A simple and reliable quick-freezing/freeze-fracturing procedure

We describe a simple method for the quickfreezing/freeze-fracturing of cells in tissues or culture monolayers. Tissue slices or cultured cells were covered with thin copper foil (10-mu m-thick), and frozen by smashing them against a liquid helium-cooled copper block. Freeze-fracturing was accomplished by mechanically separating the copper foil from the frozen specimen. The fracture faces were replicated by platinum and carbon. Replicas were processed for conventional electron microscopic observation or cytochemical labeling. This method allows the ultrastructural and cytochemical examination of large areas of fractured membrane without chemical fixation.