PF4-Dependent P-Selectin Assay Highly Accurate in Diagnosing Heparin-Induced Thrombocytopenia

A study published in Blood established the diagnostic accuracy of a PF4-dependent P-selectin expression assay (PEA) for heparin-induced thrombocytopenia (HIT), with a sensitivity similar to that of the “gold standard” serotonin release assay (SRA).

“The PEA is the first laboratory test shown to be as accurate as the SRA,” study co-author Anand Padmanabhan, MD, PhD, from Mayo Clinic in Rochester, Minnesota, told ASH Clinical News. Of note, “it is technically simpler than the SRA and uses 20-fold fewer platelets and does not require radioactive reagents.”

The study conducted by Dr. Padmanabhan and colleagues enrolled 409 consecutive adult inpatients who underwent enzyme-linked immunosorbent assay (ELISA) testing for suspected HIT from 2016 and 2017 at Mayo Clinic and the University of Washington.

Serum samples were obtained for immunoglobulin G-specific ELISA testing and were batch-tested with PEA and SRA with the same target platelets used in paired SRA-PEA runs. Samples were classified as HIT-positive, HIT-negative or HIT-indeterminate based on 4Ts scores and laboratory values, with the following manufacturer-recommended cutoffs for test positivity:

  • University of Washington: optical density (OD) >0.3
  • Mayo Clinic: OD ≥0.4

A total of 284 patients had low 4Ts scores (69.4%), while 98 and 27 had intermediate or high 4Ts scores (24% and 6.6%), respectively. Forty-nine ELISA results were positive and 360 were negative.

Using the predefined criteria, 17 patients were considered positive for HIT. People in this group had a median PEA of 88% and a median SRA of 69%. These rates decreased significantly in HIT-indeterminate patients, to 46% and 5%, respectively. The researchers added that platelet activation in the PEA and SRA was not observed in the majority of samples from HIT-negative patients.

According to receiver operating characteristic curve statistics, both the PEA and SRA correctly stratified patients into disease-positive and negative groups, with similar areas under the curve:

  • PEA: 0.94 (95% CI 0.87-1.0)
  • SRA: 0.91 (95% CI 0.82-1.0)

In a sensitivity analysis that considered indeterminate-HIT patients as disease-positive, 26 patients were classified as having HIT, and the PEA and SRA were again similar in terms of diagnostic accuracy, with AUCs of:

  • PEA: 0.88 (95% CI 0.78-0.98)
  • SRA: 0.86 (95% CI 0.77-0.96)

The concordance between PEA and SRA results was high for HIT-negative patients, and slightly lower for HIT-positive patients (TABLE). When looking at these discordant cases, 1 patient was found to have a false-negative test. Two of the 5 HIT-positive patients were re-exposed to heparin because of negative SRA results; both experienced a decrease in platelet counts following exposure confirming “true” HIT.

Dr. Padmanabhan noted that some patients with results that were “barely” positive by ELISA testing (e.g., <1 OD) had unequivocal HIT based on testing with PEA together with clinical history and platelet responses to heparin re-exposure. “Thus, while the strength of ELISA ODs generally correlates with platelet-activating antibodies, there are exceptions to this which can have important management implications,” he said.

“We hope that the PEA might someday permit hospitals to perform the assay ‘in-house’ while eliminating the need for send-out laboratory testing in patients with possible HIT. ”

—Anand Padmanabhan, MD, PhD

Because the PEA requires fewer resources than the SRA, Dr. Padmanabhan indicated that this method may offer an easier option to diagnose HIT. “The currently available laboratory testing for HIT either lacks specificity or is technically difficult to perform, often requiring a several-day delay between sample collection and final confirmation or exclusion of HIT,” he said. “While we cannot be certain about the impact of our findings on the clinical care of patients with suspected HIT, we hope that the characteristics of the PEA might someday permit some hospitals to perform the assay ‘in-house’ while eliminating the need for send-out laboratory testing in patients with possible HIT.”

In terms of limitations, Dr. Padmanabhan stated that the investigators lacked a “gold standard” assay for which they could have absolutely confirmed or refuted an HIT diagnosis. The reliance on clinical and laboratory criteria to classify patients as HIT-positive and HIT-negative represented an additional limitation. Dr. Padmanabhan suggested this could have led to misclassification, as some patients who were initially classified as disease-negative were later considered to have HIT on follow-up investigation.

Study authors report no relevant conflicts of interest.


Samuelson Bannow BT, Warad D, Jones C, et al. A prospective, blinded study of a PF4-dependent assay for HIT diagnosis. Blood. 2020 September 8. [Epub ahead of print]