Obtaining Clean Sections
This is one in a series of web pages offered as an aide
to electron microscopists, with emphasis on the special requirements
of neuroscience studies. The purpose of this page is to provide a
few tips for obtaining clean thin sections.
A common problem is a deposit on sections that is not quite black,
and that appears particulate at lower magnifications, but that is
shown to be somewhat transparent at higher ones.
Problem: an oily deposit.
Not obvious causes:
- using too much "wetting agent" in the boat water.
- trimming the block with an unclean razor blade.
Recommendations:
- When trimming the block make final cuts with a razor blade that
has been cleaned with acetone and alcohol to avoid any razor blade
grease from getting onto the block. Note: trimming the leading and
trailing edges of the block with a glass knife facilitates the
formation of
ribbons
of sections.
- Use as little wetting agent as possible during cutting.
- Rinse grids 10 seconds or so with 70% alcohol or water
before staining with lead citrate.
Either type of pre-rinse cleans grids well from any oil; alcohol
causes no harm to epon/araldite embedded material, and it should
result in a cleaner preparation than would a water pre-stain rinse,
but this has not been thoroughly investigated.
Additional precautions:
- wear gloves during every step involved in grid preparation.
- rinse every item placed into the boat during sectioning with
alcohol and water (except perhaps grids which should have been
cleaned and dried previously).
- I believe that distilled water provides consistently cleaner
preparations than with filtered water.
Notes:
As with any laboratory procedure, one or another step or
recommendation may be unnecessary; always best to omit whatever one
can.
This page prepared by Ed. White who believes scientists
should not be intimidated by the prospect of thin sectioning. For
questions or clarifications contact Ed. White at: wisrael@ bgumail.bgu.ac.il
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