Chemical reactivation of quenched fluorescent protein molecules enables resin-embedded fluorescence microimaging.

ABSTRACT: Resin embedding is a well-established technique to prepare biological specimens for microscopic imaging. However, it is not compatible with modern green-fluorescent protein (GFP) fluorescent-labelling technique because it significantly quenches the fluorescence of GFP and its variants. Previous empirical optimization efforts are good for thin tissue but not successful on macroscopic tissue blocks as the quenching mechanism remains uncertain. Here we show most of the quenched GFP molecules are structurally preserved and not denatured after routine embedding in resin, and can be chemically reactivated to a fluorescent state by alkaline buffer during imaging. We observe up to 98% preservation in yellow-fluorescent protein case, and improve the fluorescence intensity 11.8-fold compared with un

SUBMITTER: Xiong H 

PROVIDER: S-EPMC4059927 | biostudies-literature | 2014 Jun

REPOSITORIES: biostudies-literature