Re: [Histonet] Cytology CSF Cell Pellets made from Histogel
I work in a research lab, but I use histogel all the time to make FFPE blocks of cell culture material. Here is my procedure: 1. you need about 5X10^ cells per pellet 2. spin cells @ 2000 rpm for 5 minutes in a 50 ml conical tube 3. aspirate and resuspend in 15ml NBF and fix overnight @ RT 4. spin @ 2000 rpm for 5 minutes and aspirate the NBF 5. microwave the histogel (thermo #HG-4000-012) for 15-20 seconds (loosen cap before microwaving) 6. histogel should now be at liquid for and about 60C 7. resuspend cell pellet in about 350ul of histogel 8. place cell pellet in freezer or on ice for about 20 minutes to solidify 9. using spatula, remove histogel pellet and place into cassette 10. post-fix in NBF for 2 hours (this step is very important); histogel will dissolve in processor if not fixed 11. process as usual Good luck! Kim Kim Merriam, MA, HT(ASCP)QIHC Cambridge, MA From: Turner, Leandra lturn...@seton.org To: histonet@lists.utsouthwestern.edu Sent: Tuesday, February 21, 2012 12:26 PM Subject: [Histonet] Cytology CSF Cell Pellets made from Histogel Hello Everyone, I am trying to find out a few of things about making cell pellets on cerebrospinal fluids. I would like to know: 1. If anyone has ever made cell pellets from CSF's using Histogel and has any tips or procedures they could share? 2. How to process the CSF pellets made with Histogel, do we need a routine or stat process? (we use a Sakura Tissue Tek) 3. Can you do IHC staining on the pellets? Thank you for any and all help in advance. Leandra Turner, HT (ASPC)CM ___ 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] Cytology CSF Cell Pellets made from Histogel
I forgot to mention, we prepare these specifically for IHC; so yes, you can do IHC on them. Kim Kim Merriam, MA, HT(ASCP)QIHC Cambridge, MA From: Kim Merriam kmerriam2...@yahoo.com To: Turner, Leandra lturn...@seton.org; histonet@lists.utsouthwestern.edu histonet@lists.utsouthwestern.edu Sent: Wednesday, February 22, 2012 8:21 AM Subject: Re: [Histonet] Cytology CSF Cell Pellets made from Histogel I work in a research lab, but I use histogel all the time to make FFPE blocks of cell culture material. Here is my procedure: 1. you need about 5X10^ cells per pellet 2. spin cells @ 2000 rpm for 5 minutes in a 50 ml conical tube 3. aspirate and resuspend in 15ml NBF and fix overnight @ RT 4. spin @ 2000 rpm for 5 minutes and aspirate the NBF 5. microwave the histogel (thermo #HG-4000-012) for 15-20 seconds (loosen cap before microwaving) 6. histogel should now be at liquid for and about 60C 7. resuspend cell pellet in about 350ul of histogel 8. place cell pellet in freezer or on ice for about 20 minutes to solidify 9. using spatula, remove histogel pellet and place into cassette 10. post-fix in NBF for 2 hours (this step is very important); histogel will dissolve in processor if not fixed 11. process as usual Good luck! Kim Kim Merriam, MA, HT(ASCP)QIHC Cambridge, MA From: Turner, Leandra lturn...@seton.org To: histonet@lists.utsouthwestern.edu Sent: Tuesday, February 21, 2012 12:26 PM Subject: [Histonet] Cytology CSF Cell Pellets made from Histogel Hello Everyone, I am trying to find out a few of things about making cell pellets on cerebrospinal fluids. I would like to know: 1. If anyone has ever made cell pellets from CSF's using Histogel and has any tips or procedures they could share? 2. How to process the CSF pellets made with Histogel, do we need a routine or stat process? (we use a Sakura Tissue Tek) 3. Can you do IHC staining on the pellets? Thank you for any and all help in advance. Leandra Turner, HT (ASPC)CM ___ 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] Cytology CSF Cell Pellets made from Histogel
Hello Everyone, I am trying to find out a few of things about making cell pellets on cerebrospinal fluids. I would like to know: 1. If anyone has ever made cell pellets from CSF's using Histogel and has any tips or procedures they could share? 2. How to process the CSF pellets made with Histogel, do we need a routine or stat process? (we use a Sakura Tissue Tek) 3. Can you do IHC staining on the pellets? Thank you for any and all help in advance. Leandra Turner, HT (ASPC)CM ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Cytology CSF Cell Pellets made from Histogel
Hi, I haven't done a cellblock on CSF with Histogel, but I have has success with some fairly scanty cell culture specimens. A short processing cycle would be best. Try to make sure you have removed as much supernatant as possible to keep the gel from shriveling during processing. The IHC will be fine as long as the cells are suitably fixed ahead of time. Here is a link to the packaging insert for Histogel which describes preparation of cell blocks. https://www.thermo.com/eThermo/CMA/PDFs/Various/File_9759.pdf And here's a video of people using it for cytology specimens. http://tinyurl.com/7jb522e Amos On Tue, Feb 21, 2012 at 1:01 PM, histonet-requ...@lists.utsouthwestern.eduwrote: Message: 15 Date: Tue, 21 Feb 2012 11:26:10 -0600 From: Turner, Leandra lturn...@seton.org Subject: [Histonet] Cytology CSF Cell Pellets made from Histogel To: histonet@lists.utsouthwestern.edu Message-ID: 3d79f47dc92b204f9e5d35c885dfc5cb010ab...@ausex2vs1.seton.org Content-Type: text/plain; charset=us-ascii Hello Everyone, I am trying to find out a few of things about making cell pellets on cerebrospinal fluids. I would like to know: 1. If anyone has ever made cell pellets from CSF's using Histogel and has any tips or procedures they could share? 2. How to process the CSF pellets made with Histogel, do we need a routine or stat process? (we use a Sakura Tissue Tek) 3. Can you do IHC staining on the pellets? Thank you for any and all help in advance. Leandra Turner, HT (ASPC)CM ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
