Heparin Neutralization for Thrombin Generation

Samples sent to the laboratory for HIT diagnosis work up often contained heparin because blood was collected early, as soon as HIT was suspected and TG is very sensitive to presence of heparin in the plasma sample [20 (link), 21 (link)]. Consequently, we had to perform the assay after neutralization of heparin. For this, we considered two reagents: polybrene (hexadimethrine bromide, Sigma-Aldrich, France) at concentrations described elsewhere [22 (link)] and heparinase (Dade Hepzyme®, Siemens Healthcare Diagnostics, Marburg Germany), using one vial of product for 1mL of plasma, according to manufacturer’s instructions. In preliminary experiments, plasmas from two healthy controls and five non-HIT patients under unfractionated heparin (UFH) and low molecular weight heparin (LMWH) were incubated with polybrene or Dade Hepzyme® at room temperature for 15 min before checking neutralization efficiency as follows. Coagulation tests (APTT, PT) and chromogenic anti-Xa assay (STA®-Liquid Anti-Xa, Diagnostica Stago, France) were performed with a STAR® Max analyzer before and after heparin neutralization. As some residual anti-Xa levels (0.10–0.20 IU/mL) were observed in polybrene-treated samples, we performed the study treating each individual patient’s sample with Dade Hepzyme® before TG study.