Development of ELISA and enzyme-linked immunofiltration assay (ELIFA) methods for monitoring cyclodextrin glycosyltransferase (CGTase) production and bacterial growth in Bacillus macerans batch cultures.

Immunochemical methods were developed for monitoring cyclodextrin (CD) glycosyltransferase (CGTase) production and growth of an industrial CD-producing Bacillus macerans strain. Extracellular concentrations of CGTase released into a non-transparent culture medium during a 44 h long fermentation were detected by an indirect antigen inhibition enzyme-linked immunosorbent assay (ELISA). The ELISA was sensitive (minimal detection level 6 ng ml-1) and highly reproducible (coefficients of variation < or = 1.

Amplified enzyme-linked-immunofilter assays enable detection of 50-10(5) bacterial cells within 1 hour.

Two enhanced enzyme-linked-immunofilter assay (ELIFA) methods for the rapid and quantitative detection of whole bacterial cells are described. In the first method, specific antibody bound to bacterial cells was amplified using a secondary antibody and detected by the conjugated enzyme activity (peroxidase) of a third antibody in a chemiluminescent assay. In the second method, a chromogenic substrate was used in conjunction with a biotinylated secondary antibody and avidin.

A rapid and sensitive enzyme linked immunofilter assay (ELIFA) for whole bacterial cells.

An improved method is described for the detection of Escherichia coli by an enzyme linked immunofilter assay (ELIFA) using nitrocellulose membrane sandwiched between two 96-well plates. The incorporation of a pumping system permits a continuous flow of reagents and/or wash fluids through the membrane and provides an assay procedure capable of detecting 10(3) bacteria per well within 40 min. Quantitative bacterial detection was based on precipitated chromogen determined by scanning densitometry.