Our objective is to develop the scientific and engineering basis for a new reaction chamber technology that will be incorporated into a workstation for analytical cytochemistry. The technology is based on a """"""""Probe-Clip"""""""" device - a disposable solid-phase reactant plate which seals reversibly to a tissue-bearing microscope slide to form a closed high-aspect (500um in height) reaction unit in which reactants flow by gravity. Preformatted lyophilized probe is sequestered in the unit during the blocking phase of assay and is activated on-command by rotating the closed unit in an Assembler to bring blocking solution into contact. The new technology addresses the three aspects of commercially available cytochemical assay technology which mitigate their use for quantitative work: 1) an open reaction chamber; 2) a low-aspect (ca. 150um) reaction chamber characterized by capillarity flow and large static boundary layers of reagents; and 3) ad hoc manipulation and delivery of probe. Phase 1 feasibility experiments showed that the process control afforded by Probe- Clip technology is better suited to the demands of analytical cytochemistry for low-abundance regulatory targets, using non-isotopic end-points. In Phase 2, we propose to develop the scientific basis for reproducible fabrication of Probe-Clip and Assembler hardware, and to develop standard procedures for use of the technology. Automation of the technology in Phase 3 will build on the culmination of Phase 2 research, a prototype robotic cassette mechanism.
