Re: [Histonet] Losing sections
Terri this sounds entirely logical! I will try running a blank rack to warm the drying station up before running a single deeper slide. My pathologist and I will both be very happy if it makes a difference! Thank you! -Original Message- From: Terri Braud Sent: Friday, June 16, 2023 12:56 PM To: histonet@lists.utsouthwestern.edu Cc: Kolman, Kimberly D. Subject: [EXTERNAL] RE: Losing sections I may have an answer for you. Your Prisma stainer only runs the heater when needed. With full runs, the oven stays warm, but later stain runs allow for the heater to cool down. When your rack goes into the dry station, the heat comes up from the bottom to start to dry the slide, thus the bottom sections have enough dry time, but the top don't, and they wash off. We used to encounter the same problem and that is what we hypothesized was happening, because, when we ran a blank rack through 10 minutes before loading the late rack, we were fine. Or, when we dried in a 60'C oven for 15 minutes when loading that lone late rack, we were fine. Just an idea, but for us, no more wash offs of top sections. Terri L. Braud, HT(ASCP) HNL Laboratories for Holy Redeemer Hospital 1648 Huntingdon Pike Meadowbrook, PA 19046 Ph: 215-938-3689 Fax: 215-938-2021 Message: 4 Date: Fri, 16 Jun 2023 14:25:31 + From: "Kolman, Kimberly D." Subject: [Histonet] losing sections For your entertainment, I have a bizarre phenomenon to share with the group: I am occasionally losing the TOP section on a deeper cut H&E slide. Full run of daily slides do not show this issue. A deeper slide cut later in the day might, or a deeper slide cut first thing in the morning may, when I've not run a full daily run. I have switched to a different lot number of standard slides. I have used adhesive slides. Distilled water for the water bath same as I have used for 20 years. Using Sta-On adhesive in the water bath. H&E stain done on a Prisma Plus stainer, with no changes in staining procedure. There have been NO changes to any of my procedures. This is a very random happening that is boggling my mind! If any section was going to fall off, I'd think it would be the 2nd section - (last one picked up from the water bath). I've tried to make sure the slide has thoroughly dried before putting it on the stainer. Slides appear clean, and no greasy fingerprints on the slide. The one I always lose is the TOP section/very first level taken. Any ideas? Do I just have a Histo Gremlin here? Thanks for your input. Kim Kimberly D. Kolman, HT (ASCP) Eastern Kansas Health Care System Eisenhower VA Medical Center - Histology 115 4101 S. 4th St. Trfwy. Leavenworth, KS 66048 913-682-2000 x 62537 Fax: 913-758-4193 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Losing sections
I may have an answer for you. Your Prisma stainer only runs the heater when needed. With full runs, the oven stays warm, but later stain runs allow for the heater to cool down. When your rack goes into the dry station, the heat comes up from the bottom to start to dry the slide, thus the bottom sections have enough dry time, but the top don't, and they wash off. We used to encounter the same problem and that is what we hypothesized was happening, because, when we ran a blank rack through 10 minutes before loading the late rack, we were fine. Or, when we dried in a 60'C oven for 15 minutes when loading that lone late rack, we were fine. Just an idea, but for us, no more wash offs of top sections. Terri L. Braud, HT(ASCP) HNL Laboratories for Holy Redeemer Hospital 1648 Huntingdon Pike Meadowbrook, PA 19046 Ph: 215-938-3689 Fax: 215-938-2021 Message: 4 Date: Fri, 16 Jun 2023 14:25:31 + From: "Kolman, Kimberly D." Subject: [Histonet] losing sections For your entertainment, I have a bizarre phenomenon to share with the group: I am occasionally losing the TOP section on a deeper cut H&E slide. Full run of daily slides do not show this issue. A deeper slide cut later in the day might, or a deeper slide cut first thing in the morning may, when I've not run a full daily run. I have switched to a different lot number of standard slides. I have used adhesive slides. Distilled water for the water bath same as I have used for 20 years. Using Sta-On adhesive in the water bath. H&E stain done on a Prisma Plus stainer, with no changes in staining procedure. There have been NO changes to any of my procedures. This is a very random happening that is boggling my mind! If any section was going to fall off, I'd think it would be the 2nd section - (last one picked up from the water bath). I've tried to make sure the slide has thoroughly dried before putting it on the stainer. Slides appear clean, and no greasy fingerprints on the slide. The one I always lose is the TOP section/very first level taken. Any ideas? Do I just have a Histo Gremlin here? Thanks for your input. Kim Kimberly D. Kolman, HT (ASCP) Eastern Kansas Health Care System Eisenhower VA Medical Center - Histology 115 4101 S. 4th St. Trfwy. Leavenworth, KS 66048 913-682-2000 x 62537 Fax: 913-758-4193 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] losing sections
For your entertainment, I have a bizarre phenomenon to share with the group: I am occasionally losing the TOP section on a deeper cut H&E slide. Full run of daily slides do not show this issue. A deeper slide cut later in the day might, or a deeper slide cut first thing in the morning may, when I've not run a full daily run. I have switched to a different lot number of standard slides. I have used adhesive slides. Distilled water for the water bath same as I have used for 20 years. Using Sta-On adhesive in the water bath. H&E stain done on a Prisma Plus stainer, with no changes in staining procedure. There have been NO changes to any of my procedures. This is a very random happening that is boggling my mind! If any section was going to fall off, I'd think it would be the 2nd section - (last one picked up from the water bath). I've tried to make sure the slide has thoroughly dried before putting it on the stainer. Slides appear clean, and no greasy fingerprints on the slide. The one I always lose is the TOP section/very first level taken. Any ideas? Do I just have a Histo Gremlin here? Thanks for your input. Kim Kimberly D. Kolman, HT (ASCP) Eastern Kansas Health Care System Eisenhower VA Medical Center - Histology 115 4101 S. 4th St. Trfwy. Leavenworth, KS 66048 913-682-2000 x 62537 Fax: 913-758-4193 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Losing Sections While Dehydrating
Hello Everyone! I have searched the archives because I am sure this is a problem that has been encountered before, but I have been unsuccessful in finding the right thread. I ran an IHC series dilution on a new primary antibody for NeuN. I am using 35 um rat brain sections that were cut on a cryostat and stored in Cryoprotectant until ICC began. After finishing the ICC protocol, I mounted the slides that were floating in phosphate buffered saline onto slides that I subbed two weeks ago (.5% gelatin subbing). I allowed the slides to dry from Friday afternoon until Tuesday afternoon. I tried to dehydrate them for coverslipping and when they went into the dH20 bath, some of the sections began to fall off of the slides. This continued in the 50% Alcohol bath. All tissue that made it through those first two baths in the series stayed on throughout, but I did lose quite a few sections, especially those that had the lowest concentration of primary antibody. My experience has been that highly concentrated primaries make the tissue a bit sticky, but the best staining generally comes from the least concentrated. Does anyone know what the problem might be? I think it might be a subbing problem but I'm not sure. Thanks in advance. Amanda Madden Research Assistant P.S. The subbing protocol that I used only called for one "dip" into the gelatin solution, and didn't call for any type of acid wash. Also, our cryostat is a Leica CM3050S and I was wondering if any of your labs have established a working range in terms of humidity. Our lab has been experiencing very high humidity levels (up to almost 80%), so we were wondering if anyone has established that cutting cannot be done unless the humidity is under x%. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
