Immunoelectron microscopy in embryos

Immunogold labeling of proteins in sections of embryos embedded in acrylate media provides an important analytical tool when the resolving power of th e electron microscope is required to define sites of protein function. The protocol presented here was established to analyze the role and dynamics of the activated protein kinase C/Rack1 regulatory system in the patterning a nd outgrowth of limb bud mesenchyme. With mi nor changes, especially in the composition of the fixative solution, the protocol should be easily adapta ble for the postembedding immunogold labeling of any other antigen in tissu es of embryos of diverse species. Quantification of the labeling can be ach ieved by using electron microscope systems capable of supporting digital im age analysis. (C) 2001 Academic Press.