The goal of this project is to develop a portable system for the real-time detection and quantification of adenovirus in wastewater. The system is based on a small, portable flow cytometer that was developed and is being marketed by Becton Dickinson Biosciences (BDB) under the trade name FACSCount for use in selected medical and research applications. Unlike previous efforts to apply flow cytometry to the testing of wastewater, the system that will be used for this work is based on the measurement of antibody-coated fluorescent microspheres with a low-cost, user friendly, commercially tested flow cytometer, the BDB FACSCount. BDB has granted to GEOMET Technologies, Inc., exclusive license to develop and market the system for environmental applications. BDB is supporting this development program by providing the instrument, assisting in reagent formulations, and developing the specialized software needed for this application. Flow cytometry is a form of spectroscopy that interrogates individual members of a sample population for size, shape, biological, and chemical properties, thus making it a potential tool for determining the concentration of specific particles in a heterogeneous sample. Cells or fragments and a reagent blend specific to the target biologic are carried by a fluid stream through a focus of exciting light. The distribution and intensity of the scattered light and fluorescence imparted by fluorescent probes generates both a scatter and fluorescence fingerprint that is interpreted by an internal computer that determines the presence and concentration of the target biologic. The key to using flow cytometry for this purpose is development of reagents and software that are unique to adenovirus in wastewater. This project involves the development of the specific reagents and software to quantify adenovirus in wastewater. The overall system will be usable by a minimally trained technician under field conditions with a 15-minute turnaround. The Phase I research effort will develop the reagent blend, modify the software, and confirm that the system's sensitivity is adequate for wastewater analysis; preliminary indications are encouraging. The Phase II effort will simplify the programming and the reagent delivery system so that the analyses can be performed by a technician in the field, and it will validate the procedure against the standard U.S. Environmental Protection Agency method for adenovirus. In Phase III, the system will be applied to multiple microbiologics in a single sample. Supplemental
Keywords: small business, SBIR, wastewater, cytometer, adenovirus, antibody-coated fluorescent microspheres, real-time detection
