Automated in‐chamber specimen coating for serial block‐face electron microscopy

When imaging insulating specimens in a scanning electron microscope, negative charge accumulates locally (‘sample charging’). The resulting electric fields distort signal amplitude, focus and image geometry, which can be avoided by coating the specimen with a conductive film prior to introducing it into the microscope chamber. This, however, is incompatible with serial block‐face electron microscopy (SBEM), where imaging and surface removal cycles (by diamond knife or focused ion beam) alternate, with the sample remaining in place.

[1]  W. Denk,et al.  Staining and embedding the whole mouse brain for electron microscopy , 2012, Nature Methods.

[2]  N. Kasthuri,et al.  ATUM-based SEM for High-Speed Large-Volume Biological Reconstructions , 2012, Microscopy and Microanalysis.

[3]  Kei-ichiro Nakamura,et al.  Beam deceleration for block-face scanning electron microscopy of embedded biological tissue. , 2012, Micron.

[4]  Kevin L. Briggman,et al.  Wiring specificity in the direction-selectivity circuit of the retina , 2011, Nature.

[5]  R. Tsien,et al.  Enhancing Serial Block-Face Scanning Electron Microscopy to Enable High Resolution 3-D Nanohistology of Cells and Tissues , 2010 .

[6]  T. Vystavěl,et al.  Improving SEM Imaging Performance Using Beam Deceleration , 2009, Microscopy Today.

[7]  P. Echlin.  Handbook of Sample Preparation for Scanning Electron Microscopy and X-Ray Microanalysis , 2009 .

[8]  David C. Joy,et al.  A new examination of secondary electron yield data , 2005 .

[9]  W. Denk,et al.  Serial Block-Face Scanning Electron Microscopy to Reconstruct Three-Dimensional Tissue Nanostructure , 2004, PLoS biology.

[10]  E Suzuki,et al.  High‐resolution scanning electron microscopy of immunogold‐labelled cells by the use of thin plasma coating of osmium , 2002, Journal of microscopy.

[11]  J. Thong,et al.  Single-image signal-to-noise ratio estimation. , 2006, Scanning.

[12]  W. Yao,et al.  The interface diffusion and reaction between Cr layer and diamond particle during metallization , 2001 .

[13]  I. Müllerová,et al.  Strtegies for low-and ver-low-energy SEM , 1999 .

[14]  C. Mathieu,et al.  THE BEAM-GAS AND SIGNAL-GAS INTERACTIONS IN THE VARIABLE PRESSURE SCANNING ELECTRON MICROSCOPE , 1999 .

[15]  W. M. Haynes.  CRC Handbook of Chemistry and Physics , 1990 .

[16]  D. Joy.  Control of charging in low-voltage SEM , 1989 .

[17]  J. Cazaux,et al.  Some considerations on the electric field induced in insulators by electron bombardment , 1986 .

[18]  L. Reimer,et al.  Scanning Electron Microscopy , 1984 .

[19]  James Pawley,et al.  Low voltage scanning electron microscopy , 1984, Journal of microscopy.

[20]  L. Reimer.  Scanning Electron Microscopy: Physics of Image Formation and Microanalysis , 1984 .

[21]  Leighton Sb.  SEM images of block faces, cut by a miniature microtome within the SEM - a technical note. , 1981 .

[22]  S. B. Leighton.  SEM images of block faces, cut by a miniature microtome within the SEM - a technical note. , 1981, Scanning electron microscopy.

[23]  J Walton,et al.  Lead asparate, an en bloc contrast stain particularly useful for ultrastructural enzymology. , 1979, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[24]  L. B. Harris,et al.  Charge neutralisation of insulating surfaces in the SEM by gas ionisation , 1978 .

[25]  V. Robinson.  The elimination of charging artefacts in the scanning electron microscope , 1975 .

[26]  R. Wilson,et al.  Modified thiocarbohydrazide procedure for scanning electron microscopy: routine use for normal, pathological, or experimental tissues. , 1975, Stain technology.

[27]  T. J. Shaffner,et al.  'Charging' effects in the scanning electron microscope , 1971 .

[28]  M. Wineberg,et al.  The use of an electron beam evaporation source for electron microscope sample preparation , 1971 .

[29]  W. Nixon,et al.  Retarding field scanning electron microscopy. , 1968, Journal of scientific instruments.

[30]  J. Terzakis.  Uranyl acetate, a stain and a fixative. , 1968, Journal of ultrastructure research.

[31]  A. Seligman,et al.  A NEW STAINING METHOD (OTO) FOR ENHANCING CONTRAST OF LIPID-CONTAINING MEMBRANES AND DROPLETS IN OSMIUM TETROXIDE-FIXED TISSUE WITH OSMIOPHILIC THIOCARBOHYDRAZIDE (TCH) , 1966, The Journal of cell biology.