Preservation of protein fluorescence in embedded human dendritic cells for targeted 3D light and electron microscopy.

ABSTRACT: In this study, we present a correlative microscopy workflow to combine detailed 3D fluorescence light microscopy data with ultrastructural information gained by 3D focused ion beam assisted scanning electron microscopy. The workflow is based on an optimized high pressure freezing/freeze substitution protocol that preserves good ultrastructural detail along with retaining the fluorescence signal in the resin embedded specimens. Consequently, cellular structures of interest can readily be identified and imaged by state of the art 3D confocal fluorescence microscopy and are precisely referenced with respect to an imprinted coordinate system on the surface of the resin block. This allows precise guidance of the focused ion beam assisted scanning electron microscopy and limits the volume to be

SUBMITTER: Hohn K 

PROVIDER: S-EPMC4757415 | biostudies-literature | 2015 Aug

REPOSITORIES: biostudies-literature