The rapid Bethesda assay is equivalent to the standard Bethesda assay for detection of factor IX inhibitors in patients with severe haemophilia B.

INTRODUCTION: The time-dependent nature of factor VIII (FVIII) inhibitors is well described, and the standard FVIII Bethesda assay used to measure inhibitors incorporates a 2-hour incubation. Despite case reports and reviews describing the immediate-acting nature of factor IX (FIX) inhibitors, many coagulation laboratories continue to use a traditional prolonged incubation for FIX Bethesda assays. To our knowledge, a comprehensive evaluation of the FIX Bethesda assay without incubation has not been reported. AIM: The goal of this study was to evaluate the performance of a rapid FIX Bethesda (ie no incubation) compared with the standard Bethesda assay (2-hour incubation). METHODS: The analysis used a Bethesda assay configured for either immediate testing or a 2-hour incubation. Samples from 14 haemophilia B patients with inhibitors and 9 non-human controls were tested. RESULTS: The two assays yielded similar performance overall. The average per cent difference in inhibitor titre between the rapid and standard FIX Bethesda assay was -3% (range -15% to +13%; P = .175) for patient samples and -2% (range -17% to +14%; P = .376) for controls. CONCLUSION: The rapid Bethesda assay showed good agreement with the standard Bethesda assay for determination of inhibitor levels in patients with severe haemophilia B. The rapid assay allows for faster assessment of inhibitors in patients with severe haemophilia B and has the potential to improve the ability of the coagulation laboratory to perform testing from a logistical viewpoint. Further studies involving larger numbers of patients would be important to confirm our findings.

Haemophilus influenzae triggers autophagy in HEp-2 cells.

The MAP-LC3 system regulates the intracellular formation of autophagy-associated vacuoles. These vacuoles contain the LC3 protein; thus it has been utilized as a marker to identify autophagosomes. The aim of our study was to investigate whether Haemophilus influenzae strains and their supernatants could activate autophagy in human larynx carcinoma cell line (HEp-2). We demonstrate that higher expression of the LC3B-II protein was induced, particularly by nontypeable Haemophilus influenzae (NTHi) 49766 and by supernatants, containing <50 kDa proteins, of both strains. Ultrastructural studies demonstrate vacuoles with a double membrane and/or membrane material inside, showing similar features to those of autophagic vacuoles. Together, our findings demonstrate that H. influenzae strains and their supernatants trigger an autophagic process.