Tim
Sounds to me that you have a cutting problem or problems, with thickness variation and lines.
I am assuming that your tissue is relatively uniform in consistency?

I suggest problem might be one of the following.
1.	Is the knife securely clamped into the holder? If there is any movement at all on one side this might explain the thickness differences.
2.	Does the holder have wear on one side, either the knife holder or the sliding mechanism it sits on? 
3.	Are these blades reliable as regards having a sharp edge along entire edge? Rene's suggestion should take care of this.
4.	I do not believe that the wax is the problem unless you have poorly mixed wax or separation of the components. If this is the problem then this effect should be seen on different sides of different blocks in a random manner.
If this wax is too hard then Rene's suggestion of beeswax should help
Would you be kind enough to let us know what size blocks you are cutting?
thanks
Barry

-----Original Message-----
From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Rene J Buesa
Sent: Wednesday, January 17, 2007 2:53 PM
To: Tim Wheelock; Histonet@lists.utsouthwestern.edu
Subject: Re: [Histonet] LINES IN PARAFFIN SECTIONS

Tim:
  I would try first changing to another brand of disposable (JUST to see if there is a difference).
  If the problem persists I would return to your usual disposables and then try adding some bees wax (5%) to your paraffin (JUST to see if there is a difference). 
  If the problem remains it could be some dust in the paraffin or in the molds; have you had problems with the air filters in your lab?
  Just some ideas to try to isolate the problem.
  I hope this would help you!
  René J.

Tim Wheelock  wrote:
  Hi Everyone:

I have a serious problem with lines/splits forming in my 5 micron 
sections as they come off the microtome.
I just started using Paraplast Xtra Tissue Embedding Medium.
My infiltrating wax is regular Paraplast.
My disposable blades are Surgipath high profile Teflon coated blades.
The microtome is a Microm HM 355S.
I cut human brain tissue exclusively.

I had been using Surgipath embedding media.
I decided to try the X-tra because I had been having problems with the 
sections being thick and thin on just one side of the section.
Also the wax seeming to be thicker (more opaque) on one side of the block.
The blocks of tissue also were not laying flat in the mold, hence a lot 
of trimming after.
There was some problem with lines even with the Surgipath but it was 
intermittent., and not nearly as bad as the X-tra.

I notice that trimming requires more physical effort with the Paraplast 
X-tra.
And I have had much more of a problem with lines than with the Surgipath 
Embedding Media.
Does it have a higher % of plastic in it.

I cleaned out the reservoir on the embedding center and played with the 
knife angle but to no avail.
I am not sure if the lines have to do with the wax or the blades or both.

Thank you for any advice you can give me.

Tim Wheelock
Harvard Brain Tissue Resource Center
McLean Hospital

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