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Journal of Nucleic Acids
Volume 2015, Article ID 808495, 12 pages
http://dx.doi.org/10.1155/2015/808495
Research Article

In Vitro Selection of a Single-Stranded DNA Molecular Recognition Element against Clostridium difficile Toxin B and Sensitive Detection in Human Fecal Matter

1Department of Basic Pharmaceutical Sciences, West Virginia University, 1 Medical Center Drive, P.O. Box 9530, Morgantown, WV 20506, USA
2Molecular Pharmacology & Chemistry Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA

Received 18 November 2014; Revised 14 January 2015; Accepted 19 January 2015

Academic Editor: Ben Berkhout

Copyright © 2015 Ka Lok Hong et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Toxin B is one of the major virulence factors of Clostridium difficile, a bacterium that is responsible for a significant number of diarrhea cases in acute care settings. Due to the prevalence of C. difficile induced diarrhea, rapid and correct diagnosis is crucial in the disease management. In this study, we have employed a stringent in vitro selection method to identify single-stranded DNA molecular recognition elements (MRE) specific for toxin B. At the end of the 12-round selection, one MRE with high affinity  nM) for toxin B was identified. The selected MRE demonstrated low cross binding activities on negative targets: bovine serum albumin, Staphylococcus aureus alpha toxin, Pseudomonas aeruginosa exotoxin A, and cholera toxin of Vibrio cholera. A modified sandwich ELISA assay was developed utilizing the selected ssDNA MRE as the antigen capturing element and achieved a sensitive detection of 50 nM of toxin B in human fecal preparations.