Abstract
The non-dialyzable fraction of culture filtrate antigen of Aspergillus fumigatus was fractionated by anion exchange chromatography and gel filtration, connecting each column of Mono Q and Superose 6 sequentially. Fractions which had a high protein and reacted strongly with an immunoglobulin-M monoclonal antibody (MAb) 8-3C F5 G9, as determined by enzyme-linked immunosorbent assay (ELISA), were pooled to give four antigenic fractions (F1-F4). Gas-liquid chromatography analysis of the 4 antigenic fractions showed the presence of mannose and galactose. An immunoelectron microscopic experiment with MAb revealed the distribution of antigen on the surface of the hyphae. The fractions had no proteolytic activity. Fraction 4, relatively small in its molecular size but with the highest recovery of the reactivity with MAb among F1 to F4, was selected for further characterization. Antigenic reactivity of F4 with MAb was heat-stable but fully retained after sodium metaperiodate treatment. Pronase E almost destroyed the antigenic reactivity of F4 with MAb. MAb affinity chromatography of F4 gave the specific antigen, F4-AF. In both sodium dodecyl sulfate-polyacrylamide gel electrophoresis by Coomassie blue staining and Western blotting with MAb, F4-AF showed a single band with molecular mass of 66 kDa.
