Operational Technologies Corporation (OpTech) proposes to expand its current NASA Phase 2 SBIR handheld fluorometer and bone marker fluorescence resonance energy transfer (FRET)-DNA aptamer assay system to include detection of bacteria, fungi, and parasites that may contaminate astronauts' water supplies. For Gram positive bacteria, teichoic acids and peptidoglycan will serve as targets. For Gram negative bacteria, common lipopolysaccharide moieties such as 2-keto-3-deoxyoctanate (KDO antigen) will be targeted for aptamer development. Similarly, for fungi, cell wall chitin will be used to select highly specific FRET-aptamers from a randomized DNA library. Parasites such as Cryptosporidium and Giardia will require more specific whole cell or surface protein aptamer selection approaches, but OpTech has recently demonstrated detection of 30 E. coli bacteria per ml using such an approach under NSF Phase 1 SBIR funding. Prototype assays will be lyophilized in plastic cuvettes and capped under vacuum or otherwise sealed to prevent opening in negative pressure environments. Lyophilization with trehalose or other excipients will extend shelf-life to greater than 2 years for these rapid (15 minute) one step (homogeneous) FRET assays that will be quantified with an ultrasensitive commercial handheld fluorometer. Data can be displayed on the handheld reader and downloaded to a laptop computer.
