Serial Thick Section Gas Cluster Ion Beam Scanning Electron Microscopy (original) (raw)

Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) is used to volume image heavy metalstained, plastic-embedded biological samples with resolutions below 10 x 10 x 10nm, an ability that is especially important in connectomics [1]. FIB-SEM samples are typically restricted to be <50µm in the direction of the FIB beam because glancing angle milling results in artifacts over longer distances [1]. Removal rate is also restricted due to a current/spot size tradeoff. These limitations are especially problematic when one contemplates combining FIB with the increased speed offered by multibeam SEMs like the 91 beam Zeiss MultiSEM [2]. The MultiSEM's minimum field of view is ~180µm, and its imaging rate is approximately two orders of magnitude faster than FIB's milling rate. These considerations appear to preclude the integration of traditional FIB milling with MultiSEM imaging.