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| High Throughput Characterization of Amplified Nucleic Acids By ESI-TOF Mass Spectrometry: Applications of the IBIS T5000 to Pathogen Detection and Characterization |
| Wed 11:00 AM - Session: Molecular Diagnostics |
| Steve Hofstadler |
| High throughput electrospray ionization time-of-flight (ESI-TOF) mass spectrometric analysis of polymerase chain reaction (PCR) amplicons represents a novel and universal strategy for the detection and characterization of microorganisms associated with emerging infectious diseases. The process uses mass spectrometry, signal processing, and base composition analysis of PCR amplification products from biologically conserved regions of microbial genomes to simultaneously identify the organisms present in a sample without the need for culture. The sample can be derived from environmental samples, clinical specimens, or other sources. Core to this approach are broad range PCR primers that target broadly conserved regions of microbial genomes that flank variable regions. Using high-performance electrospray ionization mass spectrometry (ESI-MS), the base composition (i.e., the number of As, Gs, Cs, and Ts) of each amplicon is unambiguously determined.
This strategy distinguishes this approach from other detection/identification strategies in that it requires little or no prior knowledge about an organism in order to identify it in a sample. The approach requires that high-performance mass measurements be made on PCR products in the 80 - 140 bp size range in a high-throughput, robust modality. The base compositions from multiple primer pairs are used to 'triangulate' the identity of the organisms present in the sample. Use of species-specific primers allows rapid strain-typing of the organism. This presentation is intended to serve as a technology overview and will include examples of the technology for detection and/or strain typing of bacteria, viruses (both DNA and RNA-based), and fungi. |
| Email: shofstad@ibisbio.com |