Related Experiment Video
Updated: Jul 13, 2025
Procoagulant Platelet Characterization by Measuring Phosphatidylserine Exposure and Microvesicle Release from Human Purified Platelets
Published on: November 29, 2024
Heparin-activated procoagulant platelet assay: a flow cytometry-based functional test for heparin-induced
Lisann Pelzl1, Günalp Uzun1, Irene Marini1
1Institute for Clinical and Experimental Transfusion Medicine, University Hospital of Tübingen, Tübingen, Germany; Centre for Clinical Transfusion Medicine, Tübingen, Germany.
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.
