Scott,
This may just sound like more of what you have already heard, but all this 
extra fixation and processing is extremely inefficient when cutting a 
section thin in the first place is all that is required. 
Processing/fixation problems with thick tissues are compounded by tissues 
being squeezed into cassettes where the surface of the tissue is pressed 
against the inside of the cassette or in some cases even squeezing out 
through the slots. This contact with the cassette effectively reduces the 
surface area of the tissue by as much as 70-80% in extreme cases. (For the 
unconverted experiment with a thick slice of cellulose sponge pressed into 
a cassette and a thin one that moves freely inside a cassette, drop both 
into a dish of water, the one with the thin sponge "section" will bob and 
sink, the one with the thick section will float almost indefinitely). If 
you are going to fix breast samples overnight, breadloaf the specimen and 
fix it before blocking. This will make it easier to cut thin sections in 
the first place. I am unfamiliar with clear-rite but many of the xylene 
substitutes do not share its power as a lipid solvent. Ethanol is a poor 
lipid solvent and can become saturated rapidly. (take a sample from your 
100% container in a glass cylinder and add 1-2ccs of water. If the ethanol 
turns cloudy, it is probably saturated and will not dissolve any more fat.) 
If your clearing agent is a poor fat solvent, then you just carry the fat 
into the paraffin where it will not infiltrate. I also donot understand why 
you return to formalin after the tissues have spent several hours in 
acetone. The methanol and acetone have at least partially dehydrated you 
tissues, returning to formalin serves little purpose-carry them on to 
absolute and save some time.

Bert

-----Original Message-----
From:	Weems, Joyce [SMTP:JWEEMS@sjha.org]
Sent:	Tuesday, February 08, 2000 12:00 PM
To:	'Scott, Allison D'; histonet@pathology.swmed.edu
Subject:	RE: Processing protocol for breast specimens

Some of the pointers (all collected from the net) is to put a xylene 
between
the last two 100% alcohols on the processor. This helped a lot! Then, if 
the
tissue is still raw - press it between paper towels to extract as much
moisture as possible and let it set in paraffin for a while to 
reinfiltrate.
This has saved quite a bit of reprocessing.
Joyce Weems
Pathology Manager
Saint Joseph's Hospital of Atlanta


	-----Original Message-----
	From:	Scott, Allison D [SMTP:Allison_Scott@hchd.tmc.edu]
	Sent:	Monday, February 07, 2000 4:27 PM
	To:	histonet@pathology.swmed.edu
	Subject:	Processing protocol for breast specimens

	Does anyone have a protocol for processing breast specimens.  We get
breast
	cases at least 2 to 3 times a week.  Our pathologist is not very
happy about
	the turnaround time in getting the case out.  Our first problem is
that the
	tissues were cut in to big and thick. our residents donot listen to
reason
	or follow instructions very well.  So therefore we have to work with
some
	very big fatty pieces of tissue.  We are putting the cassettes into
Rapid
	Fixx solution(fixative for fatty tisue), for overnight fixation.
The next
	day, they are put into methanol for 2 to 3 hours. After the
methanol, we put
	them into acetone for the remainder of the day.  They are then
loaded on the
	VIP for 2 changes of formalin, 2 changes of 95% alcohol, 4 changes
of 100%
	alcohol, clear rite 3 and then paraffin.  These are for 1 hour each.
In
	some instances we still have to run some cassettes back because they
were
	still not  processed well. If someone has a suggestion please
reply.#000##000#