[Histonet] Re: floating petri dish snap freezing question

[Greg Dobbin]

Dear Kristen/ Gayle,
To see my detailed reply to a similar question on cryotomy, check 
the following reference:
Biotechnic & Histochemistry 2003, 78(5): 279.
The "Notes and Queries" section.
Cheers!
Greg

Date sent:      	Wed, 11 May 2005 10:17:30 -0600
To:             	"Kristen Broomall" <kbroomal <@t> NEMOURS.ORG>,
	Histonet <@t> lists.utsouthwestern.edu
From:           	Gayle Callis <gcallis <@t> montana.edu>
Copies to:      	Subject:        	[Histonet] Re: floating petri dish snap freezing question

> Kristen,
> 
> I have done it both ways but if you let the tissue float up, it is
> harder to find in the OCT during sectioning.   I often put a very thin
> OCT layer in mold then add tissue to have it located in bottom - then
> add OCT. After than you put mold in canoeing petri dish.  It is
> sometime hard to push a delicate or several pieces into OCT and sort
> of "glues" them to bottom of mold.
> 
> Another trick: Put tiny drop of OCT on a dark surface, add biopsies,
> then roll them into a tiny OCT ball, pick up (we like a curved eye
> forceps for this, center tissue/OCT into center/bottom of mold but
> when you add more OCT, go AROUND the little ball of tissues.  The
> pressure of adding gooey OCT around tissues tends to keep everything
> centered.  You tend to lose orientation with this method, something
> you may not like with gastric bx's.
> 
> You can dip tiny tissue in OCT, add to mold so it stays oriented, then
> add OCT slowly and gently. Whatever you do, keep tip of OCT bottle (by
> laying bottle on side) filled to prevent bubbles - the enemy.  tip of
> bottle can be cut off so hole is small and prevent huge flow of goo
> when dispensing OCT. One can always add OCT during freezing - so watch
> bottom turn white, but key word here is is during not after freezing, 
> you do not want an interface of two OCT layers, they will snap part
> during sectioning at times.  Been there, done that, and it was
> disaster.
> 
> We purchase eye forceps for embedding in OCT - these are fine round
> tips, with either straight, half curved or full curved tips.  Sharp
> points are NOT used, to easy to poke holes in tiny tissues.   Arista
> in New York has huge selection and cheap.
> 
> Another thing we are using is safety glasses that fit over
> prescription glasses, but the safety glasses have bifocal
> magnification - wonderful to see when handling tiny tissues.  They
> come in 1, 1.5, 2.5 and 5 magnification - maybe other,  available from
> Fisher, Newcomer Supply, and MarketLab.
> 
> Hopefully this helps, so many ways.
> 
> Have a good day
> 
> At 06:46 AM 5/11/2005, you wrote:
> >Gayle,
> >
> >I tried out your snap freezing method with the floating petri dish in
> >the liquid nitrogen yesterday for some itsy bitsy GI biopsies. I
> >really like it! I have a question though. When using this method, do
> >you place your sample in the bottom of your mold, then OCT on top, or
> >put a dab (or more) of OCT, then the tissue, ... or does it really
> >matter?
> >
> >Thanks,
> >
> >Kristen Broomall, HT (ASCP)
> >
> >
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> 
> 
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~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
Greg Dobbin 
Research Technologist,
Pathology Lab,
Atlantic Veterinary College, U.P.E.I.
550 University Ave.
Charlottetown, P.E.I.
Canada,  C1A 4P3
Phone: (902)566-0744
Fax: (902)566-0851
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
Happiness is a journey, not a destination.