Development and Applications of an LCOS-Based Programmable Array Microscope

Abstract:
The programmable array microscope (PAM) is a unique confocal microscope which utilizes a spatial light modulator in an image plane to generate patterns of illumination and detection, offering a combination of high speed confocal imaging with high light throughput. The PAM is useful for rapid, 3D optical sectioning of live cells. The major advantages of the PAM are: (a) speedup in optical sectioning (b) arbitrary and potentially adaptive patterns for imaging and, (c) minimal photobleaching. These characteristics enable experiments not possible with conventional systems. The PAM will be used for live cell imaging, including studies of cell membrane protein organization and signaling. The studies will also be aimed at nuclear events, especially studies of large scale chromatin organization using fluorescent protein-tagged histones. Studies will include evaluation via FRET of chromatin compaction and application of FRAP and photoactivation.

The specific aims of the current project are: 1) construction of a PAM and polarization-based live cell measurements of FRET in the cell nucleus 2) development of adaptive optical and control systems 3) fluorescence photobleaching and photoactivation experiments on proteins of the cell membrane and nucleus and 4) implementation of structured illumination microscopy for imaging beyond the Abbe limit.