Pitfalls in the determination of human acylated ghrelin plasma concentrations using a double antibody enzyme immunometric assay.

To investigate the effect of hemolysis and protease inhibitors on acylated ghrelin (AG) concentrations measured with a double antibody enzyme immunometric assay that uses an acetylcholinesterase (AChE)-Fab' conjugate.Samples were hemolysed or treated with PHMB (p-hydroxymercuribenzoate), PMSF (phenylmethanesulfonylfluoride) or AEBSF (4-(2-Aminoethyl) benzenesulfonyl fluoride) to prevent AG degradation.Hemolysis decreased AG concentrations. PHMB or PMSF did not affect the assay. The standard curve was abolished by AEBSF but rescued by addition of a washing step prior to the AChE-Fab' conjugate.Hemolysis and AEBSF may affect AG determination.