The Purification Step is not Crucial in EIA Measurements of Thromboxane B2 and 11-Dehydrothromboxane B2 in Human Plasma

Background: Thromboxane B-2 (TxB(2)) and particularly 11-dehydrothromboxane B-2 (11-dTXB(2)) are widely Used as prognostic risk markers of platelet activation in cardiovascular diseases. The main errors in TxB(2) and 11-dTxB(2) determination include either low concentrations of circulating TxB(2) (1 - 2 pg/mL) and 11-dTxB(2) (0.9 - 4.3 pg/mL) or rather high transiency (mean TxB(2) half-life is approximately 5 minutes) as well as an incorrect pre-analytical phase set up. The aim of this study was to investigate the impact of a widely used purification step on the results of enzyme immunosorbent assay (EIA) - based measurement of the two selected thromboxanes. Methods: For the purpose of this study, 20 plasma samples (10 healthy donors, 10 patients under treatment with acetylsalicylic acid) were screened for TxB(2) and 11-dTxB(2) concentrations using commercial competitive EIA kits (Cayman Chemicals (TM), Tallinn, Estonia; Neogen (TM), Lexington, KY, USA) with or without the introduction of the purification procedure. Results: The purification step does not significantly affect the results of EIA measurements of the two of TxA(2) metabolites (TxB(2), 11-dTxB(2)) in human plasma. The levels of TxB(2) and 11-dTxB(2) determined in the plasma samples were not significantly changed (p<0.05) when the purification step was omitted compared to the purified samples. Conclusions: This study establishes a protocol allowing for reliable and reproducible plasma TxB(2) and 11-dTxB(2) EIA measurement for routine basic screening of platelet function. (Clin. Lab. 2012;58:177-183)