Ronnie
the methyl benzoate celloidin really only works well with double embedding and with this there will be considerable shrinkage as the celloidin only partly infiltrates, it mostly encases the tissue to hold structures together. The paraffin wax after this still causes considerable shrinkage.
Amyl acetate can be used in place of alcohol ether as the solvent. Once infiltration is complete the solvent can be gradually removed I believe by using alcohol:chloroform mixture.
Barry

Ronnie Houston wrote:

> Baryy,
> I'm resending this as it appears we lost at least half a day's messages.
>
> I used to work with Billy, and yes the shrinkage was measured compared to both frozen and paraffin sections, albeit by a fairly archaic method. Don't recall if it was ever published, perhaps in an old, old edition of Med Lab Sciences or the Institute' Gazette; seem to recall Norman Russell's or Gordon Gentles' names in print. I'll see if I can find something.
>
> I remember amyl acetate being used somewhere in a double-embedding cycle, believe it was before methyl benzoate celloidin (boy this really takes me back!)
> Ronnie
>
> Ronnie Houston
> Cytochemistry & Molecular Pathology
> Texas Scottish Rite Hospital for Children
> 2222 Welborn Street
> Dallas, TX 75219
> (214) 559 7744
> (214) 559 7768 - fax
> ronnie.houston@tsrh.org
>
> >>> Barry Rittman  12/04/01 11:04AM >>>
> Bill
> while I agree with most of your comments here, I am not sure about the shrinkage
> that you quoted. Was this measured and/or is this personal experience. I have
> not processed a lot of brains in celloidin but have processed several other
> tissues and the sort of shrinkage that was seen and those in the literature are
> well below 10%, usually around 5%. It has been stated somewhere that a 12 micron
> celloidin section contains the same volume of tissue as a 7 micron paraffin
> section, this difference due to the differences in shrinkage between the two
> processing methods necessary.
> Perhaps this shrinkage is specific to the brain?
> As an aside, celloidin can also be dissolved in amyl acetate instead of
> alcohol:ether.
>
> To Billie
> Celloidin and LVN (Low Viscosity Nitrocellulose) blocks as has been mentioned
> are slow to infiltrate, difficult to prepare and process and technically
> difficult to section and to stain. We estimated cost to be around $200 plus for
> block of 2 teeth in a dog jaw.
> You do not state why you are considering this embedding medium?
> Barry
>
> "McMeekin, Bill" wrote:
>
> > Billie
> >
> > 25 years ago I worked in a neuropathology lab where we used celloidin
> > embedding for large brain sections.  We could obtain 2 concentrations of
> > necoloidine solution (8% and 14% from memory) at that time and prepared a
> > graded series from these in ether/alcohol (1:1).  Fixed tissue was processed
> > to absolute alcohol followed by ether alcohol and then infiltrated with
> > increasing strengths of celloidin solutions up to the 14% max.  The
> > viscosity of the thicker celloidin solutions necessitated a very slow
> > process (changes at weekly intervals).  Embedding was then carried out in
> > large flat-bottomed, circular glass troughs with a glass plate for a lid.
> > After placing blocks in fresh 14% celloidin in the trough (poured to a depth
> > of perhaps 4-5cm) they were allowed to settle.  The celloidin had to be
> > allowed to thicken very gradually by sliding the lid aside to create a small
> > vent, letting the ether/alcohol evaporate slowly.  Periodically, gas bubbles
> > had to be encouraged from under the blocks (to avoid distortion) by
> > judicious use of pressure and lifting.  Care was required to prevent a
> > 'skin' forming on the surface (by overzealous venting of solvent), leaving a
> > soft underside which was difficult to recover.  When the celloidin had
> > reached a firm, rubbery consistency, throughout its depth (assessed by
> > probing), it could be removed from the trough in one piece and individual
> > blocks trimmed using a scalpel.  Preparations were elegant and very
> > satisfying to produce.
> >
> > Some of the drawbacks:  The process was very time consuming and costly.
> > Shrinkage is significant (25-30% ?).  Ether is highly flammable and
> > explosive as a vapour (and you can get rather attached to it).  Blocks need
> > to be kept under 70% alcohol to limit hardening and this needs to be
> > regularly topped up if storing long term.  The blocks remain highly
> > flammable and make for bulky storage.
> >
> > We do not use it in my current lab and I would be loathe to introduce it for
> > some of the reasons stated.
> > Bill McMeekin
> > Senior Chief Biomedical Scientist
> > Neuropathology
> > Newcastle General Hospital
> > Phone: +44 191 256 3830
> > Fax: +44 191 256 3196
> > E-mail: bill.mcmeekin@nuth.northy.nhs.uk
> >
> >
> >  -----Original Message-----
> > From: Billie Zimmerman [mailto:bzimmerm@mail.mcg.edu]
> > Sent: 03 December 2001 15:29
> > To: histonet@pathology.swmed.edu
> > Subject: Celloidin
> >
> > Does anyone have a procedure for using celloidin an embedding media?
> >
> > Thanks,
> > Billie Zimmerman
> > MCG