FLOW CYTOMETRIC DETECTION OF THE ASSOCIATION BETWEEN CELL-SURFACE RECEPTORS AND THE CYTOSKELETON

The cytoskeleton can serve as a structure at which receptors and signa ling molecules can be immobilized to react with each other and induce signal transduction, which, consequently, leads to functional response s of the cell. Furthermore, transduction of mechanical forces into the cell can be realized by a physical linkage between receptor and cytos keleton, We present a flow cytometric approach to analyze integrin rec eptors that are physically linked to the cytoskeleton. Epithelial cell s were suspended and extracted with Triton X-100 containing lysis buff er to obtain the detergent-insoluble cytoskeletal fraction. To detect immobilized receptors, the fractions were incubated with antibodies ag ainst the receptors, We were able to measure these cytoskeletons as si ngle particles in now cytometry, The extracted fractions revealed dist inct lower forward and side light scatter intensities compared with no rmal cells. Our results demonstrated that integrin receptor cross link ing induced their association to the cytoskeleton, Incubation of cells with a receptor antibody alone had no effect, We conclude that flow c ytometry enables the evaluation of the receptor-cytoskeleton linkage o n the basis of objective fluorescence data and on a single cell level. (C) 1996 Wiley-Liss,Inc.