Hello Jessica,
Hello Natalia,

we have discussed the question of bone staining methods in our lab and we would 
recommend the following stainings, which will work on resin embedded material:

(1) For an overview and differentiation of various types of bone the 
traditional MASSON-GOLDNER or AZAN staining is quite fine, and should be the 
first try because these stains are more or less easy and fast and the staining 
solutions are not too expensive.

(2) MOVAT-Pentachrome in the original and in the modification after VERHOEFF 
also gives quite good differentiations of resin embedded bones and shows many 
details even in different types of bone. We use these staining here in many 
research projects we do for pharma companies that develop bone replacement 
materials.

(3) van KOSSA is also a very good silver impregnation to show mineralized bone, 
AND there is a possible combination with the MOVAT Pentachrom-Staining.

(4) Alizarinred can also be used to show different gradients of mineralisation; 
the staining kit contains solutions with several pH-values.

(5) Picro-Siriusred and other combinations of picric acid and other stains 
(picro fuchsin, picro polychrom, picro indigocarmin, and 
orcein-picroindicocramin can show good differentiations on tissues and 
cartilage and bone structures.
Remark: Sometimes its a bit difficult to establish picric acid based stainings 
on resin embedded tissues, if the resin has not been completely removed (which 
is possible with technovit 9100 (and some other MMA’s), but not with technovit 
7200).

(6) LEVAI-LACZKO's stain provides a good differentiation between mineralized 
bone matrix, Osteoid and synthetic bone substitutes.

(7) A special „Bone Stain“, published by Villanueva in 2009 
(https://www.tandfonline.com/doi/abs/10.3109/10520297409116928 
<https://www.tandfonline.com/doi/abs/10.3109/10520297409116928>) works best on 
fresh material. We have not yet tested it completely on resin embedded tissues, 
but some of our customers used it successfully.
For this stain, a „staining powder" has to be synthesized from various single 
stains (basic fuchsin, orange G, fastgreen and azure II) and finally mixed into 
a ready to use staining solution; we produce the stain powder and ready to use 
solution here in our lab.

(8) Last but not least HEROVICI’s stain might help beacuse it differentiates 
between new and old collagen structures. Though it was originally developed for 
diagnostics in wound repair, some of our customers use it to differentiate 
between mineralized bone, new bone (Osteoid) and synthetic bone substitutes.

For all staining procedures on resin embedded material its necessary to edge 
the surface with citric acid or hydrogen peroxide, depending on the kind of 
resin you are using. If it is technovit 7200 citric acid should work fine, for 
technovit 9100 you can also try to remove the resin with methoxyethylacetate 
and acetone. Please note, that there can be a significant difference in the 
quality of the stain between Technovit 9100 and Technovit 7200, depending on 
what you want to see. In general, the stain on Technovit 9100 sections will be 
more powerful and brilliant than that on Technovit 7200 sections. 

As mentioned in my previous mail, we have a local dealer in Oklahoma, and if 
you or somebody else is interested in our ready-to-use stainings kits, just let 
me know, I will then make the direct contact.

Kind regards
Michael



> Am 10.07.2018 um 22:15 schrieb Jessica Riggleman via Histonet 
> <histonet@lists.utsouthwestern.edu>:
> 
> Hey Jennifer,
> Thanks for your help. I have two questions:
> 
> 1. What is the point of decalcifying if they are in plastic?
> 2. Did you see the counterstain? Right now only the bone is staining. How 
> long did you leave these in the counterstain for?
> Thanks!
> Jessica
> 
> From: Mac Donald, Jennifer [mailto:jmacdon...@mtsac.edu]
> Sent: Tuesday, July 10, 2018 4:07 PM
> To: Jessica Riggleman <jriggle...@globusmedical.com>; 
> histonet@lists.utsouthwestern.edu
> Subject: Re: [Histonet] New Bone Stain
> 
> I used to do it on plastic sections of in-de-calcified bone biopsies. It’s a 
> silver reaction.
> 
> Get Outlook for iOS
> 
> 
> 
> _____________________________________________________________________
> 
> Jessica Riggleman | Research Associate
> 
> Globus Medical, Inc.
> Valley Forge Business Center
> 2560 General Armistead Avenue | Audubon, PA 19403
> Ph: (610) 930-1800 ext. 2583 | Fax:
> 
> Confidentiality Note:  This email is confidential and intended solely for the 
> use of the individual to whom it is addressed. If you are not the intended 
> recipient, be advised that you have received this email in error and that any 
> use, dissemination, forwarding, printing, or copying of this email is 
> strictly prohibited. If you have received this email in error please contact 
> the sender. Any views or opinions presented are solely those of the author 
> and do not necessarily represent those of Globus Medical, Inc. Although this 
> email and any attachments are believed to be free of any virus or other 
> defects which might affect any computer or IT system into which they are 
> received, no responsibility is accepted by Globus Medical, Inc. for any loss 
> or damage arising in any way from the receipt or use thereof.
> 
> 
> ________________________________________
> From: Jessica Riggleman <jriggle...@globusmedical.com>
> Sent: Tuesday, July 10, 2018 1:01 PM
> To: Mac Donald, Jennifer; histonet@lists.utsouthwestern.edu
> Subject: RE: [Histonet] New Bone Stain
> 
> Thank You,
> Have you ever tried this on thick histology sections? I have a titanium 
> implant in this section, so thin is impossible.
> 
> Thank You,
> Jessica
> 
> ________________________________________
> 
> Jessica Riggleman | Research Associate
> 
> Globus Medical, Inc.
> Valley Forge Business Center
> 2560 General Armistead Avenue | Audubon, PA 19403
> Ph: (610) 930-1800 ext. 2583 | Fax:
> An ISO 13485 Registered Company
> 
> 
> 
> 
> 
> 
> 
> Confidentiality Note:  This email is confidential and intended solely for the 
> use of the individual to whom it is addressed. If you are not the intended 
> recipient, be advised that you have received this email in error and that any 
> use, dissemination, forwarding, printing, or copying of this email is 
> strictly prohibited. If you have received this email in error please contact 
> the sender. Any views or opinions presented are solely those of the author 
> and do not necessarily represent those of Globus Medical, Inc. Although this 
> email and any attachments are believed to be free of any virus or other 
> defects which might affect any computer or IT system into which they are 
> received, no responsibility is accepted by Globus Medical, Inc. for any loss 
> or damage arising in any way from the receipt or use thereof.
> From: Mac Donald, Jennifer [mailto:jmacdon...@mtsac.edu]
> Sent: Monday, July 09, 2018 6:09 PM
> To: Jessica Riggleman <jriggle...@globusmedical.com>; 
> histonet@lists.utsouthwestern.edu
> Subject: Re: [Histonet] New Bone Stain
> 
> von Kossa will stain the mineralized bone black and a basic fuschin counter 
> stain the osteoid will stain deep pink.
> 
> Get Outlook for iOS
> 
> ________________________________________
> From: Jessica Riggleman via Histonet <histonet@lists.utsouthwestern.edu>
> Sent: Monday, July 9, 2018 1:17 PM
> To: histonet@lists.utsouthwestern.edu
> Subject: [Histonet] New Bone Stain
> 
> Hello,
> I am trying to assess new bone vs. old bone in an animal model. Any 
> suggestions on thick section stains? I have heard Van Gieson's?
> 
> Thank You,
> Jessica
> 
> 
> _____________________________________________________________________
> 
> Jessica Riggleman | Research Associate
> 
> Globus Medical, Inc.
> Valley Forge Business Center
> 2560 General Armistead Avenue | Audubon, PA 19403
> Ph: (610) 930-1800 ext. 2583 | Fax:
> 
> Confidentiality Note: This email is confidential and intended solely for the 
> use of the individual to whom it is addressed. If you are not the intended 
> recipient, be advised that you have received this email in error and that any 
> use, dissemination, forwarding, printing, or copying of this email is 
> strictly prohibited. If you have received this email in error please contact 
> the sender. Any views or opinions presented are solely those of the author 
> and do not necessarily represent those of Globus Medical, Inc. Although this 
> email and any attachments are believed to be free of any virus or other 
> defects which might affect any computer or IT system into which they are 
> received, no responsibility is accepted by Globus Medical, Inc. for any loss 
> or damage arising in any way from the receipt or use thereof.
> 
> 
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