Thank you, Gayle! This is exactly what I was looking for and we are willing to make this in house. We are trying just for if acid and water, but the buffering salt should be added. I will try de calcifying mouse knees next week with this protocol. Thank you for the reference, I appreciate your help!
Sincerely, Merissa > On Jul 25, 2015, at 10:24 AM, Gayle Callis via Histonet > <histonet@lists.utsouthwestern.edu> wrote: > > Merissa and Tim, > > > > This formic acid decalcifying solution is basically the classic Evans and > Krajian fluid (Sheehan and Hrapchak, Theory and Practice of > Histotechnology, 2nd edition, P.92). Shandon has added other ingredients > for some reason, and has kept those concentrations proprietary. You really > don't need to add a surfactant or PVP emulsifier when making up this > decalcifying agent. Simply use the classic recipe for successful > decalcification. This is also referred to as buffered formic acid and in > some publications an "acidic buffer". It is excellent if IHC is needed and > less damaging, obviously, than a strong mineral HCL acid decalcifiers. > > > > Sodium citrate crystals (a buffering salt) 10 g > > 90% formic acid stock 25 ml > > Distilled water 75 ml > > > > One can calculate the concentration of formic acid i.e. approx. 4.5% since > is it made from 90% formic acid stock. > > > > Don't bother with the surfactants or PVP. > > > > Enjoy an excellent in house formic acid decalcifying solution. I also > suggest you read Sheehan and Hrapchak textbook chapter on bone as a way to > familiarize yourself with decalcifiying solutions that manufacturers now > supply with some modifications. Some manufacturers will refer to these > methods but probably prefer not to do this since they want you to buy their > commercial product that is obviously a time saver with elimination of having > to store stock acid solutions. The classic methods made in house are > excellent if you have time to make them up. Formic acid with sodium > formate is another popular buffered formic acid. I suggest you look for > another source/manufacturer of the your favorite decalcifier in question as > more than one company will make it. Decal Corp, recently sold to Stat Lab, > could also be the source as Shandon isn't the only game in town. Others > are Newcomer Supply, Poly Scientific. Not having to make it up may remain > your preference. > > > > Gayle M. Callis > > HTL/HT/MT(ASCP) > > > > > > > > > > > > > > > > > > Written by Tim and Merissa: > > > > Merissa, > > > > Water 77-80 solvent > > Formic acid 21-23 active ingredient > > Fluorad >1 surfactant - a > wetting agent to make the solution wet the bone more easily > > Sodium citrate >1 emulsifier , buffer > > Polyvinyl pyrrolidone >1 emulsifier > > > > They say less than one percent of the last three, but you really have no > idea whether that is 1%, .1% or .01%. It could be any of those. > > > > But all those surfactants and emulsifiers are meant to keep the solution > viable for long periods on the shelf. When you make it fresh you don't > really need them. > > > > You can either buy a different decalcifier, or make your own. Making your > own with just the water and acid will work just fine. > > > > > > Tim Morken > > Pathology Site Manager, Parnassus > > Supervisor, Electron Microscopy/Neuromuscular Special Studies > > Department of Pathology > > UC San Francisco Medical Center > > > > -----Original Message----- > > From: M.O. via Histonet [mailto: > <http://lists.utsouthwestern.edu/mailman/listinfo/histonet> histonet at > lists.utsouthwestern.edu] > > Sent: Wednesday, July 22, 2015 1:24 PM > > To: <http://lists.utsouthwestern.edu/mailman/listinfo/histonet> histonet at > lists.utsouthwestern.edu > > Subject: [Histonet] understanding reagents in decalcifier; making it > in-house > > > > Hello Histonet > > > > The supplier for our decalcifier, TBD-2 from Shandon, is having issues with > getting the product out and we will not be receiving it for at least another > month. Our samples are piling up and I don't know what I should do, but > maybe I can make the decalcifier in-house. I am wondering if I can make my > own based on the reagents they listed and their percentages and if certain > reagents are not actually necessary. > > > > The samples we typically decalcify are mouse knees (decal time = 2 days), > mouse spines (3 days), human bone slabs about 7mm in thickness (7-12 days). > Fixation is in zinc buffered formalin, then decalcification, then 70% EtOH. > Our choice to use TBD-2 is due to the gentle decalcification for IHC and we > get GREAT results. > > > > Composition of Shandon TBD-2 Decalcifier: > > Component Weight % > > Water 77-80 > > Formic acid 21-23 > > Fluorad >1 > > Sodium citrate >1 > > Polyvinyl pyrrolidone >1 > > > > If you have any input on what reagents I should use and the percentages for > making a decalcifier myself, it would be much appreciated! > > > > Thank you for you help, > > Merissa > > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet