Fluorescent Antibody Method for the Detection of Salmonella: Development, Evaluation, and Collaborative Study

Abstract

In preliminary studies, several commercial polyvalent fluorescent antibody (FA) preparations were evaluated for specificity and crossreactivity and an FA method was developed for the screening of Salmonella in products. Approximately 4000 product samples were tested by the FA method and the results were compared to those from the official final action AOAC method, 46.013–46.026. Only 4 FA falsenegatives were found for a total of 619 confirmed positive Salmonella samples. The FA false-positive rate was 7%. The method was then subjected to a 2-phase collaborative study. In Phase I, 22 analysts tested 5 inoculated and 5 uninoculated samples of dried milk. In Phase II, 5 naturally contaminated and 5 presumably uncontaminated foods were analyzed. The study was designed to compare results from 11 analysts experienced in FA methodology with those from 11 analysts with little or no experience. Selenite cystine (SC) and tetrathionate (TT) broths were used for enrichment and both were inoculated into SC for post-enrichment. All 4 combinations (SC, TT, SC-SC, and TT-SC) were used with the FA method to determine the best technique. Results were compared to the analysis with TT and SC by the AOAC culture method. In all studies, FA analysis with SC-SC gave the highest correlation with the AOAC method. In a total of 200 samples, the experienced group found 125 AOAC positives and 127 FA positives ; no FA false-negatives and only 2 falsepositives were reported. The inexperienced group reported 9 FA false-negatives and 5 FA false-positives. All false-negatives occurred in only 3 of the inexperienced laboratories. These studies showed that enrichment and post-enrichment in SC gave the best FA results and that training in FA methodology is required for correlation with existing AOAC methodology. The FA method for the detection of Salmonella has been adopted as official first action.

A Comparison of the Fluorescent Antibody Method and a Standardized Cultural Method for the Detection of Salmonellas

“…The results reported here show that the proposed draft standard I S 0 technique for the isolation of salmonellas from foods and feedingstuffs may also be used for the FA technique at the pre-enrichment stage. No modifications, such as culture in a nonselective broth after selective enrichment ('post-enrichment')-which appears to be desirable to eliminate quenching of fluorescence by selenite or tetrathionate (Thomason & Wells 1971;Insalata et al 1972;Smyser & Snoeyenbos 1976)-are necessary although Fantasia et al (1975) recommend selenite-cystine broth for successive enrichment cultures before FA slide preparation. Application of the FA technique at an early stage in the culture method for isolation of salmonellas gave a number of false positive FA results, although these may have been false negative cultural results.…”

“…1975) based on that described by Edel & Kampelmacher (1973, 1974, both include pre-enrichment in a non-selective medium. The official first action FA method (Fantasia et al 1975), recommends, after pre-enrichment, two successive selective enrichment cultures in selenite-cystine broth, one of 18-24 h and the other of 4 h duration, before preparing FA slides. These successive selective enrichment cultures undoubtedly reduce the number of false FA positive and negative results obtained, but extend the time required for a screening result.…”

Evaluation of an automated fluorescent antibody procedure for detection of Salmonella in foods and feeds

“…Fluorescent antibody (FA) procedures (3,5,6,7,9,11,13,14) offer considerable time savings and have been gaining acceptance for screening samples for salmonellae. Recently a direct FA procedure was approved by AOAC on the basis of results found in a collaborative study in 22 laboratories (2,4). In our laboratory this manual FA method was used to test about 4,000 product samples, and results (4) were highly comparable with the AOAC cultural method.…”

Timed-Release Capsule Method for the Detection of Salmonellae in Foods and Feeds

“…Much research has been conducted to develop alternative methods that are as accurate as existing cultural procedures but allow more rapid recognition of contamination. Enrichment-serology (1,8,12,19) and fluorescent-antibody (3,7,10,13) techniques speed detection of salmonellae but require special equipment and antisera.…”