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Fluorescence photoconversion of biarsenical-labeled cells for correlated electron microscopy (EM).

Cold Spring Harbor protocols

Arsenicals, diagnostic use, Cell Line, Fluorescent Dyes, Image Processing, Computer-Assisted, methods, Microscopy, Electron, Recombinant Proteins, analysis, Staining and Labeling

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      Abstract

      Correlated light microscopy (LM)/electron microscopy (EM) analysis can be achieved by using biarsenical dyes to fluorescently label tetracysteine-tagged proteins. Once live cell imaging using LM is complete, cellular activity can be halted promptly using a glutaraldehyde-based fixative. Rapid fixation preserves cellular ultrastructure and limits diffusion of reaction products. This protocol provides details on rapid fixation of cells, followed by fluorescence photoconversion of 3,3'-diaminobenzidine tetrahydrochloride (DAB) and sample processing for EM that can be correlated with the live cell LM images.

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      21205848

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