A procedure for rupture‐free preparation of confluently grown monolayer cells for scanning electron microscopy

Abstract

Abstract Monolayers of PtK‐1 and HeLa cells grown on glass or plastic supports are extremely susceptible to lacerations, e.g., splits and cracks caused mainly by shrinkage when prepared for scanning electron microscopy (SEM). We find that a four‐step fixation procedure including glutaraldehyde, OsO 4 , tannic acid, and uranylacetate application, in combination with critical point drying, drastically reduces these structural damages. In addition, the conductivity of the specimens is enhanced, so that they can be investigated without gold coating. Transmission electron microscopy (TEM) investigation of perpendicular sections in the area of lacerations provides evidence that the subcortical cytoskeletal elements are of crucial importance in maintaining cell membrane stability during the preparations. Our relatively quick and simple procedure results in an improved structural appearance of the cells.