111/ijlh.12862
• Languages: English
• ISSNs: 1751-5521

Abstract

Abstract: Introduction: Factor VIII (FVIII) antibody formation is the greatest clinical and laboratory challenge within the haemophilia centre. The Nijmegen‐Bethesda assay (NBA) is the gold standard for inhibitor quantification, but affected by pre‐analytical variables including a patient's FVIII activity (FVIII:C). Pre‐analytical heat treatment (PHT) provides a methodology for inhibitor testing when measurable FVIII:C is present. Methods: We evaluated the effect of different PHT conditions (time/temperature) on FVIII:C as well as on potency of inhibitory activity in samples containing FVIII:C (endogenous pooled plasma and exogenous recombinant FVIII (rFL‐FVIII) concentrate) or FVIII inhibitor. Results: PHT of endogenous FVIII at 37°C, 47°C and 52°C resulted in declining measurable FVIII:C at 120 minutes (69%, 57% and 13% of the original FVIII:C, respectively). Incubation at 56°C resulted in FVIII:C ≤ 1IU/dL after 60 minutes for endogenous FVIII and 120 minutes for rFL‐FVIII. Incubation at 58°C resulted in FVIII:C < 1IU/dL at 15‐30 minutes for endogenous FVIII and at 30‐60 minutes for rFL‐FVIII. No difference was seen for inhibitor detection following PHT (56°C or 58°C) by NBA or anti‐FVIII IgG ELISA. Conclusion: PHT at 58°C for 30 minutes demonstrated consistent reduction in FVIII:C < 1IU/dL without appearing to affect inhibitor detection. Laboratory awareness of differences in thermostability of different sources of FVIII is important when choosing PHT conditions.