Localization of membrane components by freeze-fracture replica immunolabeling (FRIL).

The freezefracture technique provides large and detailed ultrastructural views of cellular membranes in the transmission electron microscope. The major goal of freezefracture cytochemistry is the identification and localization of membrane components such as proteins or lipids for studying their distribution, dynamics or their relationship to specialized membrane domains. The most versatile and effective technique in freezefracture cytochemistry is termed freezefracture replica immunolabeling (FRIL). Using this technique chemically unfixed cellular samples are rapidly cryofixed in their natural habitat. They are fractured, replicated and immobilized by platinumcarbon evaporation followed by a careful sodium dodecyl sulfate (SDS) treatment. The detergent SDS removes most of the cellular material except those molecules that are in direct contact to the platinum/carbon replica. Immunogold labeling of these replicabound biomolecules visualizes their spatial organization in high resolution on planar views of cellular membranes. [ABSTRACT FROM AUTHOR]