PB1113 - Influence of clotting trigger for functional protein S measurement

Background: Commercially available functional activity assays of protein S are clot based and incorporate activated protein C, phospholipid and a trigger reagent which could be tissue factor (Werfen), or Russell’s viper venom (Siemens) or factor Va (Stago). It was noted in recent NEQAS reports (surveys 1254 & 1255) that these three commercial methods used gave some differences in protein S determinations (table 1). These differences may be due to the calibrators used and/or the assays kits that employed different clotting triggers.

Aims: To investigate whether the method of clotting trigger influences the functional potency estimation of protein S.

Methods: An in-house protein S clotting method was devised where the trigger reagent was either a contact activator (kaolin) or tissue factor. Four commercial calibrators were tested against the International Standard for Protein S, with both methods and values compared to the quoted activity.

Results: For all samples activation by tissue factor gave a higher potency than by kaolin (see table 2). Whilst all measured activities were lower than quoted, three (Werfen, Siemens and Lot 5) were within 10% and the remaining two within 20% by tissue factor trigger.

Conclusion(s): Different activities were obtained when using different triggers. Recovery of functional protein S was consistently lower than quoted values. Except for the Siemens calibrator which gave similar values by either trigger, all other calibrators’ estimates were higher by tissue factor activation. Demonstrating TF activation can give higher estimates concurs with NEQAS results showing higher results by this method. Further investigation using different triggers is warranted to identify the mechanism of discrepancy.