Freeze-fracture cytochemistry,a new fracture-labeling method for topological analysis of biomembrane molecules

Abstract

Freeze-fracture cytochemistry allows visualization of cellular and molecular characteristics of biomembranes in situ. In this review, we discuss freezefracture cytochemistry with special reference to a new cytochemical labeling of replicas, the detergentdigestion fracture-labeling technique. In this procedure, unfixed cells are rapidly-frozen, freeze-fractured, and physically stabilized by evaporated platinum/carbon. The frozen cells are then removed from the freezefracture apparatus to thaw and are subsequently treated with detergents. After detergent-digestion, replicas are labeled with cytochemical markers. We demonstrate that the technique is a versatile tool for direct analysis of the macromolecular architecture of biomembranes and allows identification of particular intracellular membrane organelles. In addition, we demonstrate the application of ultrasmall gold to freeze-fracture immunocytochemistry. Freeze-fracture cytochemistry is a valuable technique for investigating topology and dynamics of membrane molecules.