How to keep solution on slides
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi all, This might be a very simple question for many of you. We are doing IHC with dewaxed tissue sections on slides. When we incubated the tissue sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% triton) did not stay inside the circle we drew with a diamond scribe or sharpie; it spread over the slide, sometimes the sections dried. I wonder if any of you have an easy and simple way with little effort/cost to prevent solution running off the slides. Any input is highly appreciated. Best, Greg ---------------------------- Gang (Greg) Ning, Ph.D. Director, Microscopy Facility The Huck Institutes of the Life Sciences The Pennsylvania State University N-048 MSC University Park, PA 16802 USA Phone: 814-863-0994 Fax: 914-867-2587 Email: [hidden email] http://www.huck.psu.edu/facilities/microscopy-cytometry-up |
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** One idea, stretch parafilm flat and stick to lab bench or similar. Put as little as 20 ul of solution in dot on parafilm. Put coverslip down on solution. To remove coverslip without ripping tissue/cells, touch pipette tip with buffer at edge of coverslip just slightly lifting and inject buffer slowly (100 ul tip) so coverslip floats. Grab edge with tweezrs and gently pull off to the side. Michael Cammer, Research Scientist, DART Microscopy Laboratory NYU Langone Health, 540 First Avenue, SK2 Microscopy Suite, New York, NY 10016 C: 914-309-3270 [hidden email] http://microscopynotes.com/ https://med.nyu.edu/research/research-resources/scientific-cores-shared-resources/microscopy-laboratory -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of GANG NING Sent: Wednesday, October 25, 2017 10:43 AM To: [hidden email] Subject: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: https://urldefense.proofpoint.com/v2/url?u=http-3A__lists.umn.edu_cgi-2Dbin_wa-3FA0-3Dconfocalmicroscopy&d=DQICaQ&c=j5oPpO0eBH1iio48DtsedbOBGmuw5jHLjgvtN2r4ehE&r=oU_05LztNstAydlbm5L5GDu_vAdjXk3frDLx_CqKkuo&m=xR-N9zWn5NdPFPv2of-T_K5d1r07Cyt0AEDw_l9pQ_0&s=tXQU_VJtk67BP1CpxZQNoQJYxWFjuP1Ok3Ae_t5QiaA&e= Post images on https://urldefense.proofpoint.com/v2/url?u=http-3A__www.imgur.com&d=DQICaQ&c=j5oPpO0eBH1iio48DtsedbOBGmuw5jHLjgvtN2r4ehE&r=oU_05LztNstAydlbm5L5GDu_vAdjXk3frDLx_CqKkuo&m=xR-N9zWn5NdPFPv2of-T_K5d1r07Cyt0AEDw_l9pQ_0&s=gGpb-kQYqb7YjfNf1q8iT-0UoEB1RSTJZHLq34o3h4Y&e= and include the link in your posting. ***** Hi all, This might be a very simple question for many of you. We are doing IHC with dewaxed tissue sections on slides. When we incubated the tissue sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% triton) did not stay inside the circle we drew with a diamond scribe or sharpie; it spread over the slide, sometimes the sections dried. I wonder if any of you have an easy and simple way with little effort/cost to prevent solution running off the slides. Any input is highly appreciated. Best, Greg ---------------------------- Gang (Greg) Ning, Ph.D. Director, Microscopy Facility The Huck Institutes of the Life Sciences The Pennsylvania State University N-048 MSC University Park, PA 16802 USA Phone: 814-863-0994 Fax: 914-867-2587 Email: [hidden email] https://urldefense.proofpoint.com/v2/url?u=http-3A__www.huck.psu.edu_facilities_microscopy-2Dcytometry-2Dup&d=DQICaQ&c=j5oPpO0eBH1iio48DtsedbOBGmuw5jHLjgvtN2r4ehE&r=oU_05LztNstAydlbm5L5GDu_vAdjXk3frDLx_CqKkuo&m=xR-N9zWn5NdPFPv2of-T_K5d1r07Cyt0AEDw_l9pQ_0&s=JFjtSLrqV_tGYQK_pT1MyKF7AJqeR8irdo3P2QIr8ik&e= ------------------------------------------------------------ This email message, including any attachments, is for the sole use of the intended recipient(s) and may contain information that is proprietary, confidential, and exempt from disclosure under applicable law. Any unauthorized review, use, disclosure, or distribution is prohibited. If you have received this email in error please notify the sender by return email and delete the original message. Please note, the recipient should check this email and any attachments for the presence of viruses. The organization accepts no liability for any damage caused by any virus transmitted by this email. ================================= |
 
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Virginia Bain |
Re: How to keep solution on slides
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In reply to this post by Greg Ning
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi Greg, Are you trying to use multiple antibodies of the same host on one slide? If not I would recommend using parafilm as a coverslip and using the diluted antibody on the entire slide. I found that 100 uL was sufficient to stain my sample without evaporating when incubated at 4C in a humid chamber overnight. If you are using multiple antibodies from the same host on one slide I would recommend using as little buffer as completely covers the section. I find that if I use 100 uL in a small space that would be sufficiently filled with 50 uL then the buffer will creep over the PAP pen when it contains triton. Best, Ginny On Wed, Oct 25, 2017 at 10:43 AM, GANG NING <[hidden email]> wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > Post images on http://www.imgur.com and include the link in your posting. > ***** > > Hi all, > > This might be a very simple question for many of you. We are doing IHC > with dewaxed tissue sections on slides. When we incubated the tissue > sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% > triton) did not stay inside the circle we drew with a diamond scribe or > sharpie; it spread over the slide, sometimes the sections dried. I wonder > if any of you have an easy and simple way with little effort/cost to > prevent solution running off the slides. Any input is highly appreciated. > > Best, > > Greg > > ---------------------------- > Gang (Greg) Ning, Ph.D. > Director, Microscopy Facility > The Huck Institutes of the Life Sciences > The Pennsylvania State University > N-048 MSC > University Park, PA 16802 USA > Phone: 814-863-0994 > Fax: 914-867-2587 > Email: [hidden email] > http://www.huck.psu.edu/facilities/microscopy-cytometry-up > -- This message contains information which may be confidential and/or protected by attorney-client privilege. Unless you are the addressee, you may not use, copy or disclose to anyone this message or any information contained in this message. If you have received this message in error, please send me an email and delete this message. Thank you. |
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kspencer007 |
Re: How to keep solution on slides
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In reply to this post by Greg Ning
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Pap Pen https://www.thermofisher.com/order/catalog/product/008899 Kathy The Scripps Research Institute -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of GANG NING Sent: Wednesday, October 25, 2017 7:43 AM To: [hidden email] Subject: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi all, This might be a very simple question for many of you. We are doing IHC with dewaxed tissue sections on slides. When we incubated the tissue sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% triton) did not stay inside the circle we drew with a diamond scribe or sharpie; it spread over the slide, sometimes the sections dried. I wonder if any of you have an easy and simple way with little effort/cost to prevent solution running off the slides. Any input is highly appreciated. Best, Greg ---------------------------- Gang (Greg) Ning, Ph.D. Director, Microscopy Facility The Huck Institutes of the Life Sciences The Pennsylvania State University N-048 MSC University Park, PA 16802 USA Phone: 814-863-0994 Fax: 914-867-2587 Email: [hidden email] http://www.huck.psu.edu/facilities/microscopy-cytometry-up |
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In reply to this post by Greg Ning
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Greg Boiptechs makes a device called Culture Cylinders that might do the job for you. Although they are made for plating live cells they might be useful for your application. They are a borosilicate glass cylinder similar to a cloning ring having an optically flat polished end better than 1/4 wave that hydrostatically seals to another flat optical surface like a slide. It is used when plating cells to position them for observation and improving plating time by restricting the amount of media that the cells have to re-equilibrate after being tripsinized. They are easily removed after processing with a forcep and leave no residue. Culture cylinders are reusable and autoclaveable. Let me know if you want to try one. Free samples have a 6 or 8mm ID. If it works for your application you can get any size you want. Link if interested: http://www.bioptechs.com/product/culture-cylinders/ Dan On Oct 25, 2017, at 10:43 AM, GANG NING wrote: ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi all, This might be a very simple question for many of you. We are doing IHC with dewaxed tissue sections on slides. When we incubated the tissue sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% triton) did not stay inside the circle we drew with a diamond scribe or sharpie; it spread over the slide, sometimes the sections dried. I wonder if any of you have an easy and simple way with little effort/cost to prevent solution running off the slides. Any input is highly appreciated. Best, Greg ---------------------------- Gang (Greg) Ning, Ph.D. Director, Microscopy Facility The Huck Institutes of the Life Sciences The Pennsylvania State University N-048 MSC University Park, PA 16802 USA Phone: 814-863-0994 Fax: 914-867-2587 Email: [hidden email] http://www.huck.psu.edu/facilities/microscopy-cytometry-up Dan Focht Bioptechs Inc. 3560 Beck Road Butler, PA 16002-9259 Office: 724-282-7145 Toll Free: 877-LIVE-CELL (548-3235) [hidden email] www.bioptechs.com |
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cmci_concordia |
Re: How to keep solution on slides
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In reply to this post by mcammer
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** I agree with Virginia and Michael that parafilm is the answer. I used exactly the method Virginia described for years with excellent results. A PAP pen is okay, but in my experience was less effective when using buffers containing detergent, and is *really* expensive. I have also used glass coverslips with similar efficacy, you just have to be more careful when removing them; I used a large bath of buffer and submerged the whole slide so that the coverslip floated off quickly and smoothly without pulling the tissue off the slide. As a disclaimer, this was done on cryosections, not paraffin-embedded tissue. You also cannot recover the antibody solution if you use this coverslip method. Best of luck! Chris -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of GANG NING Sent: Wednesday, October 25, 2017 10:43 AM To: [hidden email] Subject: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: https://urldefense.proofpoint.com/v2/url?u=http-3A__lists.umn.edu_cgi-2Dbin_wa-3FA0-3Dconfocalmicroscopy&d=DQICaQ&c=j5oPpO0eBH1iio48DtsedbOBGmuw5jHLjgvtN2r4ehE&r=oU_05LztNstAydlbm5L5GDu_vAdjXk3frDLx_CqKkuo&m=xR-N9zWn5NdPFPv2of-T_K5d1r07Cyt0AEDw_l9pQ_0&s=tXQU_VJtk67BP1CpxZQNoQJYxWFjuP1Ok3Ae_t5QiaA&e= Post images on https://urldefense.proofpoint.com/v2/url?u=http-3A__www.imgur.com&d=DQICaQ&c=j5oPpO0eBH1iio48DtsedbOBGmuw5jHLjgvtN2r4ehE&r=oU_05LztNstAydlbm5L5GDu_vAdjXk3frDLx_CqKkuo&m=xR-N9zWn5NdPFPv2of-T_K5d1r07Cyt0AEDw_l9pQ_0&s=gGpb-kQYqb7YjfNf1q8iT-0UoEB1RSTJZHLq34o3h4Y&e= and include the link in your posting. ***** Hi all, This might be a very simple question for many of you. We are doing IHC with dewaxed tissue sections on slides. When we incubated the tissue sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% triton) did not stay inside the circle we drew with a diamond scribe or sharpie; it spread over the slide, sometimes the sections dried. I wonder if any of you have an easy and simple way with little effort/cost to prevent solution running off the slides. Any input is highly appreciated. Best, Greg ---------------------------- Gang (Greg) Ning, Ph.D. Director, Microscopy Facility The Huck Institutes of the Life Sciences The Pennsylvania State University N-048 MSC University Park, PA 16802 USA Phone: 814-863-0994 Fax: 914-867-2587 Email: [hidden email] https://urldefense.proofpoint.com/v2/url?u=http-3A__www.huck.psu.edu_facilities_microscopy-2Dcytometry-2Dup&d=DQICaQ&c=j5oPpO0eBH1iio48DtsedbOBGmuw5jHLjgvtN2r4ehE&r=oU_05LztNstAydlbm5L5GDu_vAdjXk3frDLx_CqKkuo&m=xR-N9zWn5NdPFPv2of-T_K5d1r07Cyt0AEDw_l9pQ_0&s=JFjtSLrqV_tGYQK_pT1MyKF7AJqeR8irdo3P2QIr8ik&e= ------------------------------------------------------------ This email message, including any attachments, is for the sole use of the intended recipient(s) and may contain information that is proprietary, confidential, and exempt from disclosure under applicable law. Any unauthorized review, use, disclosure, or distribution is prohibited. If you have received this email in error please notify the sender by return email and delete the original message. Please note, the recipient should check this email and any attachments for the presence of viruses. The organization accepts no liability for any damage caused by any virus transmitted by this email. ================================= |
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Tamara Howard |
Re: How to keep solution on slides
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In reply to this post by Greg Ning
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Greg - PAP pens are great (already suggested), but in a pinch you can also use a wax pencil or vaseline/petroleum jelly on a cotton swab to draw the barrier around your sections. Tamara ................................................... Tamara Howard Dept. of Cell Biology & Physiology University of New Mexico Albuquerque, NM ________________________________ From: Confocal Microscopy List <[hidden email]> on behalf of GANG NING <[hidden email]> Sent: Wednesday, October 25, 2017 8:43 AM To: [hidden email] Subject: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy LISTSERV 16.0 - CONFOCALMICROSCOPY List at LISTS.UMN.EDU<http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy> lists.umn.edu [hidden email]: listserv archives. confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. [http://upload.wikimedia.org/wikipedia/commons/thumb/e/e9/Imgur_logo.svg/150px-Imgur_logo.svg.png]<http://www.imgur.com/> Imgur: The most awesome images on the Internet<http://www.imgur.com/> www.imgur.com Imgur is the best place to share and enjoy the most awesome images on the Internet. Every day, millions of people use Imgur to be entertained and inspired by funny ... ***** Hi all, This might be a very simple question for many of you. We are doing IHC with dewaxed tissue sections on slides. When we incubated the tissue sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% triton) did not stay inside the circle we drew with a diamond scribe or sharpie; it spread over the slide, sometimes the sections dried. I wonder if any of you have an easy and simple way with little effort/cost to prevent solution running off the slides. Any input is highly appreciated. Best, Greg ---------------------------- Gang (Greg) Ning, Ph.D. Director, Microscopy Facility The Huck Institutes of the Life Sciences The Pennsylvania State University N-048 MSC University Park, PA 16802 USA Phone: 814-863-0994 Fax: 914-867-2587 Email: [hidden email] http://www.huck.psu.edu/facilities/microscopy-cytometry-up |
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Tina Carvalho |
Re: How to keep solution on slides
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In reply to this post by Greg Ning
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** I dislike PAP pens, but love slides with frosted or, even better, Teflon circles on them. Here's a link to the frosted type: https://us.vwr.com/store/product/20296068/two-circle-microscope-slides-for-fluorescent-microscopy-springside-scientific and I can't seem to find a link to my favorite Esco fluoro slides with two Teflon (I think) circles In a pinch you can use plastic 3-ring binder paper hole reinforcements! https://www.staples.com/paper+hole+reinforcements/directory_paper+hole+reinforcements Aloha, Tina > Hi all, > > This might be a very simple question for many of you. We are doing IHC with dewaxed tissue sections on slides. When we incubated the tissue sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% triton) did not stay inside the circle we drew with a diamond scribe or sharpie; it spread over the slide, sometimes the sections dried. I wonder if any of you have an easy and simple way with little effort/cost to prevent solution running off the slides. Any input is highly appreciated. > > Best, > > Greg > > ---------------------------- > Gang (Greg) Ning, Ph.D. > Director, Microscopy Facility > The Huck Institutes of the Life Sciences > The Pennsylvania State University > N-048 MSC > University Park, PA 16802 USA > Phone: 814-863-0994 > Fax: 914-867-2587 > Email: [hidden email] > http://www.huck.psu.edu/facilities/microscopy-cytometry-up > **************************************************************************** * Tina (Weatherby) Carvalho * [hidden email] * * Biological Electron Microscope Facility * (808) 956-6251 * * University of Hawaii at Manoa * http://www.pbrc.hawaii.edu/bemf* **************************************************************************** |
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Jacqueline Ross |
Re: How to keep solution on slides
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In reply to this post by Tamara Howard
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi everyone, I would also recommend using a PAP pen but be careful about how much you put on if you want to use high NA immersion objective lenses. I have had occasions where people have used too much and then when they mount their sections (with aqueous mount), the mountant sits in a bubble, which causes an issue if the lens has a short working distance. Sometimes, I will advise them to remove the wax carefully (and safely!) by using xylene. Cheers, Jacqui -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Tamara Howard Sent: Thursday, 26 October 2017 7:09 a.m. To: [hidden email] Subject: Re: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Greg - PAP pens are great (already suggested), but in a pinch you can also use a wax pencil or vaseline/petroleum jelly on a cotton swab to draw the barrier around your sections. Tamara ................................................... Tamara Howard Dept. of Cell Biology & Physiology University of New Mexico Albuquerque, NM ________________________________ From: Confocal Microscopy List <[hidden email]> on behalf of GANG NING <[hidden email]> Sent: Wednesday, October 25, 2017 8:43 AM To: [hidden email] Subject: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy LISTSERV 16.0 - CONFOCALMICROSCOPY List at LISTS.UMN.EDU<http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy> lists.umn.edu [hidden email]: listserv archives. confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. [http://upload.wikimedia.org/wikipedia/commons/thumb/e/e9/Imgur_logo.svg/150px-Imgur_logo.svg.png]<http://www.imgur.com/> Imgur: The most awesome images on the Internet<http://www.imgur.com/> www.imgur.com Imgur is the best place to share and enjoy the most awesome images on the Internet. Every day, millions of people use Imgur to be entertained and inspired by funny ... ***** Hi all, This might be a very simple question for many of you. We are doing IHC with dewaxed tissue sections on slides. When we incubated the tissue sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% triton) did not stay inside the circle we drew with a diamond scribe or sharpie; it spread over the slide, sometimes the sections dried. I wonder if any of you have an easy and simple way with little effort/cost to prevent solution running off the slides. Any input is highly appreciated. Best, Greg ---------------------------- Gang (Greg) Ning, Ph.D. Director, Microscopy Facility The Huck Institutes of the Life Sciences The Pennsylvania State University N-048 MSC University Park, PA 16802 USA Phone: 814-863-0994 Fax: 914-867-2587 Email: [hidden email] http://www.huck.psu.edu/facilities/microscopy-cytometry-up |
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Kesavan Subburam |
Re: How to keep solution on slides
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In reply to this post by Greg Ning
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi, We use O-rings and seal it with paraffin wax to fix it on cover slip to keep solutions in the slides. You can also seal the open side with a second cover slip to minimise evaporation. Best, Kesavan Subburam On 25-Oct-2017 9:30 PM, "GANG NING" <[hidden email]> wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > Post images on http://www.imgur.com and include the link in your posting. > ***** > > Hi all, > > This might be a very simple question for many of you. We are doing IHC > with dewaxed tissue sections on slides. When we incubated the tissue > sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% > triton) did not stay inside the circle we drew with a diamond scribe or > sharpie; it spread over the slide, sometimes the sections dried. I wonder > if any of you have an easy and simple way with little effort/cost to > prevent solution running off the slides. Any input is highly appreciated. > > Best, > > Greg > > ---------------------------- > Gang (Greg) Ning, Ph.D. > Director, Microscopy Facility > The Huck Institutes of the Life Sciences > The Pennsylvania State University > N-048 MSC > University Park, PA 16802 USA > Phone: 814-863-0994 > Fax: 914-867-2587 > Email: [hidden email] > http://www.huck.psu.edu/facilities/microscopy-cytometry-up > |
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Reece, Jeff (NIH/NIDDK) [E] |
Re: How to keep solution on slides
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*****
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** More choices, on my list to try: https://www.thermofisher.com/us/en/home/references/molecular-probes-the-handbook/tables/press-to-seal-gaskets-and-secure-seal-spacers.html Cheers ________________________________ From: Kesavan Subburam <[hidden email]> Sent: Oct 26, 2017 5:03 AM To: [hidden email] Subject: Re: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi, We use O-rings and seal it with paraffin wax to fix it on cover slip to keep solutions in the slides. You can also seal the open side with a second cover slip to minimise evaporation. Best, Kesavan Subburam On 25-Oct-2017 9:30 PM, "GANG NING" <[hidden email]> wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > Post images on http://www.imgur.com and include the link in your posting. > ***** > > Hi all, > > This might be a very simple question for many of you. We are doing IHC > with dewaxed tissue sections on slides. When we incubated the tissue > sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% > triton) did not stay inside the circle we drew with a diamond scribe or > sharpie; it spread over the slide, sometimes the sections dried. I wonder > if any of you have an easy and simple way with little effort/cost to > prevent solution running off the slides. Any input is highly appreciated. > > Best, > > Greg > > ---------------------------- > Gang (Greg) Ning, Ph.D. > Director, Microscopy Facility > The Huck Institutes of the Life Sciences > The Pennsylvania State University > N-048 MSC > University Park, PA 16802 USA > Phone: 814-863-0994 > Fax: 914-867-2587 > Email: [hidden email] > http://www.huck.psu.edu/facilities/microscopy-cytometry-up > |
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Michael Delannoy |
Re: How to keep solution on slides
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In reply to this post by Jacqueline Ross
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Greg, When I have done Aby incubations on coverslips, a 50 ul drop on a 22 x 22 mm coverslips Would be enough to suspend the c/s from the slide with solution trapped in between. We would then place this in a humidity chamber (petri dish affixed with moist towels secured with nail polish on the inside cover and on the bottom). The slides would be placed on two wooden applicators (so it is not touching the moist towel) and we would incubate without solution drying out. Too rinse simply flood the slide with PBS, then remove the floating c/s horizontally. Sincerely, Michael Delannoy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Jacqui Ross Sent: Wednesday, October 25, 2017 4:51 PM To: [hidden email] Subject: Re: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi everyone, I would also recommend using a PAP pen but be careful about how much you put on if you want to use high NA immersion objective lenses. I have had occasions where people have used too much and then when they mount their sections (with aqueous mount), the mountant sits in a bubble, which causes an issue if the lens has a short working distance. Sometimes, I will advise them to remove the wax carefully (and safely!) by using xylene. Cheers, Jacqui -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Tamara Howard Sent: Thursday, 26 October 2017 7:09 a.m. To: [hidden email] Subject: Re: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Greg - PAP pens are great (already suggested), but in a pinch you can also use a wax pencil or vaseline/petroleum jelly on a cotton swab to draw the barrier around your sections. Tamara ................................................... Tamara Howard Dept. of Cell Biology & Physiology University of New Mexico Albuquerque, NM ________________________________ From: Confocal Microscopy List <[hidden email]> on behalf of GANG NING <[hidden email]> Sent: Wednesday, October 25, 2017 8:43 AM To: [hidden email] Subject: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy LISTSERV 16.0 - CONFOCALMICROSCOPY List at LISTS.UMN.EDU<http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy> lists.umn.edu [hidden email]: listserv archives. confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. [http://upload.wikimedia.org/wikipedia/commons/thumb/e/e9/Imgur_logo.svg/150 px-Imgur_logo.svg.png]<http://www.imgur.com/> Imgur: The most awesome images on the Internet<http://www.imgur.com/> www.imgur.com Imgur is the best place to share and enjoy the most awesome images on the Internet. Every day, millions of people use Imgur to be entertained and inspired by funny ... ***** Hi all, This might be a very simple question for many of you. We are doing IHC with dewaxed tissue sections on slides. When we incubated the tissue sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% triton) did not stay inside the circle we drew with a diamond scribe or sharpie; it spread over the slide, sometimes the sections dried. I wonder if any of you have an easy and simple way with little effort/cost to prevent solution running off the slides. Any input is highly appreciated. Best, Greg ---------------------------- Gang (Greg) Ning, Ph.D. Director, Microscopy Facility The Huck Institutes of the Life Sciences The Pennsylvania State University N-048 MSC University Park, PA 16802 USA Phone: 814-863-0994 Fax: 914-867-2587 Email: [hidden email] http://www.huck.psu.edu/facilities/microscopy-cytometry-up |
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In reply to this post by Greg Ning
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi Greg, I stick multiple layers of electrical tape on the back of a glass Petri dish, then cut out a square at the center with a sharp scalpel to prepare a mask and transfer the mask by sticking it firmly onto glass microscope slide. Incubate samples with antibodies in the cutout well, then remove the mask before mounting with a coverslip. I uploaded an image to Imgur for instructions (I hope the link will work) https://imgur.com/a/Ou5dj Best, Ferhan ---------------- Ferhan Ayaydin, Ph.D. Cellular Imaging Lab. Biological Research Centre Hungarian Academy of Sciences Szeged, Hungary On Wed, Oct 25, 2017 at 4:43 PM, GANG NING <[hidden email]> wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > Post images on http://www.imgur.com and include the link in your posting. > ***** > > Hi all, > > This might be a very simple question for many of you. We are doing IHC > with dewaxed tissue sections on slides. When we incubated the tissue > sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% > triton) did not stay inside the circle we drew with a diamond scribe or > sharpie; it spread over the slide, sometimes the sections dried. I wonder > if any of you have an easy and simple way with little effort/cost to > prevent solution running off the slides. Any input is highly appreciated. > > Best, > > Greg > > ---------------------------- > Gang (Greg) Ning, Ph.D. > Director, Microscopy Facility > The Huck Institutes of the Life Sciences > The Pennsylvania State University > N-048 MSC > University Park, PA 16802 USA > Phone: 814-863-0994 > Fax: 914-867-2587 > Email: [hidden email] > http://www.huck.psu.edu/facilities/microscopy-cytometry-up > |
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Luis Enrique Villegas Hernandez |
Re: How to keep solution on slides
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In reply to this post by Greg Ning
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi Greg, You can also try placing a PDMS frame around the sample, so that it creates a micro chamber while staining. When you are finished, the PDMS can be gently removed with a tweezer. Regards, Luis On Oct 26, 2017 16:38, Michael Delannoy <[hidden email]> wrote: ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Greg, When I have done Aby incubations on coverslips, a 50 ul drop on a 22 x 22 mm coverslips Would be enough to suspend the c/s from the slide with solution trapped in between. We would then place this in a humidity chamber (petri dish affixed with moist towels secured with nail polish on the inside cover and on the bottom). The slides would be placed on two wooden applicators (so it is not touching the moist towel) and we would incubate without solution drying out. Too rinse simply flood the slide with PBS, then remove the floating c/s horizontally. Sincerely, Michael Delannoy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Jacqui Ross Sent: Wednesday, October 25, 2017 4:51 PM To: [hidden email] Subject: Re: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Hi everyone, I would also recommend using a PAP pen but be careful about how much you put on if you want to use high NA immersion objective lenses. I have had occasions where people have used too much and then when they mount their sections (with aqueous mount), the mountant sits in a bubble, which causes an issue if the lens has a short working distance. Sometimes, I will advise them to remove the wax carefully (and safely!) by using xylene. Cheers, Jacqui -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Tamara Howard Sent: Thursday, 26 October 2017 7:09 a.m. To: [hidden email] Subject: Re: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Greg - PAP pens are great (already suggested), but in a pinch you can also use a wax pencil or vaseline/petroleum jelly on a cotton swab to draw the barrier around your sections. Tamara ................................................... Tamara Howard Dept. of Cell Biology & Physiology University of New Mexico Albuquerque, NM ________________________________ From: Confocal Microscopy List <[hidden email]> on behalf of GANG NING <[hidden email]> Sent: Wednesday, October 25, 2017 8:43 AM To: [hidden email] Subject: How to keep solution on slides ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy LISTSERV 16.0 - CONFOCALMICROSCOPY List at LISTS.UMN.EDU<http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy> lists.umn.edu [hidden email]: listserv archives. confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. [http://upload.wikimedia.org/wikipedia/commons/thumb/e/e9/Imgur_logo.svg/150 px-Imgur_logo.svg.png]<http://www.imgur.com/> Imgur: The most awesome images on the Internet<http://www.imgur.com/> www.imgur.com<http://www.imgur.com> Imgur is the best place to share and enjoy the most awesome images on the Internet. Every day, millions of people use Imgur to be entertained and inspired by funny ... ***** Hi all, This might be a very simple question for many of you. We are doing IHC with dewaxed tissue sections on slides. When we incubated the tissue sections with antibodies, esp o/n at 4C, the antibody solution (with 0.1% triton) did not stay inside the circle we drew with a diamond scribe or sharpie; it spread over the slide, sometimes the sections dried. I wonder if any of you have an easy and simple way with little effort/cost to prevent solution running off the slides. Any input is highly appreciated. Best, Greg ---------------------------- Gang (Greg) Ning, Ph.D. Director, Microscopy Facility The Huck Institutes of the Life Sciences The Pennsylvania State University N-048 MSC University Park, PA 16802 USA Phone: 814-863-0994 Fax: 914-867-2587 Email: [hidden email] http://www.huck.psu.edu/facilities/microscopy-cytometry-up |
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Stanislav Vitha-2 |
Re: How to keep solution on slides
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In reply to this post by Greg Ning
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. ***** Another vote for Parafilm. Make sure you cut the piece of Parafilm narrower than the slide, so that the antibody solution does not run down the sides into the filer paper or whatever the slide is sitting on. Also, do not overlap with the frosted, writing area on the end of the slide, if there is one - the capillary action would wick the solution away from under the Parafilm. Here is a plug for my book chapter -Fig. 2N shows the Parafilm use: Vitha, S. and Osteryoung, K. W. (2011) Immunofluorescence Microscopy for Localization of Arabidopsis Chloroplast Proteins. In Chloroplast Research in Arabidopsis (Jarvis, R. P., ed.). pp. 33-58, Humana Press, New York https://link.springer.com/protocol/10.1007%2F978-1-61779-234-2_3 Stan Vitha Microscopy and Imaging Center Texas A&M University |
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