Confocal Unit

Keller, Raymond

Abstract

The objective of this proposal is to provide state of art confocal laser scanning microscopy capabilities for key, ongoing NIH-supported research programs at the University of Virginia. To accomplish this goal, purchase of a confocal laser scanning microscope (CLSM), specifically a Biorad MRC 1024 scanning unit with an Olympus IX70 inverted microscope, is proposed. This instrument will be integrated into a central core facility, the Advanced Cellular Imaging Facility (ACIF), as a multi-user system, serving 6 investigators, all of whom have pressing, unmet needs for CLSM in their research. Use of green fluorescent protein (GFP) and its color variants as fusion proteins, as well as other fluorescent markers, to study the localization, function, and interaction of gene products, has resulted in many ground-breaking applications in cell and developmental biology and genetics. In this context, the participating faculty need a CLSM of high sensitivity, capable of doing time- lapse, repeated imaging of living material with minimal photoxicity, and capable of imaging several fluorescent markers of different colors. The 6 major users (Adler, Barroso, Corwin, Green, Hays, and Keller) all have applications requiring imaging multiple fluorescent labels in 3-D, both static imaging of fixed preparations and dynamic time-series imaging of living cells. These NIH-supported research projects involve imaging the locations, movements, and interactions of molecules critical in several aspects of cell structure and function. Other applications involve imaging important aspects of morphogenesis and cell movement in developmental biology. Currently, the available instruments suffer from out of date software, inadequate sensitivity, no time-lapse capability, or are limited to single color imaging and thus are inadequate to meet the users' needs. The MRC 1024 will fill the need for an instrument with high sensitivity and fast image acquisition times for near real time and time-lapse imaging of living cells with minimal phototoxic effects. These capabilities are essential for our faculty to remain competitive in research and in research training.