ASH recommends that positive immunoassay results should be confirmed by a functional assay , with the serotonin release assay (SRA) being the gold standard. Generally, the optical density of ELISA-based methods or arbitrary concentration units of other immunoassay formats correlate with SRA positivity; that is, the higher the immunoassay result the more likely that the SRA will yield a positive result . Consequently, ASH guidelines state that in patients with a high 4T score and a strongly positive immunoassay, a functional assay may not be necessary for diagnosis . Serotonin is released from platelet dense granules upon platelet activation, and the percentage release of serotonin can be used to quantify the magnitude of platelet activation induced by HIT antibodies. In brief, platelet rich plasma (PRP) is prepared from pedigree blood donors whose platelets are known to release high amounts of serotonin. The PRP is incubated with 14C-serotonin and heat inactivated patient serum is then added, followed by a low and high dose of unfractionated heparin (for example, low dose: 0.1–0.3 U/mL and high dose: 100 U/mL). The SRA is considered negative if less than or equal to 20% release is observed. In order for the test to be positive, the low dose needs to display more than 20% release, while the high dose displays at least a 50% reduction compared to the low dose. High-dose heparin causes inhibition of platelet activation in the SRA, as it imbalances the stoichiometry required for heparin-antigen formation. Very few laboratories perform functional assays for HIT diagnosis, as the assays are lab-developed and labour-intensive. These assays offer high sensitivity and a higher specificity for HIT compared to immunoassays. False positive SRA results have been reported , and can be mitigated through reflexive testing algorithms in which only antibody-positive specimens are tested by SRA or the SRA and immunoassay testing is performed concurrently.
Management of patients with HIT
Once a HIT diagnosis is confirmed, the patient is labelled as having acute HIT. Management of acute HIT involves discontinuation of heparin and initiation of a non-heparin anticoagulant. Heparin discontinuation is not sufficient to eliminate the risk for thrombosis in patients with HIT, as the 30-day risk for thrombosis after heparin withdrawal is estimated to be between 19 and 52% . HIT therapy should be individualized, depending on the type of patient, kidney and liver function, likelihood of additional procedures and bleeding risk. Direct thrombin inhibitors (bivalirudin, argatroban, dabigatran), indirect factor Xa inhibitors (danaparoid, fondaparinux) and direct factor Xa inhibitors (apixaban, rivaroxaban) have been recommended for management of acute HIT . Complete platelet recovery is seen in approximately 65% of patients within one week of heparin cessation, although thrombotic risk remains high for 4 to 6 weeks after diagnosis. Patients may be monitored by HIT immunoassays to determine when antibodies are no longer present. The median time to HIT seronegativity is 85–90 days .
Current challenges and future considerations
Early diagnosis and intervention is critical in mitigating morbidity and mortality associated with HIT. As previously mentioned, calculation of the 4T score can be challenging, which is further compounded by the fact that thrombocytopenia occurs frequently in hospitalized, heparin-treated patients. Inaccurate estimation of the 4T score can lead to inappropriate HIT testing in patients with a low pre-test probability for HIT and an increase in the number of false positive results.
Another key challenge in HIT diagnosis is that only a small fraction of seropositive patients develops clinically significant HIT. Since functional assays are most often performed off-site, future immunoassays should focus on improved specificity for HIT. It is encouraging that the degree of immunoassay test positivity correlates with functional assay-positivity; however, further multicentre, assayspecific studies are required to evaluate the reliability of this measure alone or in combination with the 4T score in the prediction of HIT. Lastly, variation in the performance characteristics of the same functional assay format may exist between labs, since these assays are lab-developed. Since diagnosis hinges heavily on the functional assay result, it is critical that clinicians are familiar with the performance characteristics of the functional assay used at their institute and consider the entire clinical picture in making a diagnosis.