Real World Data on the Utility of P-Selectin Expression Assay; Correlation of Test Results and Disease Expression
Authors: Sweidan Aroob , Brancamp Rachel L , Lopez-Plaza Ileana , Jacobsen Gordon , Kuriakose Philip 
Published: December 2024

Background:

Serotonin Release Assay (SRA); the gold standard confirmatory test for Heparin Induced Thrombocytopenia (HIT), is usually a reflex test done after the initial screening test detecting platelet factor 4 (PF4) antibodies using enzyme-linked immunosorbent assay (ELISA). SRA testing uses radioactive labeled serotonin, is time and labor consuming, and is only done in a limited number of laboratories around the United States. Given the difficulty in performing this confirmatory test, there has been a growing need for innovative testing methods.

P Selectin Expression Assay (PEA) is a relatively simple method for detecting platelet-activating HIT antibodies based on the expression of p-selectin/CD62p by normal platelets mixed with patient serum and exogenous PF4 in the absence of heparin. It is based on a non-radioactive, less labor-intensive, readily available technology (flow cytometry), with a shorter turnaround time.

A PEA test is considered positive only if the following 2 conditions are met: 1) P Selectin expression is greater than 35% when the patient's serum is mixed with 30mcg of PF4, AND 2) P-Selectin expression is suppressed greater than 50% after mixing with 100 units of heparin. We noticed that some patients with high clinical suspicion for HIT had initial P Selectin expression of less than 35% but had more than 50% suppression after mixing with 100U heparin.

We incorporated PEA testing as the primary reflex test at Henry Ford Hospital (HFH) following any positive anti-PF4 ELISA test (defined as Optical Density (OD) &gt; 0.4) in December of 2021. Only selected cases undergo SRA testing as a secondary confirmatory test when PEA results are negative in the presence of strong clinical suspicion.

Methods:

A retrospective review was performed on all patients who underwent PEA testing from January 2023 to May 2024. Data included admission diagnosis, previous exposure to heparin, initial platelet count, platelet nadir, interval time until the drop of platelets, thrombotic events, bleeding events, anti-PF4 ELISA OD value, PEA test values and interpretation, type of anticoagulation used, duration of anticoagulation, survival of hospital admission, and etiology of death if occurred. We calculated individual 4Ts scores for all patients. All anti-PF4 ELISA tests were performed at HFH special coagulation labs, and all PEA tests were performed at Versiti diagnostic laboratories (Milwaukee, WI).

Results:

We had a total of 61 PEA tests, of which 25 (40.9%) were positive and 36 (59%) were negative. Of the negative tests, 24 (66.6%) had P selectin expression less than 35% when serum was mixed with 30mcg of PF4 but had more than 50% suppression in P-Selectin expression after mixing with 100 units of heparin. Of these 24 patients, 5 (20.8%) had 4Ts score &gt; 5, 3 (12.5%) had 4Ts score of 7; 9 (37.5%) had PF4 antibody ELISA OD &gt;1.0, 5 (20.8%) with OD &gt;1.5 (2 of whom had OD values of 3.862 and 2.652); and 9 (37.5%) had a P Selectin expression of 10-20% when serum was mixed with 30mcg of PF4 and &gt;50% suppression when mixed with 100U of heparin. 15/24 (62.5%) were treated as HIT based on clinical suspicion, 4Ts scores, and OD values despite negative PEA results, most commonly with Bivalirudin followed by Warfarin or direct oral anticoagulants (DOACs).

Conclusion:

Though limited by small sample size, our study suggests that a finite number of patients with high clinical suspicion for HIT had negative PEA results. We found that these patients had a P Selectin expression of 10-20% (&lt;35%) when mixed with 30mcg of PF4 but more than 50% suppression when mixed with 100U of heparin. These results emphasize the importance of continuously reassessing binary tests. We propose reviewing larger data sets to assess if a separate intermediate PEA risk group might need to be defined while incorporating clinical parameters to finalize the diagnosis and help guide appropriate management.