Re: Mineralised Bone biopsies



I would like to reinterate that GMA can be a good resin of choice for Mineralized
Bone Biopsy work.  I find GMA to be easier to incorporate into a routine histo
lab than PMMA.  It is not as volatile and therefore less toxic when used in less
than adequate ventilation or chemical fume hoods.  Although it is not the resin
of choice for IHC because it cannot be removed like PMMA, you may not be doing
IHC on the mineralized bone samples anyway.  Most people are doing mineralized
bone histology to evaluate bone formation features for Metabolic Bone Disease
studies, such as Osteoporosis, Osteomalacia and Aluminum toxicity.  GMA will work
well for these studies including fluorchrome label evaluation and
histomorphometery measurements.  I think GMA has gotten a bad rap in the past
because people have used the embedding kits (JB4) which do not allow for
manipulation of resin hardness.  By making GMA yourself you will not only make it
much, much cheaper, but also be able to control how hard it sets up for your
needs.  It is extremely easy to make up on your own.  This is my nearly 30 years
worth of experience and plug for GMA for mineralized bone work.  You can do
fairly large, dense mineralized bone samples in GMA, I think very easily in a
routine histo lab.
Patsy Ruegg

Gayle Callis wrote:

> You can make up your own polymethylmethacrylate (PMMA, and preferred by
> many for bone, microtomed sections where PMMA can be removed for
> immunostaining, ie Neil Hand protocols).  You need tungsten carbide tipped
> blades, d profile, there are c profile available, d is excellent and a
> microtome that will be strong enough to do the cutting, Olympus has one (do
> not recall model) Leica 2155 and 2165.  As for a good sledge for
> mineralized bone in PMMA, a Polycut, but it is pricey as are all
> microtomes/knives for these protocols.
> It is cheaper to not use kits, and you can use PMMA methods where washing
> inhibitor away is avoided, getting rid of tedious, messy, handling of toxic
> substance is reduced.  You will need a fume hood vented to outside, a
> refrigerator, waterbath for polymerization.  Not sure if Technovits has a
> PMMA kit suitable for microtoming, if so, it would be worth a try.
> You can use an automated processor for dehydration and clearing, but not
> infiltration with PMMA, use to vacuum dessicators for hands on infiltration
> with PMMA.  The polymerization is best done in a waterbath for even
> heating, ovens do not provide even heating with this step.
> There are many publications in J of Histotechnology concerning these
> methods, including Hand's antigen retrieval, look for these authors, Diane
> Sterchi, J. Eurell, Cathy Sanderson (now Mayton), Chappell (hope I spelled
> that correctly).
> Working with undecalcified bone samples is more time consuming, there are
> no STAT protocols, or you end up with a mess of bubbly, polymerized goo.
> At 03:37 PM 2/27/02 +1100, you wrote:
> >Hi Histonetters,
> >
> >We are in the process of setting up a Mineralised Bone Histo section in our
> >Department and we need some advice. We are continuing the work of another
> >department who have handed over this job to us and would like to streamline
> >the process a bit. The way they make up their resin is long, tedious and
> >tricky, so that is the first thing I would like to "fix". Secondly we use
> >"spare time" on another departments' old sledge microtome to cut the bones
> >and I would like to purchase our own equipment to get away from having to
> >fit in with another lab.
> >So firstly, has anyone had any experience cutting Mineralised (not
> >decalcified!) bone on a heavy duty rotary microtome or should we be using a
> >sledge?
> >Secondly, what resins do you use and are they available in easy to use kit
> >forms?
> >
> >Thanks for any help in advance
> >
> >Andrew Kennedy
> >
> >Senior Science Officer in charge - Histopathology.
> >Department of Anatomical Pathology
> >Concord Repatriation General Hospital
> >Hospital Road
> >Concord, NSW, Australia
> >2139
> >Phone: +612 9767 6115
> >Fax: +612 9767 8427
> >
> >"Noli illegitimi carborundum"
> >
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> >
> >
> >
> Gayle Callis
> Histopathology Supervisor
> Veterinary Molecular Biology - Marsh Lab
> Montana State University - Bozeman
> 19th and Lincoln St
> Bozeman MT 59717-3610
> 406 994-6367
> 406 994-4303 (FAX)

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