The Potential of TaqMan Array Cards for Detection of Multiple Biological Agents by Real-Time PCR

The TaqMan Array Card architecture, normally used for gene expression studies, was evaluated for its potential to detect multiple bacterial agents by real-time PCR. Ten PCR assays targeting five biological agents (Bacillus anthracis, Burkholderia mallei, Burkholderia pseudomallei, Francisella tulare...

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Main Authors:	Rachwal, Phillip A., Rose, Helen L., Cox, Victoria, Lukaszewski, Roman A., Murch, Amber L., Weller, Simon A.
Format:	Online
Language:	English
Published:	Public Library of Science 2012
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3334355/

id	pubmed-3334355
recordtype	oai_dc
spelling	pubmed-33343552012-04-26 The Potential of TaqMan Array Cards for Detection of Multiple Biological Agents by Real-Time PCR Rachwal, Phillip A. Rose, Helen L. Cox, Victoria Lukaszewski, Roman A. Murch, Amber L. Weller, Simon A. Research Article The TaqMan Array Card architecture, normally used for gene expression studies, was evaluated for its potential to detect multiple bacterial agents by real-time PCR. Ten PCR assays targeting five biological agents (Bacillus anthracis, Burkholderia mallei, Burkholderia pseudomallei, Francisella tularensis, and Yersinia pestis) were incorporated onto Array Cards. A comparison of PCR performance of each PCR in Array Card and singleplex format was conducted using DNA extracted from pure bacterial cultures. When 100 fg of agent DNA was added to Array Card channels the following levels of agent detection (where at least one agent PCR replicate returned a positive result) were observed: Y. pestis 100%, B. mallei & F. tularensis 93%; B. anthracis 71%; B. pseudomallei 43%. For B. mallei & pseudomallei detection the BPM2 PCR, which detects both species, outperformed PCR assays specific to each organism indicating identification of the respective species would not be reproducible at the 100 fg level. Near 100% levels of detection were observed when 100 fg of DNA was added to each PCR in singleplex format with singleplex PCRs also returning sporadic positives at the 10 fg per PCR level. Before evaluating the use of Array Cards for the testing of environmental and clinical sample types, with potential levels of background DNA and PCR inhibitors, users would therefore have to accept a 10-fold reduction in sensitivity of PCR assays on the Array Card format, in order to benefit for the capacity to test multiple samples for multiple agents. A two PCR per agent strategy would allow the testing of 7 samples for the presence of 11 biological agents or 3 samples for 23 biological agents per card (with negative control channels). Public Library of Science 2012-04-23 /pmc/articles/PMC3334355/ /pubmed/22540014 http://dx.doi.org/10.1371/journal.pone.0035971 Text en Rachwal et al. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Rachwal, Phillip A. Rose, Helen L. Cox, Victoria Lukaszewski, Roman A. Murch, Amber L. Weller, Simon A.
spellingShingle	Rachwal, Phillip A. Rose, Helen L. Cox, Victoria Lukaszewski, Roman A. Murch, Amber L. Weller, Simon A. The Potential of TaqMan Array Cards for Detection of Multiple Biological Agents by Real-Time PCR
author_facet	Rachwal, Phillip A. Rose, Helen L. Cox, Victoria Lukaszewski, Roman A. Murch, Amber L. Weller, Simon A.
author_sort	Rachwal, Phillip A.
title	The Potential of TaqMan Array Cards for Detection of Multiple Biological Agents by Real-Time PCR
title_short	The Potential of TaqMan Array Cards for Detection of Multiple Biological Agents by Real-Time PCR
title_full	The Potential of TaqMan Array Cards for Detection of Multiple Biological Agents by Real-Time PCR
title_fullStr	The Potential of TaqMan Array Cards for Detection of Multiple Biological Agents by Real-Time PCR
title_full_unstemmed	The Potential of TaqMan Array Cards for Detection of Multiple Biological Agents by Real-Time PCR
title_sort	potential of taqman array cards for detection of multiple biological agents by real-time pcr
description	The TaqMan Array Card architecture, normally used for gene expression studies, was evaluated for its potential to detect multiple bacterial agents by real-time PCR. Ten PCR assays targeting five biological agents (Bacillus anthracis, Burkholderia mallei, Burkholderia pseudomallei, Francisella tularensis, and Yersinia pestis) were incorporated onto Array Cards. A comparison of PCR performance of each PCR in Array Card and singleplex format was conducted using DNA extracted from pure bacterial cultures. When 100 fg of agent DNA was added to Array Card channels the following levels of agent detection (where at least one agent PCR replicate returned a positive result) were observed: Y. pestis 100%, B. mallei & F. tularensis 93%; B. anthracis 71%; B. pseudomallei 43%. For B. mallei & pseudomallei detection the BPM2 PCR, which detects both species, outperformed PCR assays specific to each organism indicating identification of the respective species would not be reproducible at the 100 fg level. Near 100% levels of detection were observed when 100 fg of DNA was added to each PCR in singleplex format with singleplex PCRs also returning sporadic positives at the 10 fg per PCR level. Before evaluating the use of Array Cards for the testing of environmental and clinical sample types, with potential levels of background DNA and PCR inhibitors, users would therefore have to accept a 10-fold reduction in sensitivity of PCR assays on the Array Card format, in order to benefit for the capacity to test multiple samples for multiple agents. A two PCR per agent strategy would allow the testing of 7 samples for the presence of 11 biological agents or 3 samples for 23 biological agents per card (with negative control channels).
publisher	Public Library of Science
publishDate	2012
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3334355/
_version_	1611523873240514560

The Potential of TaqMan Array Cards for Detection of Multiple Biological Agents by Real-Time PCR

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