9] Cytomechanics applications of optical sectioning microscopy

Abstract

This chapter describes the cytomechanics applications of optical sectioning microscopy. In the past half-century since the purification of actin, the fields of biochemistry and biophysics have produced a vast trove of information on the protein components and regulatory mechanisms of the cytoskeleton. The action or mechanics of the cytoskeleton must be measured and matched to identifiable assemblies. To analyze this cellular machine, an elastic collagen-based model extracellular matrix (ECM) can be utilized as a strain gauge. The light microscope, when suitably automated and interfaced to a digital imaging system, then serves a dual purpose: (1) to image the configuration of the cytoskeleton with molecular specificity through fluorescence labeling and (2) to image the deformations produced by the cell in the surrounding matrix. The basic instrument consists of a fluorescence microscope equipped with a cooled charge-coupled device (CCD) camera system, programmable shutter, grating slider with drive mechanism, and desktop computer for control, image acquisition, and processing. The cooled CCD camera provides the high linearity, dynamic range, geometric precision, and low noise needed for digital processing.