High-resolution SE-I imaging of biological membranes using a Schottky field-emission in-lens SEM

Previous notes have described efforts to routinely collect quality SE-I image contrasts of biologically significant particulate membrane features (1-10 nm) in the context of whole cells and tissue fragments. Utilizing SEMs equipped with in-lens specimen stages and field emission sources (Schottky and cold cathode) operated at 15-30 kV and in conjunction with specimens coated with 1 nm Cr films (Z=24), nanometer resolution of biological samples may be attained. This note describes the definition of optimal electron source conditions for the high resolution imaging of cell membrane features μ 10 nm. The Schottky field emitter equipped ISI DS-130F SEM, in-house at the Yerkes Research Facility, was operated at 5-20 kV accelerating voltages (A.V.) and at 4 or 4.8 kV extraction voltages (E.V.). Although we have published images of soft and hard biological sections and isolated cells containing 1-10 nm particle contrasts by operating the SEM at 25-30 kV A.V., we have maintained 4 kV extraction voltage and not attempted using 4.8 kV above 25 kV A.V.