Labeling properties of sucrose-infiltrated cryosections

Plastic transverse sections of labeled semithick and ultrathin cryosections prepared by the Tokuyasu procedure demonstrate that section surfaces can be smooth or rough, sometimes showing fracture-like structures. These surface irregularities can arise from specimen properties, sectioning/fracturing and compression during sectioning, loss of cytoplasmic material during labeling, and from the final drying step. Nevertheless, small and sensitive marker molecules like 4 nm gold-protein A, ferritin-IgG and 1 nm gold-IgG bind predominantly to the cryosection surface of even mildly fixed specimens as long as the section is well preserved. In our experiments, marker penetration is enhanced by section damage. Marker incubation of ultrathin cryosections obtained by cryo transverse sectioning of IgG-labeled semithick cryosections shows also that penetration of uncoupled antibodies into cytoplasm and nuclei of chicken erythrocytes is limited.