Preparation and evaluation of an immunoaffinity sorbent with Fab′ antibody fragments for the analysis of opioid peptides by on-line immunoaffinity solid-phase extraction capillary electrophoresis–mass spectrometry

Abstract

An immunoaffinity (IA) sorbent with antibody fragments was prepared for the analysis of opioid peptides by on-line immunoaffinity solid-phase extraction capillary electrophoresis–mass spectrometry (IA-SPE-CE–MS). The antibody fragmentation was evaluated by MALDI-TOF-MS. Fab′ fragments obtained from a polyclonal IgG antibody against Endomorphins 1 and 2 (End1 and End2) were covalently attached to succinimidyl silica particles to prepare the IA sorbent. An IA-SPE-CE–MS methodology was established analyzing standard solutions of End1 and End2 and acceptable repeatability, linearity ranges and LODs (0.5 and 5ngmL−1, respectively) were obtained. The LOD of End1 was slightly better than that previously obtained using an IA sorbent with intact antibodies (1ngmL−1). In human plasma samples, End1 and End2 could be detected at 1 and 50ngmL−1, respectively, which meant an improvement of 100 and 2-fold with regard to the LODs using an IA sorbent with intact antibodies (100ngmL−1).