Fluorescent staining of microfilaments with heavy meromyosin labeled with N-(7-dimethylamino-4-methylcoumarinyl) maleimide.

Abstract

For fluorescent staining of microfilaments in cells, heavy meromyosin (HMM) or subfragment-1 (S-1) was labeled with a novel thiol-directed fluorescent dye, N-(7-dimethylamino-4-methylcoumarinyl) maleimide (DACM), instead of the usual dyes, such as fluorescein-isothiocyanate (FITC). DACM-labeled HMM or S-1 gave characteristic fluorescence patterns to a variety of cell types similar to those reported with the use of FITC-labeled HMM or S-1 or with immunofluorescence techniques using anti-actin antibody. The fluorescence of DACM was fairly photoresistant as compared with FITC, so that HMM or S-1 required only 1 mol of the dye per myosin head. Consequently, F-actin need not be used to preserve the actin binding activity of the myosin fragments when labeling with the dye.