PB0701 - Automated immunoassay vs. Flow cytometry-based functional assay in the diagnosis of HIT

Background: Heparin-induced thrombocytopenia (HIT) is an immune complication of heparin therapy caused by antibodies to complexes of platelet factor 4 (PF4) and heparin. Pathogenic antibodies to PF4/heparin bind and activate cellular FcγRIIA on platelets and monocytes leading to thrombocytopenia and life-threatening thrombosis in some patients. Laboratory assays detect the presence of HIT antibodies using functional platelet activation assays or immunoassays.

Aims: To determine the utility and challenges of a functional flow cytometry-based test in comparison with a semi-quantitative immunoassay for the diagnosis of HIT.

Methods: Citrated plasma from 45 patients with clinically suspected HIT were processed using the Emo-test HIT CONFIRM test (Emosis SAS, France) on the DxFlex flow cytometer (Beckman Coulter, USA), and a fully automated latex enhanced immunoassay test (HemosIL immunoassay; IL, Massachusetts, USA).

Results: The immunoassay was considered positive when the HemosIL HIT antibody levels were ≥1.0 U/ml. The flow-based Emo-test HIT assay uses a HEparin induced PLAtelet activation index (%HEPLA) calculated as follows: [(%H0.3 − %H100)/ (% positive control − % negative control)] × 100. Cutoffs defined by the manufacturer were used [≥13%: positive result; ≥9.6%: inconclusive results, < 9.6: negative] (Figure.1). Using the immunoassay as the standard, the sensitivity, specificity, positive predictive value, and negative predictive value of the flow-based assay was 75%, 92.7%, 50% and 97.4% respectively (Table. 1).

Conclusion(s): Flow-based assay had high specificity and negative predictive value for a diagnosis of HIT with moderate sensitivity. Donor platelets play a critical role in the assay, and it is therefore imperative to identify a pool of donors whose platelets are sensitive to activation by Heparin. Availability of such donors at the time of testing and the technically challenging nature of the assay poses the biggest challenges of this functional assay. We are limited by not performing a gold standard test such as Serotonin release assay.