Re: [Histonet] Tape Transfer Methods For Cryosectioned Brains
Hi Heather, I can see why you’re having trouble. 30 micron sections are inherently unstable. Like paraffin, the thicker a section is the more difficult it is to cut. Plus since your practice samples are old they tend to be more brittle. Try cutting at 5 microns and see what happens. Remember to let the samples warm up in the cryostat if they have been stored at -70 or -80 degrees. If they are too cold they are brittle too. I have not tried the tape method. We just place the sections directly on the slide. Paula Sent from my iPhone > On Apr 15, 2021, at 11:24 AM, Heather Deziel wrote: > > > Hi Paula, > > I am cutting at -24, but have tried going as warm as -18. I am currently > learning with 30uM sections with the ultimate goal of moving towards 5 or > 10uM. Our lab standard has been collecting into millonigs solution and doing > free floating IHC. We generally have no issue with this technique, but do > lose some of the peripheral damaged tissue near the infarct in our stroke > brains. We're trying to work up painting the samples directly onto slides > and skipping free floating staining to get a better end product. > > My current samples are very old, they were collected into 4%PFA in 2017 and > cryoprotected by freezing in OCT cryomatrix in a little dish floating on LN2 > in 2019. They've been stored at -80 since then. Usually we process the > brains within a few weeks of collecting them so this particular tissue isn't > our ideal situation. We're using old tissue to practice technique, so the > current samples aren't going to be used for any actual analysis. When we > heard about the tape method of collecting we were very curious as the the > opinion other labs have about it. Have you tried it? > > Thanks for the answer! > > Heather > > Heather Deziel, MSc. > Laboratory Technician, CNS|CRO > 550 University Ave, Charlottetown, PE C1A 4P3 > > (a subsidiary of Neurodyn Life Sciences Inc.) > > > >> On 2021-04-15 10:29, Patpxs wrote: >> >> Good Morning Heather, >> >> I have some questions about how you cut frozen brains. >> >> What temperature are you cutting at? >> How thick are your sections? >> How are your samples frozen? Flash freezing, slow freezing, iso-pentane in >> LN2? >> >> Your answers may provide clues to help you get better cryosections. >> >> Paula >> >> Sent from my iPhone >> >>> On Apr 15, 2021, at 5:39 AM, Heather Deziel via Histonet >>> wrote: >>> >>> Hello Histonet, >>> >>> I'm looking into working up a tape transfer method of collecting >>> cryosections of brain while preserving infarct to be used in IHC. I >>> find that when I try and section heavily damaged regions of the brain >>> the tissue tears and and I lose it. Has anyone got any recommendations >>> about the the Section-lab transfer tape (Kawamoto method), using the >>> circuit plating tape recommended here >>> (https://www.future-science.com/doi/full/10.2144/btn-2018-0021) or the >>> cryojane system from Leica? >>> >>> Thank you, >>> >>> Heather >>> >>> Heather Deziel, MSc. >>> >>> Laboratory Technician, CNS|CRO 550 University Ave, Charlottetown, PE C1A >>> 4P3 >>> >>> (a subsidiary of Neurodyn Life Sciences Inc.) >>> ___ >>> 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] Filed slides sticking together
We used to use a drying oven set at 37 degrees C. We stopped using it because some people thought it smelled, plus our slide volume increased to the point where all the flats couldn’t fit. Now we dry them for two days, minimum, before filing. I prefer to let them dry for at least 4 days. Paula Sent from my iPhone > On Apr 16, 2021, at 5:26 PM, John Garratt via Histonet > wrote: > > Use tape if you want to have a lean mean operation. Instant filing, no > sticking, no ovens. > > John > >> On Fri, Apr 16, 2021 at 2:55 PM, Jay Lundgren via Histonet >> wrote: >> >> I've seen labs that do this, like 30C for 24 hours, but if you use xylene, >> make sure to put the oven under a hood. >> >> It's a lot easier just to leave them out, in a well ventilated place, for >> 48hours. Buy more slide folders/trays. >> >> I worked one place where the slides were held for 24 hours before being >> given to the paths, because they were concerned about being exposed to >> xylene fumes. But that lab/hospital system had no competition within 100 >> miles and didn't care about turn around times. >> ___ >> 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] Filed slides sticking together
Use tape if you want to have a lean mean operation. Instant filing, no sticking, no ovens. John On Fri, Apr 16, 2021 at 2:55 PM, Jay Lundgren via Histonet wrote: > I've seen labs that do this, like 30C for 24 hours, but if you use xylene, > make sure to put the oven under a hood. > > It's a lot easier just to leave them out, in a well ventilated place, for > 48hours. Buy more slide folders/trays. > > I worked one place where the slides were held for 24 hours before being > given to the paths, because they were concerned about being exposed to > xylene fumes. But that lab/hospital system had no competition within 100 > miles and didn't care about turn around times. > ___ > 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] Filed slides sticking together
I've seen labs that do this, like 30C for 24 hours, but if you use xylene, make sure to put the oven under a hood. It's a lot easier just to leave them out, in a well ventilated place, for 48hours. Buy more slide folders/trays. I worked one place where the slides were held for 24 hours before being given to the paths, because they were concerned about being exposed to xylene fumes. But that lab/hospital system had no competition within 100 miles and didn't care about turn around times. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Filed slides sticking together
We leave ours in slide trays for at least 2 days prior to filing. We are a smaller derm lab so that’s probably not feasible for larger labs. Sent from my iPhone > On Apr 16, 2021, at 12:05 PM, Thomas Podawiltz via Histonet > wrote: > > I never have. Usually let them sit for two days before we filed them. > > > Sent from Yahoo Mail for iPhone > > > On Friday, April 16, 2021, 10:05 AM, Martha Ward-Pathology via Histonet > wrote: > > I am posting this question for our Histology manager: > > Does anyone dry coverslipped slides in an oven before filing and if so how > long and at what temperature. We are having issues with filed slides > sticking together. > > > Thanks in advance for your help with her question. > > Martha Ward, MT ASCP (QIHC) > Manager, Molecular Diagnostics Lab > Wake Forest Baptist Medical Center > Winston-Salem, NC 27157 > > ___ > 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] Filed slides sticking together
I never have. Usually let them sit for two days before we filed them. Sent from Yahoo Mail for iPhone On Friday, April 16, 2021, 10:05 AM, Martha Ward-Pathology via Histonet wrote: I am posting this question for our Histology manager: Does anyone dry coverslipped slides in an oven before filing and if so how long and at what temperature. We are having issues with filed slides sticking together. Thanks in advance for your help with her question. Martha Ward, MT ASCP (QIHC) Manager, Molecular Diagnostics Lab Wake Forest Baptist Medical Center Winston-Salem, NC 27157 ___ 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] Filed slides sticking together
I am posting this question for our Histology manager: Does anyone dry coverslipped slides in an oven before filing and if so how long and at what temperature. We are having issues with filed slides sticking together. Thanks in advance for your help with her question. Martha Ward, MT ASCP (QIHC) Manager, Molecular Diagnostics Lab Wake Forest Baptist Medical Center Winston-Salem, NC 27157 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
