Validation of a cell-free translation assay for detecting shiga toxin 2 in bacterial culture
Submitted by Beatriz Quiñones on
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Title | Validation of a cell-free translation assay for detecting shiga toxin 2 in bacterial culture |
Publication Type | Journal Article |
Year of Publication | 2009 |
Authors | He, X, Quiñones, B, Carter, JM, Mandrell, RE |
Journal | J Agric Food ChemJ Agric Food ChemJ Agric Food Chem |
Volume | 57 |
Pagination | 5084-8 |
Date Published | Jun 10 |
ISBN Number | 1520-5118 (Electronic)0021-8561 (Linking) |
Accession Number | 19489630 |
Abstract | A cell-free translation (CFT) assay for detecting Shiga toxin (Stx) at different levels of purity has been validated. The limits of detection for pure Stx2 (PStx2) and partially pure Stx2 (PPStx2) in water reached 20 and 3.5 pg/microL, respectively, without the artificial process of proteolytic activation and reduction of the pro-toxin. The specific detection of Stx2 was confirmed by a neutralization test using Stx2-specific mouse monoclonal antibody. This assay can be used for differentiation of Stx-producing Escherichia coli from non-Stx-producing E. coli. Four E. coli O157:H7 strains genotypically different for Stx were tested. The translational inhibition of Stx-producing E. coli was significantly higher than that of non-Stx-producing E. coli when bacterial culture supernatants were used for the analysis. Inhibition occurred even with supernatants diluted 1000-fold. The thermal stability of Stx2 was studied using the CFT assay, and significant differences were observed among three Stx2 preparations heated at 70 degrees C for 60 min. It was concluded that the CFT assay is a rapid, specific, and sensitive method for detecting Stx2 activity. |
Short Title | Journal of agricultural and food chemistryJournal of agricultural and food chemistry |
Alternate Journal | Journal of agricultural and food chemistry |