[Histonet] New to paraffin cutting - seeking advice

[Marc DeCarlo]

Merissa,

I can't help with your paraffin needs, but have a suggestion for your
plastic.  Try Weigerts Hematoxylin.  It's available commercially through
several vendors and in my experience stains just like what your looking for.

If you need help just contact me.

Marc

On Monday, April 9, 2012, M.O. wrote:

> Hello everyone! I have had such wonderful feedback regarding plastic
> embedded specimens and now I am moving onto paraffin.  The majority of the
> specimens I am going to be cutting are paraffin embedded decal rabbit
> femurs.  I am using a Leica RM2255 and there is an option for retraction
> (after each section) and blade angle.  I need to practice a lot because
> right now my sections are crinkly even though I have my samples on ice, but
> hopefully practice makes perfect!
>
> So I want to ask you all a few questions regarding settings and technique
> and one about H&E staining.  What angle should the blade be at - I have it
> set at 0 for the moment.  What about the retraction option - I have it set
> at 15um.  The sections are being cut at 4um.
>
> When the blade has pieces of paraffin on it, do I need to clean that off?
> How do I remove this paraffin because I don't want to dull the blade? If
> not, does that impact the section quality and should I be moving to a
> different area of the blade?
>
> I am looking into flotation baths that are relatively inexpensive.  In
> particular, I am looking for a simple bath with a glass pyrex dish.  Do you
> have any suggestions on where to purchase one from?
>
> Lastly, I am going back to H&E staining of plastic embedded undecalcified
> bone, specifically hematoxylin, for a brief moment.  When I use a clearing
> solution I know that I am trying to destain a bit from the undecalcified
> bone to make a lighter stain, but does this also significantly destain the
> nuclei?  Right now, I need to leave the sample staining in Harris's
> hematoxylin for more than 4 mins to get the nuclei stained nicely, but I
> need to destain because the section is too dark. I just don't want to
> destain the nuclei too much, but want a lighter stain on the undecal bone.
>
> Again, thank you all for your support and advice, it is much appreciated!
>
> Sincerely,
> Merissa
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