A Novel Semiquantitative Fluorescence-Based Multiplex Polymerase Chain Reaction Assay for Rapid Simultaneous Detection of Bacterial and Parasitic Pathogens.

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A Novel Semiquantitative Fluorescence-Based Multiplex Polymerase Chain Reaction Assay for Rapid Simultaneous Detection of Bacterial and Parasitic Pathogens from Blood Angamuthu Selvapandiyan, Katie Stabler, Nasim A. Ansari, Stephen Kerby, Jenny Riemenschneider, Poonam Salotra, Robert Duncan, Hira L. Nakhasi The Journal of Molecular Diagnostics Volume 7, Issue 2, Pages 268-275 (May 2005) DOI: 10.1016/S1525-1578(10)60554-5 Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 1 A: Melt derivative analysis of the multiplex fluorescence PCR showing the amplification of specific products of B. anthracis (Sterne) (BA) with Tm 82.2°C, L. donovani (LD) with Tm 86.1°C, Y. pseudotuberculosis (YP) with TM 87.3°C, and the internal human control (HS) with Tm 88.8°C. B: Agarose gel confirmation of the products of the multiplex PCR stained with ethidium bromide from A showing the amplified products of: LD, L. donovani; HS, human; YP, Y. pseudotuberculosis; and BA, B. anthracis (Sterne). The Journal of Molecular Diagnostics 2005 7, 268-275DOI: (10.1016/S1525-1578(10)60554-5) Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 2 A: Melt derivative analysis of the multiplex PCR assay with human blood carrying a known number of spiked B. anthracis (Sterne) spores. B: Agarose gel confirmation of the products of the multiplex PCR showing the linear relationship between the logarithm of the number of spores spiked and the amount of amplified band intensity of B. anthracis (Sterne). C: Quantitation graph showing the linear relationship between the height of the fluorescence peak above baseline of the specific amplicons (y axis) and the logarithm of the number of bacterial or parasitic pathogen cells individually spiked in 200 μl of blood (x axis). BA, B. anthracis (Sterne); YP, Y. pseudotuberculosis; LD, L. donovani; a.v., arbitrary value. The data represent the means ± SD of three independent experiments. The Journal of Molecular Diagnostics 2005 7, 268-275DOI: (10.1016/S1525-1578(10)60554-5) Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions