The purpose of this proposal is to acquire NIH funding for a Carl Zeiss Elyra S1 SIM system for the CNR Biological Imaging Facility (BIF). This super-resolution microscope system will augment the research and training capabilities of the BIF, and specifically, will permit the research goals of the six (6) Major Users listed in this proposal to proceed forward with their active NIH-funded research. The BIF is the main focus for research involving optical microscopy for a great many scientists at UC Berkeley (379 PIs/2431 users). A major limitation, however, is the inability to resolve fluorescent targets at a """"""""super-resolution"""""""" level. At the present time the highest resolution optical imaging technique available to these users is deconvolution (API DeltaVision Elite). There is only one super-resolution system (PALM) available on campus. Our researchers require 2-3 channel 3D super-resolution imaging, and thus only a Structured Illumination Microscope (SIM) is appropriate for our needs. STED does not work with long wavelength probes and is therefore disqualified for all of our users. PALM/STORM yields little or no 3D information. In this proposal we have shown that the research of our Major Users is crippled by the lack of the ability to resolve structures smaller than diffraction-limited resolution of 200nm. The requested instrument will fill a much-needed gap in optical imaging technology by providing reproducible 100nm resolution using any color probe. This critical capability will facilitate the research of scientists in the College of Natura Resources, the College of Letters and Sciences, and the general scientific community at the University of California, Berkeley. Over the last year we performed a preliminary SIM experiments on biological samples using a Carl Zeiss Elyra S1, Nikon N-SIM, and the API OMX, and we compared the results to that obtained using the BIF's deconvolution and LSM 710 confocal microscopes. For each Major User's research project we present unequivocal evidence showing that SIM imaging far surpasses the capabilities of our existing microscopes in resolution, sensitivity, and the ability to image in 3D.
