Re: bone marrow trephines

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From:Katri Tuomala <> (by way of histonet)
To:histonet <>
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Mick Rentsch wrote:
>  Dear Marjorie,
> we've been using FormalSaline/Formic Acid for more years than I can remember
> on all our trephines. At the time of Aspiration we put the trephine directly
> into this Fixative/Decal for a minumum of four hours, but more often then
> not it's about 8 -10 hours as we normally do our own trephines at about
> 0800hrs. At four hours decalcifying is close to complete, and even though
> sometimes a bit "crispy" you can still get good sections. If you leave the
> trephines overnight in this Fixative/Decal, there doesn't appear to be much
> deterioration and detail is still excellent.
> It may well be that any acid decalcifying agent will be adverse for your
> studies including Trichloracetic Acid, while I've only used EDTA on rare
> occasions and then only to ensure the best possible detail, I only did so
> for PSA and AcP, which were excellent but required two days to decalcify the
> trephines.
> The recipe I use for  5 LitresFormal Saline/Formic Acid is as follows:-
> Sodium Chloride Analar 42.5g
> Formaldehyde 38/40% Analar 500mls
> Formic Acid 90-95% Analar 1600mls
> Make up to 5 litres with RO Water >4MOhm.
> Label as UN2810 Corrosive Liquid N.O.S. Decal Reagent. Give a two year
> expiry and pack in 1 Litre DGA approv'd bottles.
> Decant 35mls/ 70ml specimen jar labelled expressly for Bone Marrow
> Trephines.
> Hope this is of some help, Regards
> Mike Rentsch (Downunder)
> -----Original Message-----
> From: Hagerty, Marjorie A. <>
> To: Histonet <>
> Date: Friday, 13 November 1998 8:29
> Subject: bone marrow trephines
> >
> >Histonetters,
> >We are currently using RDO for one hour to decal our BM core bxs. This
> works
> >perfectly for us, they are easy to cut, and the nuclear detail is
> excellent.
> >I think, however, that the HCL in the decalcifier may be interfering with
> >our Kappy/Lambda immunoperoxidase staining. I tried an antigen retrieval
> >solution specifically for decalcified tissue but that did not seem to
> >correct the problem. Doing this on one case does not count as any kind of
> >"study" so this may be an excellent product. We would like to try a non-HCL
> >decalcifier. Alex metioned formal citrate but our clinicians would not
> >tolerate the length of time it takes. Does anyone know of a decalcifier
> that
> >takes no more than an hour for very small core bxs and does not use HCL?
> >Thanks
> >Marg
> >EMC, Rancho Mirage, CA
> >

Hi Mike (Downunder),
We use the same decalcifying solution you do, but always do a complete
fixation in 10% formalin first. I was always taught that the complete
fixation is a must before decalcifying. Is there a reference to this
fixative/decalcifyer and how it actually works? Obviously it is not
destroying your bone biopsies, but does the formalin still form
crosslinkages under these conditions therefore requiring some type of
epitope retrieval before doing IHC? It is funny, I've been working with
this solution for several years, without really questioning the role of
the formaldehyde in it, except that I was told it is there to prevent
tissue destruction in case the fixation was not quite complete. I enjoy
the cans of worms the Histonet opens! Live and learn!
Sincerely, Katri (Canada)

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