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

Cássia Reika Takabayashi, Luciana Hayashi, Tatiane Martins de Oliveira, Simone Fujii, Carlos Kemmelmeier, Elisabete Yurie Sataque Ono, Eiko Nakagawa Itano, Osamu Kawamura, Elisa Yoko Hirooka

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.


Keywords


Monoclonal antibody; Hybridoma; Biotool; AFB



DOI: http://dx.doi.org/10.5433/1679-0359.2010v31n4p933

Semina: Ciênc. Agrár.
Londrina - PR
E-ISSN 1679-0359
DOI: 10.5433/1679-0359
E-mail: semina.agrarias@uel.br
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