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The Journal of the Japanese Society for Clinical Microbiology |
Biblioraphy Information
| ArticleTitle |
Improvement of HEp-2 cell vacuolation assay for Bacillus cereus emetic toxin |
| Language |
J |
| AuthorList |
Kumiko Kawamura1), Yumi Hirama1), Norio Agata2), Hideo Ito1) |
| Affiliation |
1) Department of Medical Technology Nagoya University School of Health Science
2) Nagoya City Public Health Research Institute |
| Publication |
J.J.C.M.: 14 (3), 146-152, 2004 |
| Received |
June 17, 2004 |
| Accepted |
August 10, 2004 |
| Abstract |
Cereulide is the causative toxin of emetic type food poisoning by Bacillus cereus. Because of its hydrophobic character, it does not induce specific antibodies. Therefore, HEp-2 cell vacuolation assay is usually used for the detection of cereulide. This method is highly sensitive as the minimum detection sensitivity of 5 ng/ml. However, there are several problems such as requirement for skill in the judgment of vacuolation and poor reproducibility in the quantification of cereulide in this method. Our laboratory has been trying to establish a new practicable method for the cereulide detection. In this study we improved the reproducibility of the standard HEp-2 cell vacuolation assay and examined the concentrations of cereulide production by B. cereus. We found that culture media, temperature and culture stages remarkably affected the cereulide production. The highest cereulide secretion was observed in mid- to late-stationary phases, when B. cereus was incubated in 50 ml of 10% skim milk medium for 30 h at 30° C with a rotary shaker at 200 rpm. When HEp-2 cell vacuolation assay was performed under this condition, the result of between-run reproducibility was improved dramatically to CV 35.3% from before modification of CV 64.0%. Since the cereulide production changed markedly depending on the culture conditions, false-negative results can be brought about by inadequate culture conditions. Therefore, the optimal culture condition will lead to improvement of detection sensitivity as well as reproducibility. |
| Keywords |
Bacillus cereus, emetic toxin cereulide, HEp-2 cell vacuolation assay, cereulide production, culture condition |
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