Development of rapid detection method of viable Escherichia coli and coli-form

1996 Fiscal Year Final Research Report Summary

• Project/Area Number: 07660168
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: 食品科学・栄養科学
• Research Institution: KYUSHU UNIVERSITY
• Principal Investigator: MIYAMOTO Takahisa Kyushu Univ.Food Sci.& Technol. Associate Prof., 農学部, 助教授 (70190816)
• Co-Investigator(Kenkyū-buntansha): HONJOH Ken-ichi Kyushu Univ.Food Sci.& Technol. Assistant Prof., 農学部, 助手 (00264101) ; HATANO Shoji Kyushu Univ.Food Sci.& Technol. Full Prof., 農学部, 教授 (30038260)
• Project Period (FY): 1995 – 1996
• Keywords: Escherichia coli / coli-form / monoclonal antibody / Rapid detection

Research Abstract

Three anti-E.coli monoclonal antibodies were prepared. Among them, MAb-C was most specific to E.coli. The MAb did not react with other bacteria such as Enterobacter aerogenes, Klebsiella pneumoniae, Proteus vulgaris, Serratia marcescens, and Salmonella typhimurium. As a preliminary experiment to develop rapid detection of viable cells, ATP assay was performed on E.coli harvested on a membrane filter. By measuring light emission, about 10^4 CFU of E.coli was detected. The same membrane was treated with peroxidase-labeled MAb-C and light emission was measured after reaction with luminol solution (MAb assay). About 10^3 CFU of E.coli were detected with the Mab assay. When the membrane was incubated in Nutrient broth for 6 hr at 35ﾟC , E.coli was detected at 1 CFU by both methods. Initial cell nember and total light emission counts or total light emission points on the filter was strongly correlated. Correlation coefficients were calculated to be 0.95 and 0.89, respectively, for total light emission counts and total light emission points. In the case of MAb assay, correlation was not so strong between initial cell number and total light emission points (r=0,76). Further work is under way to obtain more specific MAb to E.coli.