An Enzyme-Linked Immunosorbent Assay for the Detection of Aspergillus parasiticus and Aspergillus flavus
Abstract:An enzyme-linked immunosorbent assay (ELISA) was established for the specific detection of Aspergillus parasiticus and Aspergillus flavus. A New Zealand white rabbit was immunized intravenously with 100 μg of A. parasiticus CCRC 30117 mycelial protein extracts. The antibodies were separated and purified. The optimal concentration of the antibody and antibody-peroxidase conjugate used in the established ELISA was 10 μg/ml with a detection limit of 1 μg/ml. Among the 126 strains tested (including 21 strains of A. parasiticus, 11 strains of A. flavus, 34 isolates of A. parasiticus/A. flavus from cereals, and 60 strains of non-A. parasiticus/A. flavus fungi), the false-negative and false-positive rates were 1.5 and 3.3%, respectively. Strains of Aspergillus flavofrucatis and Aspergillus sojae produced false-positive reactions. However, their antigens had much lower cross-reactivity with the antibodies raised against A. parasiticus, as shown from I50 values. The molecular weights of the main antigens of A. parasiticus were 94, 82, and 40 kDa. The two heavier antigens had higher sugar contents, as demonstrated by SDS-PAGE and immunoblotting. A good correlation (r = 0.97) was found between mycelium measurement by weighing and by ELISA for A. parasiticus grown in yeast extract sucrose broth (YESB) at 25°C.
Document Type: Research Article
Affiliations: Department of Marine Food Science, National Taiwan Ocean University, Keelung, Taiwan 202, ROC
Publication date: August 1, 1997
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