Prevention of electron beam transmittance for biological cell imaging using electron beam excitation-assisted optical microscope

Abstract

We demonstrated the high-spatial-resolution imaging of label-free biological cells using an electron beam excitation-assisted optical (EXA) microscope without irradiation damage by the electron beam. An EXA microscope can be used to observe a specimen with a nanometric light source excited in the \(\mathrm{Si}_{\mathrm{3}}\mathrm{N}_{\mathrm{4}}\) membrane by an electron beam. The incident electron beam penetrates the \(\mathrm{Si}_{\mathrm{3}}\mathrm{N}_{\mathrm{4}}\) membrane and damages the specimen. To suppress the irradiation damage of the specimen, we prevented the transmittance of the electron beam by coating the \(\mathrm{Si}_{\mathrm{3}}\mathrm{N}_{\mathrm{4}}\) membrane with a gold thin film. To obtain an electron beam transmittance through the \(\mathrm{Si}_{\mathrm{3}}\mathrm{N}_{\mathrm{4}}\) of 0%, a gold film of 15 nm thickness was required. By adding the gold layer, a label-free cellular structure was observed with 135-nm spatial resolution.