Evaluation of discrepancy between clot-based and chromogenic factor IX coagulation assays in non-severe hemophilia B patients and identification of the causing mutations.

Background: Hemophilia B, or Christmas disease, is a hemorrhagic inherited disorder. Previous studies have reported measurement discrepancies in factor VIII activity between clot-based and chromogenic assays in approximately one-third of patients with non-severe hemophilia A. However, similar discrepancies in hemophilia B have been less extensively studied. This research compares clot-based and chromogenic assays in 33 patients with non-severe hemophilia B and investigates the mutations associated with these discrepancies.

Methods: Citrate and EDTA samples were collected from 33 patients with non-severe hemophilia B at Iran's hemophilia comprehensive care center. Clinical information was also gathered. Both clot-based and chromogenic assays were performed on these patients. DNA was extracted from the EDTA samples for those with discrepancies in the test results, and PCR was conducted to sequence their genes to find mutations.

Results: Among 33 plasma samples from patients with non-severe hemophilia B, 7 showed a measurement discrepancy according to the definition of ISTH (<0.5, >2.0, or an absolute difference >10), which includes both reverse and classic types of discrepancies. In this study, mutations that previously did not show contradictory results now exhibit discrepancies. A difference in classification was observed in 21 % of the patients.

Conclusions: The findings indicate that the impact of specific mutations varies depending on the assay conditions. In addition to mutations, other factors also play a role in this discrepancy. Both types of assays are essential for the accurate diagnosis and classification of hemophilia B.