CONFOCAL MICROSCOPE RESOURCE FACILITY

The projects described by the users in this application share the common
approach of using a combination of cell biology and quantitative optical
techniques to explore a variety of processes. These investigators are most
frequently dependent on the resolution of fluorescent indicators in living
cells. The instrument chosen is a Leica CLSM confocal laser scan with a
Fluorvert inverted microscope that will be 50% cost shared with the
university. The design includes a complete two-channel detector system
that allows simultaneous measurements of two different fluorescent labels
in precise register for all image points. The high focusing accuracy
provided will enable more precise measurements of cell volume than that
afforded by other instruments and the Leica images are the best that we
have observed. All of the users in this application have been funded by
NIH for their work and for the experiments described in this proposal. The
confocal microscope will bring a new dimension to their research by
providing better resolution, removing uncertainty about co-localization,
making measurements easier and in some cases, possible, for the first time.
Several different types of measurements are described. In some cases the
investigators will be exploring new territory in trying to make sense of
a complicated pathogenesis pathway. In Forte's case, the ability to
optically section entire Drosophila embryo is the best way to follow the
developmental regulation of specific neural components. The Westbrook
experiments will test a specific model for activation of a neural receptor
by making simultaneous measurements of intracellular calcium levels and the
activation state of the receptor -- difficult measurements to make without
the resolving power of the confocal. Low's group currently makes many
measurements using traditional fluorescence to compare co-localization of
specific stains. The same measurements that take all day can be made more
accurately using optical sectioning and the data can be stored on disk in
just minutes. Thornberg's group need to make cell volume measurements that
cannot be achieved in any other way than with a confocal.