Full Description

There has been a long-standing need to develop autonomous pathogen detection systems forwater samples that will eliminate the time, variability, and operator involvement needed to obtainaccurate results. Construction and validation of an autonomous system for pathogen detection inwater supplies is a significant challenge that must take into account: the representative volumeof water that must be sampled and examined to have statistical confidence in the results; themethods used for concentrating the sample into reasonable volumes for analyses; theprocessing and purification steps needed for detection; the detection platform chosen (e.g. cellculture, PCR, fluorescent antibody); and, the engineering required to accomplish thesetasks that will provide precise and accurate results with limited false positive and false negativeerror rates. The authors are currently testing a breadboard field device that was originallydesigned for aerosolized bioterrorism agents. However, the guiding biochemistry that drives thesystem can be readily adapted for water quality monitoring. The prototype field device iscurrently being developed for E. coli O157:H7. The device can autonomously sample up to 100mL volumes of water. Processing to remove inhibitors is accomplished by flow-throughimmunomagnetic separation. The purified cells, immobilized on the magnetic beads, are thenmoved to a flow through PCR system, and PCR is performed directly off of the beads. Theeluate is collected and the PCR sample is hybridized to an array with specific probes for thedetection of these products. Both specificity and sensitivity of the automated protocol areexcellent. Consistent recovery of 10 spiked cells into matrix water (Columbia River water) isroutinely achieved. The system architecture and PCR thermal cycling protocols virtuallyeliminate PCR carryover allowing the system to be used many times before tubing and othereasy user replaceable parts is required. The next generation system will be able to collect muchlarger volumes of water (up to 10 L), and link highly multiplexed PCR for multiple agents, witha flow through bead-array system for true autonomous detection of any waterborne pathogen. Includes 31 references, figures.

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Edition: Vol. - No. Published: 11/02/2003 Number of Pages: 10File Size: 1 file , 540 KB