Rapid enumeration of Staphylococcus aureus in foods by direct demonstration of enterotoxigenic colonies on membrane filters by enzyme immunoassay

Abstract

Based on enzyme-linked immunosorbent assay, a convenient method has been devised for the direct demonstration of enterotoxin B production by Staphylococcus aureus colonies grown for 24 h on membrane filters. The problem of false-positive reactions due to binding of immunoglobulin G to protein A was turned to advantage by conjugating horseradish peroxidase directly to protein A, which then mediated the labeling of the antitoxin. The test requires 3 h to complete and yields a purple stain at the site of enterotoxin Bproducing colonies, thus allowing direct enumeration of confirmed S. aureus in foods within 27 h. The method should be applicable to other enterotoxins of S. aureus.

Improved immunological membrane filter method for detection of food-borne Salmonella strains

“…In recent years, many approaches have been made toward automated analytical food microbiology, including the use of HGMF for detection and enumeration of microorganisms (7,15,17). The present study shows the applicability of the ELA stain method performed directly on HGMF for the demonstration of Salmonella strains in foods.…”

“…Two previous studies of the ability of the ELA stain method to detect microorganisms on HGMF (7,15) involved the use of a blotting step in which colony material was transferred to a nitrocellulose membrane for staining. When ELA staining was carried out directly on the HGMF, results varied widely.…”

“…When ELA staining was carried out directly on the HGMF, results varied widely. No staining, staining in only a narrow ring around colonies, and completely unstained colonies on a stained background were all observed (7,15). At the time it was believed that metabolic products released from colonies during the long incubation interfered with stain reaction and that blotting (which transfers only the tops of colonies) was more efficient because of the uncontaminated substrate introduced by the process.…”

“…These methods require a blotting step to transfer colony material to nitrocellulose, which binds proteins much more strongly than does the polysulfone polymer from which HGMFs are made (15). However, the computerized count-* Corresponding author.…”

Detection of Listeria monocytogenes by direct colony hybridization on hydrophobic grid-membrane filters by using a chromogen-labeled DNA probe

“…Supporting this system, HGMF-based methods that use enzyme-labeled polyclonal and monoclonal antibodies have been developed for the detection of Salmonella spp. (1), enterotoxigenic Staphylococcus aureus (19), and Escherichia coli 0157 (24) in foods. Methods that use monoclonal antibodies and DNA probes for bacterial detection are specific and do not require time-consuming biochemical testing for confirmed identification of the organism (13).…”