title: Rapid and effective method for preparation of fecal specimens for PCR assays.
abstract: We have developed a novel method for the preparation of fecal specimens for PCR assays. Approximately 100 mg of solid stool or 200 microliters of liquid fecal sample was thoroughly suspended in 1 ml of water. Fecal debris was removed by low-speed centrifugation (2,800 x g for 2 min). The supernatant was then boiled for 10 min in a water bath and further clarified by high-speed centrifugation (12,000 x g for 5 min). Fifty microliters of the clarified supernatant was then purified by Sepharose CL-6B spin column chromatography, and a portion of the purified supernatant was used for PCR. By this method, stools containing enterotoxigenic Escherichia coli H10407 were amplified by colonization factor antigen I fimbrial gene PCR, with a sensitivity of 100 organisms per reaction. The method was also effective for processing stool specimens for Clostridium difficile toxin A and B gene PCRs. This method is rapid, effective, and simple to perform and will improve the applications of PCR to stool specimens for diagnostic purposes.
authors.affiliation: Department of Pediatrics, Indiana University School of Medicine, Indianapolis 46202, USA.
authors.forename: Q
authors.initials: Q
authors.lastname: Lou
authors.forename: S K
authors.initials: SK
authors.lastname: Chong
authors.forename: J F
authors.initials: JF
authors.lastname: Fitzgerald
authors.forename: J A
authors.initials: JA
authors.lastname: Siders
authors.forename: S D
authors.initials: SD
authors.lastname: Allen
authors.forename: C H
authors.initials: CH
authors.lastname: Lee
mesh_terms: D015373:Bacterial Typing Techniques; D005243:Feces / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods*