Re: [Histonet] Sudanblack B on FFPET
Many thanks to all for your helpful hints. Kind regards Gudrun Lang -Ursprüngliche Nachricht- Von: Tony Henwood via Histonet [mailto:histonet@lists.utsouthwestern.edu] Gesendet: Dienstag, 28. März 2023 21:13 An: Bob Richmond; Histonet@lists.utsouthwestern.edu Betreff: Re: [Histonet] Sudanblack B on FFPET I would also let the saturated solution stand for a few days. Like Oil Red O, it probably needs time to mature. I would also use a frozen section of skin as a control. Regards, Tony Henwood Sydney, Australia From: Bob Richmond via Histonet<mailto:histonet@lists.utsouthwestern.edu> Sent: Wednesday, 29 March 2023 4:51 AM To: Histonet@lists.utsouthwestern.edu<mailto:histonet@lists.utsouthwestern.edu> Subject: Re: [Histonet] Sudanblack B on FFPET > > Gudrun Lang in Austria asks: > >>Has anyone experience with Sudanblack B on paraffin slides for staining [lipofuscin]? A doctor wants the demonstration of the lipoid content of foamy cells or granulocytes in lung. I've found protocols that have incubation-times from 10 minutes to over-night. - I've made a saturated Sudan black B-solution in 70% ethanol and tried it with10 min on liver without real success.<< The main thing you need to do is demonstrate that it isn't hemosiderin with an iron stain (Perls prussian blue reaction), and perhaps also that it isn't acid-fast. Lipofuscin can be identified an H & E staining, except for these considerations. Bob Richmond Maryville, Tennessee ___ 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 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Sudanblack B on FFPET
It's true that Sudan black B won't stain ordinary lipids (fat, phospholipids etc) that are absent from paraffin sections. The lipid in lipofuscin is bound to protein strongly enough to resist extraction during passage through all the solvents used in preparing paraffin sections. Churukian's adaptation of the Lillie & Ashburn method (link to a free download methods book in my recent Histonet email) is OK. It is one of the Biological Stain Commission's tests for certifying Sudan black B. John Kiernan London, Ontario = = = From: AJ Cabral via Histonet Sent: March 28, 2023 4:12 PM To: Tony Henwood Cc: Histonet@lists.utsouthwestern.edu ; Bob Richmond Subject: Re: [Histonet] Sudanblack B on FFPET Sudan Black staining won’t work on FFPET. The alcohols and xylenes used in the tissue processing dissolves the lipids in the tissue. However, you can used formalin fixed tissue as an alternative if no frozen section is available. Rinse the tissue in distilled water for several minutes, pat dry, freeze the tissue on OCT, cut frozen sections and stain in Sudan black. Have you considered looking into acid phosphatase staining for lipofucshin? It is non specific but it can be demonstrated in muscle biopsy. Cheers, Joanna On Tue, Mar 28, 2023 at 12:22 PM Tony Henwood via Histonet < histonet@lists.utsouthwestern.edu> wrote: > I would also let the saturated solution stand for a few days. Like Oil Red > O, it probably needs time to “mature”. I would also use a frozen section of > skin as a control. > > Regards, > > Tony Henwood > Sydney, Australia > > From: Bob Richmond via Histonet<mailto:histonet@lists.utsouthwestern.edu> > Sent: Wednesday, 29 March 2023 4:51 AM > To: Histonet@lists.utsouthwestern.edu histonet@lists.utsouthwestern.edu> > Subject: Re: [Histonet] Sudanblack B on FFPET > > > > > Gudrun Lang in Austria asks: > > > > >>Has anyone experience with Sudanblack B on paraffin slides for staining > [lipofuscin]? A doctor wants the demonstration of the lipoid content of > foamy cells or granulocytes in lung. I've found protocols that have > incubation-times from 10 minutes to over-night. - I've made a saturated > Sudan black B-solution in 70% ethanol and tried it with10 min on liver > without real success.<< > > The main thing you need to do is demonstrate that it isn't hemosiderin with > an iron stain (Perls prussian blue reaction), and perhaps also that it > isn't acid-fast. Lipofuscin can be identified an H & E staining, except for > these considerations. > > Bob Richmond > Maryville, Tennessee > ___ > 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 > ___ 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
Re: [Histonet] Sudanblack B on FFPET
Sudan Black staining won’t work on FFPET. The alcohols and xylenes used in the tissue processing dissolves the lipids in the tissue. However, you can used formalin fixed tissue as an alternative if no frozen section is available. Rinse the tissue in distilled water for several minutes, pat dry, freeze the tissue on OCT, cut frozen sections and stain in Sudan black. Have you considered looking into acid phosphatase staining for lipofucshin? It is non specific but it can be demonstrated in muscle biopsy. Cheers, Joanna On Tue, Mar 28, 2023 at 12:22 PM Tony Henwood via Histonet < histonet@lists.utsouthwestern.edu> wrote: > I would also let the saturated solution stand for a few days. Like Oil Red > O, it probably needs time to “mature”. I would also use a frozen section of > skin as a control. > > Regards, > > Tony Henwood > Sydney, Australia > > From: Bob Richmond via Histonet<mailto:histonet@lists.utsouthwestern.edu> > Sent: Wednesday, 29 March 2023 4:51 AM > To: Histonet@lists.utsouthwestern.edu histonet@lists.utsouthwestern.edu> > Subject: Re: [Histonet] Sudanblack B on FFPET > > > > > Gudrun Lang in Austria asks: > > > > >>Has anyone experience with Sudanblack B on paraffin slides for staining > [lipofuscin]? A doctor wants the demonstration of the lipoid content of > foamy cells or granulocytes in lung. I've found protocols that have > incubation-times from 10 minutes to over-night. - I've made a saturated > Sudan black B-solution in 70% ethanol and tried it with10 min on liver > without real success.<< > > The main thing you need to do is demonstrate that it isn't hemosiderin with > an iron stain (Perls prussian blue reaction), and perhaps also that it > isn't acid-fast. Lipofuscin can be identified an H & E staining, except for > these considerations. > > Bob Richmond > Maryville, Tennessee > ___ > 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 > ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Sudanblack B on FFPET
I would also let the saturated solution stand for a few days. Like Oil Red O, it probably needs time to “mature”. I would also use a frozen section of skin as a control. Regards, Tony Henwood Sydney, Australia From: Bob Richmond via Histonet<mailto:histonet@lists.utsouthwestern.edu> Sent: Wednesday, 29 March 2023 4:51 AM To: Histonet@lists.utsouthwestern.edu<mailto:histonet@lists.utsouthwestern.edu> Subject: Re: [Histonet] Sudanblack B on FFPET > > Gudrun Lang in Austria asks: > >>Has anyone experience with Sudanblack B on paraffin slides for staining [lipofuscin]? A doctor wants the demonstration of the lipoid content of foamy cells or granulocytes in lung. I've found protocols that have incubation-times from 10 minutes to over-night. - I've made a saturated Sudan black B-solution in 70% ethanol and tried it with10 min on liver without real success.<< The main thing you need to do is demonstrate that it isn't hemosiderin with an iron stain (Perls prussian blue reaction), and perhaps also that it isn't acid-fast. Lipofuscin can be identified an H & E staining, except for these considerations. Bob Richmond Maryville, Tennessee ___ 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
Re: [Histonet] Sudanblack B on FFPET
> > Gudrun Lang in Austria asks: > >>Has anyone experience with Sudanblack B on paraffin slides for staining [lipofuscin]? A doctor wants the demonstration of the lipoid content of foamy cells or granulocytes in lung. I've found protocols that have incubation-times from 10 minutes to over-night. - I've made a saturated Sudan black B-solution in 70% ethanol and tried it with10 min on liver without real success.<< The main thing you need to do is demonstrate that it isn't hemosiderin with an iron stain (Perls prussian blue reaction), and perhaps also that it isn't acid-fast. Lipofuscin can be identified an H & E staining, except for these considerations. Bob Richmond Maryville, Tennessee ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Sudanblack B on FFPET
You probably would get better results with a supersaturated solution of the dye in 60% isopropanol with added dextrin. For details see https://www.urmc.rochester.edu/urmc-labs/pathology/StainsManual/. Click on Table of Contents and follow the links to fined the method. Be sure to use a batch of Sudan black B certified by the Biological Stain Commission. John Kiernan London, Canada = = = From: Gudrun Lang via Histonet Sent: March 28, 2023 10:30 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Sudanblack B on FFPET Hallo! Has anyone experience with Sudanblack B on paraffin slides for staining lipofuszin? A doctor wants the demonstration of the lipoid content of foamy cells or granulocytes in lung. I've found protocols that have incubation-times from 10 minutes to over-night. I've made a saturated Sudanblack B-solution in 70% ethanol and tried it with 10 min on liver without real success. I would appreciate any input and help. Thanks in advance Gudrun Lang Austria ___ 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
[Histonet] Sudanblack B on FFPET
Hallo! Has anyone experience with Sudanblack B on paraffin slides for staining lipofuszin? A doctor wants the demonstration of the lipoid content of foamy cells or granulocytes in lung. I've found protocols that have incubation-times from 10 minutes to over-night. I've made a saturated Sudanblack B-solution in 70% ethanol and tried it with 10 min on liver without real success. I would appreciate any input and help. Thanks in advance Gudrun Lang Austria ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
