Identifying patients at high risk of heparin-induced thrombocytopenia-associated thrombosis with a platelet activation assay using flow cytometry

Abstract

To diagnose heparin-induced thrombocytopenia (HIT), detection of platelet-activating antibodies (HIT antibodies) is crucial. However, serum platelet activation profiles vary across patients and depend on test conditions. We evaluated the association between clinical outcomes and platelet-activating profiles assessed by a platelet microparticle assay (PMA), which detects activation of washed platelets induced by HIT antibodies, in 401 consecutive patients clinically suspected of having HIT. We made modifications to the assay, such as donor selection for washed platelets that increased sensitivity. Serum that activated platelets at a therapeutic (but not high) heparin concentration was defined as positive. Of these, serum that activated platelets within 30 minutes or in the absence of heparin was defined as strongly positive. The remaining samples were considered weakly positive. As a result, 97 % and 93 % of patients who tested strongly and weakly positive had clinical profiles consistent with HIT, respectively. The incidence of thromboembolic events (TEEs) after heparin exposure in patients who tested strongly positive, weakly positive, and negative was 61 %, 40 %, and 29 %, respectively. Among patients who did not experience a TEE on the day HIT was suspected, there was no significant difference in the cumulative incidence of subsequent TEEs between patients who tested strongly and weakly positive when argatroban was initiated on the same day (19.0 % vs 7.1 %, p=0.313), but there was a significant difference when argatroban therapy was delayed by one or more days (61.1 % vs 17.6 %, p=0.007). The modified PMA is effective in diagnosing HIT and identifying patients at high risk for HIT-associated TEEs.

Heparin‐induced thrombocytopenia and thrombosis in primary lymphedema patients who underwent vascularized lymph node transplantations

“…Therefore, developing a sensitive approach to identify patients at high risk of HITT is critical for microsurgeries 25 . One of the clinical presentations of HITT is platelet consumption, which is directly related to the disease severity of HITT 54,55 . In this study, we found statistically significant differences in the percentage of platelet reduction on the first day postoperatively (21.0% in HITT and 14.0% in non‐HITT, p < 0.01).…”

Deterioration of vaccine‐induced immune thrombotic thrombocytopenia treated by heparin and platelet transfusion: Insight from functional cytometry and serotonin release assay

“…[16][17][18] We also measured platelet-activating antibodies by flow cytometric assay based on a platelet microvesicle assay (PMA), which detected the activation of washed platelets induced by activating heparin-dependent antibodies. 19 The PMA was performed as previously described by Maeda et al 19 by the evaluation of plateletactivating profiles, which detects the activation of washed platelets induced by activating antibodies in plasma or serum. In the SRA and PMA, platelet activation was challenged by an Fc-receptor-blocking monoclonal antibody (anti-CD32/FcγRIIA, clone IV.3; STEMCELL Technologies, Vancouver, BC, Canada) to confirm FcγRIIA engagement in platelets.…”

“…The platelet activation index (iPA) was calculated as previously described. 19 This iPA was obtained after sample analysis with flow cytometry to quantify the extent of washed-platelet activation. iPA is defined as the ratio (percentage) of the number of fluorescent events in the left upper quadrant to the total number of fluorescent events in the left and right upper quadrants combined.…”

Association of Trauma Severity with Antibody Seroconversion in Heparin-Induced Thrombocytopenia: A Multicenter, Prospective Observational Study

“…(13) The washed platelet activation assay was performed as described in detail elsewhere. (2,14) In brief, washed platelets, prepared from HIT antibody-sensitive healthy volunteers, were used. Platelet microparticles, activated by HIT antibodies in a heat-inactivated serum, were quanti ed by ow cytometry.…”

“…(1) A segment of anti-PF4/heparin, an IgG with high titers, has platelet-activating properties which cause HIT. (2,3) In addition, these antibodies activate monocytes and endothelial cells leading to thrombocytopenia and a thrombin-induced hypercoagulable state. (4)(5)(6)(7) HIT is an atypical immune response when compared to other immune-mediated diseases caused by adaptive immunity (8-10): heparin-naïve patients can develop IgG antibodies as early as day 4, like a secondary adaptive immune response; evidence for an anamnestic response on heparin re-exposure is lacking; HIT antibodies are relatively short-lived unlike those from a secondary adaptive immune response.…”