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Journal of Histochemistry and Cytochemistry, Vol. 51, 19-29, January 2003, Copyright © 2003, The Histochemical Society, Inc.

ARTICLE

Simultaneous Imaging and Functional Assessment of Cytoskeletal Protein Connections in Passively Loaded Single Muscle Cells

Sameer B. Shaha and Richard L. Liebera
a Departments of Bioengineering and Orthopaedics, Biomedical Sciences Graduate Group, University of California and Veterans Administration Medical Centers, San Diego, California

Correspondence to: Richard L. Lieber, Dept. of Orthopaedics (9151), VA Medical Center and UC San Diego, 3350 La Jolla Village Drive, San Diego, CA 92161. E-mail: rlieber@ucsd.edu

We describe a novel system that permits simultaneous confocal imaging of protein interactions and measurement of cell mechanical properties during passive loading. A mechanical apparatus was designed to replace the stage of a confocal microscope, enabling cell manipulation, force transduction, and imaging. In addition, image processing algorithms were developed to quantify the degree of connectivity between subcellular structures. Using this system, we examined the interactions among three cellular structures thought to be linked by the muscle's intermediate filament system: Z-disks, nuclei, and the costamere protein complexes located at the muscle cell surface. Fast Fourier transforms (FFTs) and autocorrelations (ACs) were implemented to quantify image periodicity and relative phase shifts among structures. We demonstrated in sample wild-type muscle cells that there was significant connectivity among Z-disks in the same fiber at various sarcomere lengths, as well as between Z-disks and the costamere complexes. This approach can be applied to any cell system in which structural periodicity and mechanical connectivity are of interest. (J Histochem Cytochem 51:19–29, 2003)

Key Words: costamere, talin, {alpha}-actinin, sarcomere length, image analysis


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S. B. Shah, J. Davis, N. Weisleder, I. Kostavassili, A. D. McCulloch, E. Ralston, Y. Capetanaki, and R. L. Lieber
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