title: Detection of biological threat agents by real-time PCR: comparison of assay performance on the R.A.P.I.D., the LightCycler, and the Smart Cycler platforms.
abstract: BACKGROUND
  Rapid detection of biological threat agents is critical for timely therapeutic administration. Fluorogenic PCR provides a rapid, sensitive, and specific tool for molecular identification of these agents. We compared the performance of assays for 7 biological threat agents on the Idaho Technology, Inc. R.A.P.I.D., the Roche LightCycler, and the Cepheid Smart Cycler.
  
  
  METHODS
  Real-time PCR primers and dual-labeled fluorogenic probes were designed to detect Bacillus anthracis, Brucella species, Clostridium botulinum, Coxiella burnetii, Francisella tularensis, Staphylococcus aureus, and Yersinia pestis. DNA amplification assays were optimized by use of Idaho Technology buffers and deoxynucleotide triphosphates supplemented with Invitrogen Platinum Taq DNA polymerase, and were subsequently tested for sensitivity and specificity on the R.A.P.I.D., the LightCycler, and the Smart Cycler.
  
  
  RESULTS
  Limit of detection experiments indicated that assay performance was comparable among the platforms tested. Exclusivity and inclusivity testing with a general bacterial nucleic acid cross-reactivity panel containing 60 DNAs and agent-specific panels containing nearest neighbors for the organisms of interest indicated that all assays were specific for their intended targets.
  
  
  CONCLUSIONS
  With minor supplementation, such as the addition of Smart Cycler Additive Reagent to the Idaho Technology buffers, assays for DNA templates from biological threat agents demonstrated similar performance, sensitivity, and specificity on all 3 platforms.
authors.affiliation: United States Army Medical Research Institute of Infectious Diseases, Fort Detrick, MD 21702, USA.
authors.forename: Deanna R
authors.initials: DR
authors.lastname: Christensen
authors.forename: Laurie J
authors.initials: LJ
authors.lastname: Hartman
authors.forename: Bonnie M
authors.initials: BM
authors.lastname: Loveless
authors.forename: Melissa S
authors.initials: MS
authors.lastname: Frye
authors.forename: Michelle A
authors.initials: MA
authors.lastname: Shipley
authors.forename: Deanna L
authors.initials: DL
authors.lastname: Bridge
authors.forename: Michelle J
authors.initials: MJ
authors.lastname: Richards
authors.forename: Rebecca S
authors.initials: RS
authors.lastname: Kaplan
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Garrison
authors.forename: Carson D
authors.initials: CD
authors.lastname: Baldwin
authors.forename: David A
authors.initials: DA
authors.lastname: Kulesh
authors.forename: David A
authors.initials: DA
authors.lastname: Norwood
mesh_terms: D001419:Bacteria / Q000145:classification* / Q000235:genetics; D001431:Bacteriological Techniques; D001694:Biological Warfare; D004269:DNA, Bacterial / Q000235:genetics; D005470:Fluorometry; D016133:Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity
chemical_list: D004269:DNA, Bacterial