caa a22 4500 477069711 CHVBK 20180405111421.0 cr unu---uuuuu 170330e19960601xx s 000 0 eng 10.1007/BF01691317 doi (NATIONALLICENCE)springer-10.1007/BF01691317 Single-tube nested polymerase chain reaction assay based on flagellin gene sequences for detection of Borrelia burgdorferi sensu lato [Elektronische Daten] [M. Picken, R. Picken, D. Han, Y. Cheng, F. Strle] An inherent drawback of nested PCR systems to increase sensitivity of PCR-based assays is that tubes must be opened after the first round of amplification in order to transfer template molecules to the second amplification reaction; this procedure introduces the risk of carry-over contamination of negative specimens. To obviate this disadvantage, a nested PCR assay for detection ofBorrelia burgdorferi in which both amplifications are performed in a single tube that remains closed throughout the entire process was devised. The assay is based on flagellin gene sequences with previously determined species-wide and species-specific properties. The nested PCR system proved to be 1000 times more sensitive than the conventional assay. Using the nested PCR system, ten spirochaetes could be routinely detected by agarose gel electrophoresis alone, whereas the conventional PCR system could detect only 104 spirochaetes under these conditions. After Southern transfer of amplification products and hybridization with32P- or chemiluminescent-labeled probes, the nested PCR system could easily detect a single spirochaete by both means, whereas the sensitivity of the conventional PCR assay varied from 101 (32P) to 103 (chemiluminescence) spirochaetes. This single-tube nested PCR system should be a useful addition to the current range of diagnostic assays for Lyme borreliosis. MMV Medizin Verlag GmbH, 1996 Picken M. Department of Pathology, Room 2242 Building 110, Loyola University Medical Center, 2160 S. First Avenue, 60153, Maywood, Illinois, USA aut Picken R. Section of Infectious Disease, Rush-Presbyterian-St. Luke's Medical Center, 600 South Paulina, Chicago, Illinois, USA aut Han D. Section of Infectious Disease, Rush-Presbyterian-St. Luke's Medical Center, 600 South Paulina, Chicago, Illinois, USA aut Cheng Y. Section of Infectious Disease, Rush-Presbyterian-St. Luke's Medical Center, 600 South Paulina, Chicago, Illinois, USA aut Strle F. Department of Infectious Diseases, University Medical Centre, Ljubljana, Slovenia aut European Journal of Clinical Microbiology and Infectious Diseases Springer-Verlag 15/6(1996-06-01), 489-498 0934-9723 15:6<489 1996 15 10096 https://doi.org/10.1007/BF01691317 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/BF01691317 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Picken M. Department of Pathology, Room 2242 Building 110, Loyola University Medical Center, 2160 S. First Avenue, 60153, Maywood, Illinois, USA aut NATIONALLICENCE 700 1- Picken R. Section of Infectious Disease, Rush-Presbyterian-St. Luke's Medical Center, 600 South Paulina, Chicago, Illinois, USA aut NATIONALLICENCE 700 1- Han D. Section of Infectious Disease, Rush-Presbyterian-St. Luke's Medical Center, 600 South Paulina, Chicago, Illinois, USA aut NATIONALLICENCE 700 1- Cheng Y. Section of Infectious Disease, Rush-Presbyterian-St. Luke's Medical Center, 600 South Paulina, Chicago, Illinois, USA aut NATIONALLICENCE 700 1- Strle F. Department of Infectious Diseases, University Medical Centre, Ljubljana, Slovenia aut NATIONALLICENCE 773 0- European Journal of Clinical Microbiology and Infectious Diseases Springer-Verlag 15/6(1996-06-01), 489-498 0934-9723 15:6<489 1996 15 10096 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer