Colorimetric and fluorometric substrate immunoassays for detection of Salmonella in all foods: comparative study.

Two modified fluorescent enzyme immunoassays for the detection of Salmonella in food have been developed. Both of the new procedures, which substitute a colorimetric substrate for the fluorescent substrate and in which results are read visually or with a photometer, are modifications of AOAC method 989.15. The visually read procedure uses the same antibody-coated wells as in method 989.15. The colorimetric end point of the assay is determined by comparing the solution color to a color chart. The assay result may also be read in a photometer, if the solution is first transferred to a transparent microtiter well. The second procedure designed to be read in a photometer substitutes clear, antibody-coated wells for those used in the fluorescent assay. The colorimetric assays employ identical monoclonal antibodies for capture and detection of Salmonella as used in the fluorescent assay. In this comparative study, the performance of each new assay was consistent with the performance of method 989.15. These methods have been adopted official first action by AOAC as alternative methods for the detection of Salmonella in foods.

Colorimetric deoxyribonucleic acid hybridization assay for rapid screening of Salmonella in foods: collaborative study.

A collaborative study was performed in 11 laboratories to validate a colorimetric DNA hybridization (DNAH) method for rapid detection of Salmonella in foods. The method was compared to the standard culture method for detection of Salmonella in nonfat dry milk, milk chocolate, soy isolate, dried whole egg, ground black pepper, and raw ground turkey. Samples inoculated with high (0.4-2 cells/g) and low (0.04-0.2 cells/g) levels of Salmonella and uninoculated control samples were included in each food group analyzed. There was no significant difference in the proportion of samples positive by DNAH and culture procedure for any of the 6 foods. The colorimetric DNA hybridization assay screening method has been adopted official first action as a rapid screening method for detection of Salmonella in all foods.

Comparison of colorimetric monoclonal enzyme immunoassay screening methods for detection of Salmonella in foods.

A colorimetric enzyme immunoassay (EIA) method for detection of Salmonella in foods has been compared to the AOAC colorimetric monoclonal EIA screening method (986.35, 15th ed.; 46.B21-46.B29, 14th ed.). The assays use the same monoclonal antibodies and have similar reactivity toward Salmonella. However, the new assay uses antibody-coated microtiter wells instead of coated magnetic beads to capture Salmonella antigens. Compared with the bead assay, the coated-well assay format requires significantly less time to complete, and was consistently able to detect lower levels of Salmonella in mixed culture. Compared to the standard AOAC culture method for food samples, the plate assay was as productive. No false negatives were obtained by the immunoassay; the false negative rate was 1.1% by the culture method. The rate of agreement between the 2 methods was 99.1%. The official final action bead assay method for Salmonella in foods, 986.35, and the same assay for use with low-moisture foods, 987.11, have been modified official first action to use antibody-coated microtiter strip-wells.

Immunodiffusion screening method for detection of motile Salmonella in foods: collaborative study.

A collaborative study was performed to validate the performance of the 1-2 TEST for detection of motile salmonellae in foods. Detection is based on observation of an immobilized band of cells. Twenty-three laboratories participated in the study. The 1-2 TEST (immunodiffusion test) was compared with the standard culture procedure (BAM/AOAC; FDA Bacteriological Analytical Manual) for detection of Salmonella in 6 food types: ground black pepper, soy flour, dried whole egg, milk chocolate, nonfat dry milk, and raw deboned turkey. Uninoculated and inoculated samples were included in each food group analyzed. After the tests on the 6 foods were completed, analysis of the data for turkey and soy flour showed that certain collaborators obtained data inconsistent with the data from the majority of collaborators. No specific method deviations to account for the inconsistencies were reported by those collaborators, so the collaborative testing of these 2 foods was repeated. Analysis of data for pepper, chocolate, nonfat dry milk, dried whole egg, and the second set of soy flour and turkey indicated 96.1% agreement between the BAM/AOAC and immunodiffusion test methods. The false negative rates for the immunodiffusion test and BAM/AOAC methods were 3.6 and 1.7%, respectively. There was no significant difference in the productivity of the immunodiffusion test and BAM/AOAC method at the 5% level for any of the 6 foods. The immunodiffusion screening method has been approved official first action for detection of motile Salmonella in foods.

Fluorescent enzyme immunoassay for rapid screening of Salmonella in foods: collaborative study.

A collaborative study was performed in 13 laboratories to validate an enzyme immunoassay (EIA) procedure for rapid detection of Salmonella in foods. The EIA was compared with the standard culture procedure for detection of Salmonella in 6 food types: ground black pepper, soy flour, dried whole eggs, milk chocolate, nonfat dry milk, and raw deboned turkey. Uninoculated and inoculated samples were included in each food group analyzed. There was no significant difference in the proportion of samples positive by the EIA and culture procedures at the 5% level for any of the 6 foods. The enzyme immunoassay screening method has been adopted official first action as a rapid screening method for detection of Salmonella.

Visual immunoassay for detection of Salmonella in foods: collaborative study.

A collaborative study was performed in 13 laboratories to validate a visual enzyme immunoassay (EIA) procedure, TECRA, for rapid detection of Salmonella in foods. The EIA method was compared with the standard culture procedure for detection of Salmonella in 6 food types: ground black pepper, soy flour, dried whole eggs, milk chocolate, nonfat dry milk, and raw deboned turkey. Uninoculated and inoculated samples were included in each food group analyzed. There was no significant difference in the productivity of the EIA and culture procedures at the 5% level for any of the 6 foods. The enzyme immunoassay screening method has been approved interim official first action.

Evaluation of abbreviated enzyme immunoassay method for detection of Salmonella in low-moisture foods.

A modified enzyme immunoassay method (EIA) utilizing an 18 h pre-enrichment, a 6-8 h selective enrichment, and a 14 h M-broth post-enrichment is compared to the standard culture method (AOAC/BAM) on selected low-moisture foods. Tested samples included 238 inoculated, 30 naturally contaminated, and 30 uninoculated foods. By EIA, 235 samples were positive (optical densities greater than 0.2 at 405 nm), 233 of which were confirmed culturally. By the culture methods, 221 samples were positive. The EIA method was more productive in detecting salmonellae in inoculated samples of dry cheese powder, chocolate, and nonfat dry milk, whereas the culture method gave better recovery from naturally contaminated meat and bone meal. The modified EIA could be completed in 40 h and required no centrifugation.

DNA hybridization assay for detection of Salmonella in foods: collaborative study.

A collaborative study was performed in 11 laboratories to validate a DNA hybridization (DNAH) procedure for detection of Salmonella in foods. The DNAH procedure was compared to the standard culture method for detection of Salmonella in 6 foods: ground pepper, soy flour, dry whole egg, milk chocolate, nonfat dry milk, and raw deboned turkey. With the exception of turkey which was naturally contaminated, uninoculated and inoculated samples of each food group were analyzed. Results for the DNAH method were significantly better than for the standard culture method at the 5% probability level for the detection of Salmonella in turkey. There was no significant difference between the methods for the other 5 foods. The method has been adopted official first action.

Enzyme immunoassay for detection of Salmonella in low-moisture foods: collaborative study.

A collaborative study was performed in 15 laboratories to evaluate a modification of the enzyme immunoassay (EIA) method for detection of Salmonella in foods (46.B21-46.B29). The modified EIA requires 18-24 h pre-enrichment, 6-8 h selective enrichment, and 14-18 h M-broth post-enrichment prior to performing the assay, which requires 1-2 h. Total assay time is 40-52 h. The modified method was compared with the standard culture method for detection of Salmonella in 5 low-moisture foods: nonfat dry milk, milk chocolate, meat and bone meal, dry whole egg, and ground pepper. The modified method has been adopted official first action for use with low-moisture foods.