Evaluation of the Magnetic Immuno PCR assay for rapid detection ofSalmonella
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A new technique, the Magnetic Immuno PCR Assay (MIPA), has been developed for the detection ofSalmonella. The assay utilizes magnetic particles coated with monoclonal antibodies againstSalmonella to extract these bacteria from the sample. Trapped bacteria are lysed, and the supernatant, which contains bacterial DNA, is then subjected to the polymerase chain reaction (PCR) using primers from theSalmonella typhimurium origin of DNA replication to amplify a 163 bp region. The specificity of the primer set was tested in the PCR; amplification occurred with all 25Salmonella strains tested but not with 19 other species ofEnterobacteriaceae tested. A sensitivity of 100 cfuSalmonella typhimurium was achieved for the MIPA by visualization of the amplified products by ethidiumbromide stained agarose gel electrophoresis. A tenfold higher sensitivity was obtained by Southern blotting of the amplified products. The presence of 107 cfuEscherichia coli did not interfere with these detection levels. The MIPA thus specifically detected 100 cfu ofSalmonella within 5 h and may be potentially useful for rapid detection ofSalmonella in clinical specimens and food.
KeywordsPolymerase Chain Reaction Monoclonal Antibody Agarose Ethidiumbromide Southern Blotting
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- 1.Helrich K (ed Official methods of analysis. Association of Official Analytical Chemists, Arlington, VA, 1990, p. 467–492.Google Scholar
- 2.Van Leusden FM, van Schothorst M, Beckers HJ The standardSalmonella isolation method. In: Corry JEL, Roberts D, Skinner FA (ed): Isolation and identification methods for food poisoning organisms. Academic Press, London, 1982, p. 35–49.Google Scholar
- 4.Eckner KF, Flowers RS, Robison BJ, Mattingly JA, Gabis DA, Silliker JH Comparison ofSalmonella Bio-Enza Bead immunoassay method and conventional culture procedure for detection ofSalmonella in foods. Journal of Food Protection 1987, 50: 379–385.Google Scholar
- 13.Kornberg A Supplement to DNA replication. W.H. Freeman, San Fransisco, 1982, p. 145–152.Google Scholar
- 15.Maniatis T, Fritch EF, Sambrook J Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, 1982.Google Scholar