Imaging of immunolabeled membrane receptors in uncoated sem specimens

Abstract

Summary: Epidermal growth factor receptors (EGFR) were labeled with I0 n m immunogold and examined on uncoated specimens of A43 I human epidermoid carcinomacells. A field emission gun and a high-sensitivity YAG ring detector were used to demonstrate the affinity labeling simultaneously in the secondary-electron (SE) and backscattered-electron (BSE) modes with a low accelerating voltage (V,). At V0=2 kV, the SE and BSE signals were too weak to identify all markers, while at Vc,=3-7 kV labeling was observed unambiguously in both the SE and BSE modes with smaller and higher working distances. Increaing the V, to above 7 kV sometimes provokes instability of the specimens. A V,, of> 10 kV produces charging artifacts in the SE image, but permits a BSE image of the gold markers providing additional topographic information. In conclusion, immunogold labeling can be used with good results for uncoated specimens.