A sensitive and specific functional flow cytometric assay for the diagnosis of heparin-induced thrombocytopenia

A functional flow cytometric assay (FCA) for the immediate diagnosis of heparin-induced thrombocytopenia (HIT), with simultaneous compatibility testing for alternative anticoagulant therapies, has been developed to provide rapid and reliable results which effectively support patient management. The assay provides results within 1–2 h, uses readily available non-radioactive reagents, and employs standard equipment. Using the highly sensitive annexin V protein probe, the method detects activated platelets induced by heparin immune-complexes, with 300-fold increased binding to activated platelets. Twenty-five samples from patients clinically-suspected of having HIT (131 tests) and 10 normal control (NC) samples (36 tests) were simultaneously tested with unfractionated heparin (UH) and low-molecular-weight heparin (LMWH), and by the radioactive serotonin-release assay (SRA) (62 and 16 tests respectively). The FCA highly correlated with the SRA, showing 100% specificity and 95% sensitivity. Moreover, the FCA exhibited higher resolution between positive and negative samples (an average value of 8.6-fold the NC versus 4.0-fold the NC by SRA). The LMWH showed concordant results with UH ( r =0.95). We conclude that the functional FCA for HIT is practical, specific and sensitive, thereby permitting the rapid diagnosis of HIT and the suitability of alternative therapies.