Re: Staining Spurr's???
<< Previous Message | Next Message >>
From: | tflore@lsumc.edu (Flores, Teresa) (by way of histonet) |
To: | histonet <histonet@magicnet.net> |
Reply-To: | |
Content-Type: | text/plain; charset="us-ascii" |
Hildegard, Our EM lab has been using Spurr epoxy for the past 20 years.
Please provide your "get rid of spurr" with references. Thanks, Teresa
>------------------------------------------------------------------------
>The Microscopy ListServer -- Sponsor: The Microscopy Society of America
>To Subscribe/Unsubscribe -- Send Email to ListServer@MSA.Microscopy.Com
>On-Line Help http://www.msa.microscopy.com/MicroscopyListserver/FAQ.html
>-----------------------------------------------------------------------.
>
>
>Hi,
>
>Spurr's is the most highly crosslinked of all embedding media used for
>LM-TEM. The higher the crosslinkage, the less likely any stain, whether
>for LM or heavy metal for TEM, is to penetrate well enough to be called
>successfull.
>First: Get rid of Spurr's. It contains a potent carcinogen, VCD. Do you
>need this?
>Second: If you cannot get rid of Spurr's (try real hard), then use the
>softest mixture you can tolerate for cutting. Polymerize at 60deg C
>overnight, and see if this is adequate. This will reduce the final
>crosslinkage.
>Third: Blocks in existence already! Don't stain well? Try soaking the
>sections prior to staining for extended periods of time in water, then 5
>min in alcohol, in increasing concentrations until you reach 95%. Then go
>back down to water stepwise. Try staining.
>Fourth: Use as alkaline as possible a vehicle for your stain. pH 12 is
>about right.
>Fifth: Combine all of the above. Does not work? Soak the sections in
>water and gradually bring to 95% alcohol.. Expose to stain dissolved in
>alcohol. Does not work? Forget it. Start over.
>
>If you have very valuable sections and you must stain them for TEM, use
>alcoholic UA for 10min at 60deg C. Use Reynolds lead citrate at a pH of
>about 9 or 10. This last trick is truly a last resort, since the lead may
>dump erratically (or stain easily) at this low pH.
>
>If you polymerize a block at a low power for 45 minutes in a microwave,
>you crosslink the resin to such an extent that nothing, nothing, nothing
>you do will stain it. (Lost my best meat loaf dish this way). For TEM we
>do not polymerize resins totally, about 10% of the monomers are left
>unreacted with one another. If you "drive" the crosslinkage, the block
>will be harder, less elastic, and impenetrable for liquids (except if you
>boil it for a year or so in water). I am not exaggerating. I got this
>info out of one my very favorite materials science books.
>
>Don't use Spurr's!
>
>
>Bye,
>hildy
<< Previous Message | Next Message >>