Neutralization of Heparin in Plasma by Platelet Factor 4 and Protamine Sulphate

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Abstract
The neutralization of heparin by protamine sulphate and platelet factor 4 (PF4) has been studied using kaolin cephalin clotting time (KCCT), thrombin clotting time (TCT) and anti‐Xa assays to measure residual heparin levels. Protamine sulphate and purified PF4 had almost equivalent neutralizing ability on a weight basis regardless of which assay system was used. Plasma samples that were heparinized in vitro could be totally and readily neutralized by both agents in all of the assay systems used. However, when heparinized plasma samples were obtained following intravenous injection of the drug different results for neutralization were obtained depending on the heparin assay used. When residual heparin was measured by anti‐Xa assay partial neutralization of heparin was observed even in the presence of a large excess of neutralizing agent. The same in vivo derived heparinized plasma samples had no heparin activity following neutralization if residual heparin was measured by KCCT and TCT assays. Further ion exchange chromatography experiments demonstrated that the heparin‐like activity that could not be neutralized in the anti‐Xa assay was not adsorbed by the resin ECTEOLA‐cellulose, and therefore could not be removed from plasma by this technique. These results suggest, therefore, that part of the plasma anti‐Xa activity produced following intravenous injection of heparin differs from the anti‐Xa activity obtained following addition of the drug to plasma in vitro.