Monoclonal antibody anti-AFB1: scale-up in vitro for biotools development

Abstract

Aflatoxin B₁ (AFB₁) is a mycotoxin classified as group 1 (human carcinogen) by International Agency for Research on Cancer - IARC, causing hazardous contamination in a wide variety of food and feed, where the monitoring depends on precision and accuracy of analytical method. The culture of AFB₁ specific monoclonal antibody (mAb) secreting hybridoma was performed for further development of immunochemical methods. The growth of hybridoma AF2 was carried out in RPMI medium + 15 % fetal bovine serum (FBS), as well as the same medium gradually amended with H-SFM medium (25, 50, 75 and 100 % H-SFM). The protein concentration in the culture supernatant ranged from 1.80 to 10.88 mg/mL. The culture amended with FBS-free synthetic H-SFM medium reached production of reagent with higher degree of purity and lower risk, in addition to lower protein content (2.29 mg/mL reached with 100 % H-SFM), which approaches the real content of pure mAb. The indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and SDS-polyacrylamide gel electrophoresis (SDS-PAGE) showed anti-AFB₁ activity and IgG corresponding bands, respectively, indicating feasible application of mAb produced in 100, 75 and 50 % H-SFM for further use in the development of AFB₁ detecting biotools. This mAb production can be an initial step that can supply the self-sufficient immune-reagent in the rapid diagnosis at national condition, which is essential in the food quality and safety.