Influence of Endogenous Factors of Food Matrices on Avidin—Biotin Immunoassays for the Detection of Bacitracin and Colistin in Food

Abstract

(Strept)avidin–biotin technology is frequently used in immunoassay systems to improve their analytical properties. It is known from clinical practice that many (strept)avidin–biotin-based tests provide false results when analyzing patient samples with a high content of endogenous biotin. No specific investigation has been carried out regarding possible interferences from avidin (AVI) and biotin (B7) contained in food matrices in (strept)avidin–biotin-based immunoanalytical systems for food safety. Two kinds of competitive ELISAs for bacitracin (BT) and colistin (COL) determination in food matrices were developed based on conventional hapten–protein coating conjugates and biotinylated BT and COL bound to immobilized streptavidin (SAV). Coating SAV–B7–BT and SAV–B7–COL complexes-based ELISAs provided 2- and 15-times better sensitivity in BT and COL determination, corresponding to 0.6 and 0.3 ng/mL, respectively. Simultaneously with the determination of the main analytes, these kinds of tests were used as competitive assays for the assessment of AVI or B7 content up to 10 and 1 ng/mL, respectively, in food matrices (egg, infant milk formulas enriched with B7, chicken and beef liver). Matrix-free experiments with AVI/B7-enriched solutions showed distortion of the standard curves, indicating that these ingredients interfere with the adequate quantification of analytes. Summarizing the experience of the present study, it is recommended to avoid immunoassays based on avidin–biotin interactions when analyzing biosamples containing these endogenous factors or enriched with B7.

Oral delivery of glutathione: antioxidant function, barriers and strategies

“…Insulin is well protected from proteolytic degradation (triggered by trypsin and chymotrypsin) when encapsulated with these enzyme inhibitors, as compared with unadulterated insulin solutions, in vitro [110]. Bacitracin, a cyclic polypeptide antibiotic with a molecular weight of 1422.7 Da, is another enzyme inhibitor that effectively inhibits various proteases, including trypsin, pepsin and aminopeptidase [111]. An in vitro study has reported that bacitracin, camostat mesilate and sodium glycocholate decrease insulin degradation in rat large intestinal homogenate [112].…”

Application of Antibody and Immunoassay for Food Safety

“…Assessing the matrix interference on the assay sensitivity is thus an important issue in the development of methods. Burkin et al [ 3 ] firstly evaluated the influence of avidin (AVI) and biotin (B7) contained in food matrices on two kinds of (Strept)avidin–biotin-based enzyme-linked immunosorbent assays (ELISAs) for bacitracin (BT) and colistin (COL) determination, with simultaneous assessment of extraneous AVI/B7 and AVI/B7 from different matrices (egg, infant milk formulas enriched with B7, and chicken and beef liver). Summarizing the experience of the present study, it is recommended to avoid immunoassays based on avidin–biotin interactions when analyzing samples containing these endogenous factors or enriched with B7.…”