The goal of the proposed research is to develop novel methods for the detection and measurement of hormone secretion from a single cell. Such methods require the both ability to recover secreted materials at the single cell level and the capability to measure the target analytes at extremely low levels. BioTraces' proprietary Multi Photon Detection (MPD) technique permits detection of minute (less than 10-18 mole) quantities of biological material. This unitque capability will be combined with micro-dialysis sampling apparatus at George Washington Unviersity Medical Center (GWUMC) to quantify hormone secretion at the single cell level. The GWUMC micro-dialysis apparatus will be modified to accommodate dialysis probes capable of work at the single cell level. Samples obtained from this single apparatus will be anlyzed by MPD enhanced high performance liquid chromatography (HPLC) and by BioTraces' proprietary SuperIRMA immunoassay protocol. In Phase I this system will be used to quantify the synthesis of insulin from rat pancreatic cells and insulin-like growth factor-I from human fibroblasts. MPD-enhanced HPLC and SuperIRMA assays for these targets will be developed and their accuracy and precision will be validated in Phase I. POTENTIAL COMMERCIAL APPLICATIONS The ability to obtain and analyze samples from single living cells is of immense importance in the study of the pathogenesis and treatment of many diseases, including diabetes. The system porposed herein will provide a generic tool for studying peptide production at the cellular level, and thus will be an indispensable tool for understanding physiological function and disease response at the cellular level.
