부산대학교 도서관

To purchase or authenticate to the full-text of this article, please visit this link: http://dx.doi.org/10.1111/j.1574-695X.2006.00125.x Byline: Younes Maaroufi (1), Jean-Marc de Bruyne (1), Valerie Duchateau (1), Robert Scheen (2), Francoise Crokaert (1,2) Keywords: multiple internal control; real-time PCR; false-negative PCR results Abstract: Abstract A major pitfall in most published genomic amplification methods for the detection and identification of human pathogens is that they do not include an internal amplification control in order to achieve an acceptable level of confidence for the absence of false-negative results. By applying composite primer technology, a single multiple internal amplification control DNA molecule was constructed to detect and quantify the hepatitis B virus, human polyomavirus, Epstein-Barr virus, Toxoplasma gondii and cytomegalovirus using real-time PCR. The multiple internal amplification control contains all forward and reverse primer binding regions targeted in the five distinct duplex PCRs, but with a unique probe hybridization site. Multiple internal amplification control detection sensitivity, assessed by Probit analysis, was 58 copies per PCR, associated with an extremely wide dynamic range (8 log.sub.10 units). Moreover, in testing 614 patient samples, PCR inhibition occurred at a frequency of 0-8.8%. Similar multiple internal amplification controls for quantitative PCR-based assays could be designed to accommodate any infectious profiles in a particular institution as they are easy to make and inexpensive. Author Affiliation: (1)Department of Microbiology and Infectious Diseases, Institut Jules Bordet, Brussels, Belgium (2)Department of Microbiology, C.H.U. Saint-Pierre, Brussels, Belgium Article History: Received 22 March 2006; revised 3 May 2006; accepted 29 May 2006.First published online 12 September 2006. Article note: Correspondence: Francoise Crokaert, Department of Microbiology and Infectious Diseases, Institut Jules Bordet, Rue Heger-Bordet 1, 1000 Brussels, Belgium. Tel.: +322 541 3700/3706; fax: +322 541 3295; e-mail: fcrokaer@ulb.ac.be