Rat Skeletal Muscle Tissue Labeled with Wheat Germ Agglutinin and Phalloidin

In order to localize a red fluorescent tag to F-actin in the sample of rat skeletal muscle tissue presented in the digital image above, the specimen was labeled with Alexa Fluor 568 conjugated to a phallotoxin (phalloidin). Oregon Green 488 conjugated to the lectin wheat germ agglutinin, which selectively binds to N-acetylglucosamine and N-acetylneuraminic residues, was also applied to the tissue sample, as was the nuclear counterstain Hoechst 33342. Images were recorded in grayscale with a 12-bit digital camera coupled to a Nikon Eclipse 80i microscope equipped with bandpass emission fluorescence filter optical blocks. During the processing stage, individual image channels were pseudocolored with RGB values corresponding to each of the fluorophore emission spectral profiles.