Get 20M+ Full-Text Papers For Less Than $1.50/day. Start a 14-Day Trial for You or Your Team.

Learn More →

Application of cell culture enrichment for improving the sensitivity of mycoplasma detection methods based on nucleic acid amplification technology (NAT)

Application of cell culture enrichment for improving the sensitivity of mycoplasma detection... Herein, we present data demonstrating that the application of initial cell culture enrichment could significantly improve mycoplasma testing methods based on the nucleic acid amplification technology (NAT) including a polymerase chain reaction (PCR)/microarray method. The results of the study using Vero cells demonstrated that this cell culture is able (1) to support efficient growth of mycoplasmas of primary interest, i.e., species found to be cell line contaminants, (2) to increase the sensitivity of NAT assay to the detection limits of the conventional broth/agar culture methods, and (3) to reduce the time required for mycoplasma testing fourfold in comparison with the conventional methods. Detection and identification of mycoplasmal agents were conducted using a modified PCR/microarray assay based on genetic differences among Mollicutes in the 16S-23S rRNA intergenic transcribed spacer (ITS). The application of nano-gold/silver enhancement technology instead of previously used fluorescent dyes significantly simplified the readout of microarray results and allowed us to avoid using expensive scanning equipment. This modification has the potential to expand the implementation of microarray techniques into laboratories involved in diagnostic testing of mycoplasma contamination in cell substrates and potentially in other biological and pharmaceutical products. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Applied Microbiology and Biotechnology Springer Journals

Application of cell culture enrichment for improving the sensitivity of mycoplasma detection methods based on nucleic acid amplification technology (NAT)

Loading next page...
 
/lp/springer-journals/application-of-cell-culture-enrichment-for-improving-the-sensitivity-Ib8My0FcaW

References (51)

Publisher
Springer Journals
Copyright
Copyright © 2007 by Springer-Verlag
Subject
Chemistry; Microbial Genetics and Genomics; Microbiology ; Biotechnology
ISSN
0175-7598
eISSN
1432-0614
DOI
10.1007/s00253-007-1135-1
pmid
17717660
Publisher site
See Article on Publisher Site

Abstract

Herein, we present data demonstrating that the application of initial cell culture enrichment could significantly improve mycoplasma testing methods based on the nucleic acid amplification technology (NAT) including a polymerase chain reaction (PCR)/microarray method. The results of the study using Vero cells demonstrated that this cell culture is able (1) to support efficient growth of mycoplasmas of primary interest, i.e., species found to be cell line contaminants, (2) to increase the sensitivity of NAT assay to the detection limits of the conventional broth/agar culture methods, and (3) to reduce the time required for mycoplasma testing fourfold in comparison with the conventional methods. Detection and identification of mycoplasmal agents were conducted using a modified PCR/microarray assay based on genetic differences among Mollicutes in the 16S-23S rRNA intergenic transcribed spacer (ITS). The application of nano-gold/silver enhancement technology instead of previously used fluorescent dyes significantly simplified the readout of microarray results and allowed us to avoid using expensive scanning equipment. This modification has the potential to expand the implementation of microarray techniques into laboratories involved in diagnostic testing of mycoplasma contamination in cell substrates and potentially in other biological and pharmaceutical products.

Journal

Applied Microbiology and BiotechnologySpringer Journals

Published: Aug 24, 2007

There are no references for this article.