A rapid point-of-care test for the diagnosis of factor VIII inhibitors in hemophilia A patients.

BACKGROUND

The development of neutralizing antibodies or inhibitors against exogenous factor (F)VIII represents a major adverse event in hemophilia A treatment. The conventional assay for the diagnosis of FVIII inhibitors is the Nijmegen-modified Bethesda assay. This test is time-consuming, resource-intensive, and cost-prohibitive. The test lacks sensitivity and shows wide variation across quality surveys.

OBJECTIVES

To develop a rapid, cost-effective, functionally relevant, sensitive, and specific test for FVIII inhibitors.

METHODS

We developed and validated a cost-effective, accurate, and equipment-free point-of-care (POC) test for FVIII inhibitors in a paper-strip format using full-length recombinant FVIII and gold nanoparticle-labeled antibodies.

RESULTS

The diagnostic sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of the FVIII inhibitor POC test were 0.93, 0.99, 0.94, 0.99, and 0.98, respectively, compared with the Nijmegen-modified Bethesda assay. The test was rapid (10 minutes), cost-effective (USD ≈$1-$1.5), equipment-free (naked-eye test), and required only a small volume (50 μL) of citrated whole blood or citrated plasma. The POC test was unaffected by nonneutralizing antibodies, residual FVIII, emicizumab, lupus anticoagulants, or heparin.

CONCLUSION

The POC test shows promise as a valuable screening tool in resource-limited settings for timely diagnosis of FVIII inhibitors and can be applied even at the lowest tier of the health care system, where sophisticated equipment or technical expertise may not be available. With a turnaround time of 10 minutes, this POC test will be highly useful for timely clinical decision-making.