A Direct Competitive Immunoassay for Fumonisins Detection in Maize Feed
Kadir*, M.K.A., Ishak, Z. and Awaludin, N.
Biotechnology Research Centre, Malaysian Agricultural Research and Development Institute, Serdang, Selangor
Malaysian J. Anim. Sci. 2013 16(1): 23-38
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The development of a direct competitive immunoassay formats for the Enzyme-linked immunosorbent assay (ELISA) method was undertaken for fumonisins (Fms) determination in feed. The use of polyclonal antibody was conducted on the microtitre plate for a rapid and sensitive detection. The direct format assay was based on the competition between Fms labelled with houseradish peroxidase (HRP) and unlabelled Fms for binding sites first of immobilised antibody (AbFms) followed by precoating of secondary anti-antibody IgG (Anti-IgG). A spectrophotometric assay was developed as a first step procedure for optimization of concentrations and conditions using micro-titre plates. Optimal parameters established for direct assay were 20 ?g mL-1 of anti-IgG, 1:50 dilution of AbFms and 1:5 dilution of Fumonisins-HRP. The ELISA exhibited detection limit of 100 ?g?L?1 fumonisins with a dynamic range from 100 - 2000 ?g?L?1. The achieved detection range for Fms was within the required legislative limit of analyses. Samples analysis involved the rapid extraction (without clean-up) and pre-treatment using solid phase extraction (clean-up) before measuring using the developed ELISA. The results achieved were average of 97.6 % (clean-up extraction) and 77.9 % (without cleanup). The developed ELISA had a satisfactory agreement with HPLC (Confirmatory method). It also showed very high sensitivity and a potential method for a rapid, simple and sensitive detection of Fms in maize feed.
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