PCR–Electrospray Ionization Mass Spectrometry

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PCR–Electrospray Ionization Mass Spectrometry Donna M. Wolk, Erin J. Kaleta, Vicki H. Wysocki The Journal of Molecular Diagnostics Volume 14, Issue 4, Pages 295-304 (July 2012) DOI: 10.1016/j.jmoldx.2012.02.005 Copyright © 2012 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 1 A: The Ibis T5000 Biosensor System is the PCR-ESI/MS prototype. B: The Abbott PLEX-ID system is the current instrument. Reprinted with permission from Ibis Biosciences. The Journal of Molecular Diagnostics 2012 14, 295-304DOI: (10.1016/j.jmoldx.2012.02.005) Copyright © 2012 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 2 Conserved sequences serve as priming sites on which broad-range primers amplify sequences of various sizes and compositions, because of combinations of conserved regions that flank intercalated regions of sequence diversity, found in the space between common regions. Reprinted with permission from Ibis Biosciences. The Journal of Molecular Diagnostics 2012 14, 295-304DOI: (10.1016/j.jmoldx.2012.02.005) Copyright © 2012 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 3 Representative mass spectra from PCR-ESI/MS. Data resulting from ESI TOF MS of each microtiter well containing one primer pair and the target DNA. A: The 96-well view of spectra collected. B: Two charge-state distributions are seen in products produced with primer 346 for each forward and reverse primer, indicating the presence of two microorganisms in this sample. Blue, Bacteroides thetaiotaomicron; green, S. aureus. C: Deconvolution of collected mass spectra, resulting in a mol. wt. for the forward and reverse strand. The resulting base compositions are as follows: [30A-27G-23C-19T] (top panel) and [27A-30G-21C-21T] (bottom panel). Reprinted with permission from Kaleta et al,10 copyright American Society for Microbiology. AI, average intensity. The Journal of Molecular Diagnostics 2012 14, 295-304DOI: (10.1016/j.jmoldx.2012.02.005) Copyright © 2012 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 4 The ESI/MS generates amplicon base composition values from the intact mol. wt. measured by MS for several genetic regions and compare results to the Ibis microbial sequence database. Composition values will vary by microorganism. Reprinted with permission from Ibis Biosciences. The Journal of Molecular Diagnostics 2012 14, 295-304DOI: (10.1016/j.jmoldx.2012.02.005) Copyright © 2012 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions