Heparin-activated procoagulant platelet assay: a flow cytometry-based functional test for heparin-induced thrombocytopenia.

Background: Functional platelet activation assays are required for the diagnosis of heparin-induced thrombocytopenia (HIT). Due to their sophisticated methodology, they are only available in reference centers.
Objectives: To evaluate the diagnostic accuracy of the flow cytometry-based heparin-activated procoagulant platelet (HAPP) assay in the laboratory diagnosis of HIT.
Methods: Procoagulant platelets (PCP), defined by the expression of phosphatidylserine and CD62-P, were evaluated by flow cytometry in platelet-rich plasma from healthy donors after incubation with patient sera in the absence and presence of heparin. A sample was considered positive in HAPP assay, if the following 3 criteria were met: 1) the percentage of PCPs was ≥10.3% after incubation with 0.2 IU/mL heparin, 2) the fold increase in presence of 0.2 IU/mL heparin compared with buffer was ≥1.5, and 3) 100 IU/mL of heparin resulted in ≥50% inhibition of PCP. HAPP assay was validated in a prospective cohort (n = 202) of consecutive specimens submitted to our laboratory for serologic diagnosis of HIT. Heparin-induced platelet activation (HIPA) assay was used as the reference standard.
Results: HIT-positive sera induced PCPs in the presence of 0.2 IU/mL heparin, which was inhibited with 100 IU/mL of heparin. In the prospective validation cohort, there were 15 HIPA+ and 187 HIPA- sera. HAPP was positive in 20 samples in this cohort. Using optimized cut-offs, HAPP assay had a sensitivity of 93.3% and specificity of 96.8%.
Conclusion: HAPP assay is promising as a simple and reliable functional assay for HIT; however, further studies are needed to confirm our results in larger cohorts.
Competing Interests: Declaration of competing interests Tamam Bakchoul has received research funding from CoaChrom Diagnostica GmbH, DFG, Robert Bosch GmbH, Stiftung Transfusionsmedizin und Immunhämatologie e.V.: Ergomed, DRK Blutspendedienst, Deutsche Herzstiftung, Ministerium fuer Wissenschaft, Forschung und Kunst Baden-Wuerttemberg, has received lecture honoraria from Aspen Germany GmbH, Bayer Vital GmbH, Bristol-Myers Squibb GmbH & Co, Doctrina Med AG, Meet The Experts Academy UG, Schoechl medical education GmbH, Stago GmbH, Mitsubishi Tanabe Pharma GmbH, Novo Nordisk Pharma GmbH, Leo Pharma GmbH, Swedish Orphan Biovitrum GmbH, has provided consulting services to: Terumo, has provided expert witness testimony relating to HIT and non-HIT thrombocytopenic and coagulopathic disorders. All of these are outside the current work. Tamam Bakchoul, Karina Althaus and Jan Zlamal have a pending patent application for flow cytometry-based assays to detect HIT and VITT antibodies. Other authors declare no competing financial interests.
(Copyright © 2023 International Society on Thrombosis and Haemostasis. Published by Elsevier Inc. All rights reserved.)