Field emission scanning electron microscopy (FESEM) entering the 21st century: Nanometer resolution and molecular topography of cell structure

The field of scanning electron microscopy (SEM) has undergone a revolution in terms of resolution obtainable on biological samples in the last 4 decad es. In the late 1960s, the introduction of electron guns using tungsten fil aments for SEM provided resolution on the order of 10-20 nm and valuable in formation on cell shape and morphology was obtained that had hitherto been inaccessible. In the late 1980s, the development of stable cold field emiss ion SEM (FESEM) with in-lens specimen positioning and above the lens collec tion for secondary electron (Se) imaging provided a tremendous advance with resolution in the nm range being possible. Accompanying these changes in r esolution have been improved procedures in thin film technology with metal film thickness being reduced from 10-20 nm to less than 1 nm for FESEM. Dev elopments in cryoSEM have led to interpretable structure on biological samp les of about 1-2 nm. Improvements in instrumentation, refinements in ultrat hin film technology, and specimen preparation may lead to atomic level reso lution exceeding 1 nm in the next decade.