The development of methods in the diagnostics of heparin-induced trombocytopenia

Objective: Heparin-induced thrombocytopenia (HIT II) is an immune-mediated disorder observed in up to 0.5-5% of heparin-treated patients. Laboratory testing for the diagnosis of HIT II includes a 14C-serotonin release test, PF4/heparin enzyme-immunoassay, platelet aggregation and flow cytometric assay with various specificities or sensitivities for the diagnosis of HIT II. The aim of this work was to develop an HPLC method with electrochemical detection (EC) and prove the possibility of using a non-radioactive serotonin release test for the diagnosis of HIT II. Material and Methods: Two groups were tested. First group of patients suspect for HIT II, and second control group with or without heparin therapy. Serotonin release was analyzed by a newly developed HPLC-EC method. Results: Analytical performance of this method was satisfactory. The method was linear from 5 to 7000 nmol/l with a limit of detection of 1 nmol/l. The intra- and inter-assay coefficients of variation were 7.7% and 12%, respectively, for a concentration of serotonin 1460 nmol/l. Conclusion: In the present work we describe rapid, non-radioactive, specific and sensitive HPLC-ED methods to determine serotonin release from the donor platelets