A Platelet Factor 4-Dependent Platelet Activation Assay Facilitates Early Detection of Pathogenic Heparin-Induced Thrombocytopenia Antibodies

Abstract

Heparin-induced thrombocytopenia (HIT) is a dangerous complication of heparin therapy. HIT diagnosis is established by recognizing thrombocytopenia and/or thrombosis in an affected patient and from the results of serological tests such as the platelet factor 4 (PF4)/heparin immunoassay (PF4 ELISA) and serotonin release assay (SRA). Recent studies suggest that HIT antibodies activate platelets by recognizing PF4 in a complex with platelet glycosaminoglycans (and/or polyphosphates) and that an assay based on this principle, the PF4-dependent P-selectin expression assay (PEA), may be even more accurate than the SRA for HIT diagnosis. Here, we demonstrate that the PEA detected pathogenic antibodies before the SRA became positive in two patients with HIT studied serially, in one case even before seropositivity in the PF4 ELISA. In one of the patients treated with plasma exchange, persistent dissociation between the PEA and SRA test results was observed. These results support a role for the PEA in early HIT diagnosis.

Keywords: HIT; early diagnosis of HIT; heparin; plasma exchange, plasmapheresis; thrombocytopenia.

Figure 1

PEA detects pathogenic HIT antibodies earlier than the SRA, and results dissociate during TPE treatment. HIT serologic results and platelet counts during the early part of patient 1’s hospitalization (A) and during TPE treatment (B) are shown. Time (hours, days after heparin exposure) is depicted on the abscissa; platelet count (percent of baseline [A] or absolute count [B], inverted gray triangle), PEA (% of maximal activation, orange circle), and SRA (% serotonin release, red diamonds) are depicted on the left ordinate; PF4 ELISA (OD, blue circle) is depicted on the right ordinate. The first positive result in each assay is shown in black squares in (A). Anticoagulation used is indicated on top of the horizontal bars shown in the figure. ELISA = enzyme-linked immunosorbent assay; HIT = heparin-induced thrombocytopenia; OD = optical density; PEA = PF4-dependent P-selectin expression assay; Pos = positive; SRA = serotonin release assay; TPE = therapeutic plasma exchange.

Figure 2

PEA detects pathogenic HIT antibodies before the SRA and PF4 ELISA. HIT serologic results and platelet counts during the hospitalization for patient 2 are shown. Time (days after heparin exposure) is depicted on the abscissa; platelet count (percent of baseline, inverted gray triangle), PEA (% of maximal activation, orange circle), and SRA (% serotonin release, red diamonds) are depicted on the left ordinate; PF4 ELISA (OD, blue circle) is depicted on the right ordinate. The first positive result in each assay is shown in black squares. Anticoagulation used is indicated on top of the horizontal bars shown in the figure. See Figure 1 legend for expansion of abbreviations.