Re: [Histonet] rolling sections
Try changing the angle of the knife blade, so that the clearance angle (angle between the knife blade and the block face) is larger. In other words, tip the top of the blade towards the block more. If there are numbers on the side of the knife holder, you want to move it to a larger number (like from 5 to 10). Before sectioning, remember to move the block holder towards you, since the blade will now be closer to the block, and you don't want to ker-chunk the block. And remember, when done, to return the clearance angle back to it's usual location, so you aren't curling all your ribbons. So look at the number it is usually set at, or, if there is no number, make some marks on the side of the knife holder, that you can line up again. Peggy A. Wenk, HTL(ASCP)SLS -Original Message- From: Roberta Horner Sent: Thursday, June 12, 2014 10:42 AM To: Histonet (histonet@lists.utsouthwestern.edu) Subject: [Histonet] rolling sections I have some researchers that want to do PCR. They want 10 - 10u sections in a micro-centrifuge tube. The only way to get the sections in the tube is for the sections to roll. How do you get sections to roll when you want them to roll? I've tried room temperature, on ice, brand new sharp blade, dull blade and I can still get some really nice ribbons. When I want a thick ribbon it will roll, darn that Murphy and his laws. Roberta Horner Animal Diagnostic Lab Penn State University ___ 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] Rolling sections
If your cryostat is set up correctly (ie, the roll plate isn't causing the rolling up), then I would try 100% EtOH instead of 70%, I have no idea why it works but sometimes, it does. Also, when our cryostat starts doing this, I just clean the blade and the knife and walk away for a few minutes--but our rolling-up is due to static electricity. You can also ground the blade by touching it with something metal. I wouldn't change your section thickness--rolling up has nothing to do with that if you've got your cryostat set right. Also don't breathe on your sections--that's not a particularly scientific way to get what you need, and you might need RNAse free sections in the future (RNAse free, your breath is not.) Emily prometheus, thief of light, giver of light, bound by the gods, must have been a book. -mark danielewski, house of leaves > ___ > 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 > > > _ > Confidential information. Unauthorized use or disclosure prohibited. > Refer http://www.singhealth.com.sg/ContactUs/#Disclaimer > > ___ > 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] Rolling sections
May I suggest trying the following. I too have experienced cryosectioned slices rolling up. 1) try cutting sections at -20 instead of -25 (it may be too cold) 2) try cutting sections at 6 microns instead of 8-12 microns 3) allow some frost to build up on the metal plate surface and gently with two brushes hold either side down until it gently sticks then quickly pick the specimen up with your slide. This can be very tedious and slow going but at least I was able to get the sections I needed. 4) the "breath" suggestion also worked, although not consistently, for me. 5) take your gloved "thumb" and quickly place over your sample then immediately section. This works too, but again not consistently-is worth a try. Good luck, Linda >-Original Message- >From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet- >boun...@lists.utsouthwestern.edu] On Behalf Of Robyn Vazquez >Sent: Wednesday, April 29, 2009 9:04 AM >To: Dearolf, Jennifer; histonet@lists.utsouthwestern.edu >Subject: RE: [Histonet] Rolling sections > >Jennifer, >It sounds like the sections are a little thick. Have you tried just >barely blowing on the tissue with your breath to warm the OCT? > >-Original Message- >From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet- >boun...@lists.utsouthwestern.edu] On Behalf Of Dearolf, Jennifer >Sent: Tuesday, April 28, 2009 3:52 PM >To: histonet@lists.utsouthwestern.edu >Subject: [Histonet] Rolling sections > >Greetings, Histonetters! > >First, I wanted to thank all of you that responded to my e-mail a few >years back about freezing small pieces of muscle tissue. We have found >a method that works for us, and if anyone is interested, I would be >happy to share. It still involves the wonderfully explosive isopentane, >but it allows us to freeze fetal guinea pig muscle without artifact. > >I am writing today to ask a question about cutting frozen sections with >a cryostat. We are having problems with the sections rolling once they >come off the knife and before we can get them on a slide. We have a >Microm 505E cryostat, and we cut our OCT mounted specimens at around -25 >degrees C. We use Accuedge high profile blades, cut sections between 8 >and 12 microns thick, and use a brush to pull the sections off. But, >when we remove the brush, the sections roll up. Sometimes, they just >arc up and other times they completely roll into a jellyroll. > >I have tried putting 70% EtOH in a beaker in the cryostat. This method >was suggested to us by a vendor, but it doesn't seem to work >consistently. We can also flatten the sections with a brush, but unless >we are really quick, the sections roll up before we can get them on the >slide. It makes it difficult to get serial sections. > >Any advice would be appreciated. Thanks again for all your help so far. > >Sincerely, >Jenn > >Jennifer Dearolf, Ph.D. >Associate Professor >Biology Department >Hendrix College >1600 Washington Ave. >Conway, AR 72032 >(501) 450-4530 (office) >___ >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] Rolling sections
Jennifer, It sounds like the sections are a little thick. Have you tried just barely blowing on the tissue with your breath to warm the OCT? -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Dearolf, Jennifer Sent: Tuesday, April 28, 2009 3:52 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Rolling sections Greetings, Histonetters! First, I wanted to thank all of you that responded to my e-mail a few years back about freezing small pieces of muscle tissue. We have found a method that works for us, and if anyone is interested, I would be happy to share. It still involves the wonderfully explosive isopentane, but it allows us to freeze fetal guinea pig muscle without artifact. I am writing today to ask a question about cutting frozen sections with a cryostat. We are having problems with the sections rolling once they come off the knife and before we can get them on a slide. We have a Microm 505E cryostat, and we cut our OCT mounted specimens at around -25 degrees C. We use Accuedge high profile blades, cut sections between 8 and 12 microns thick, and use a brush to pull the sections off. But, when we remove the brush, the sections roll up. Sometimes, they just arc up and other times they completely roll into a jellyroll. I have tried putting 70% EtOH in a beaker in the cryostat. This method was suggested to us by a vendor, but it doesn't seem to work consistently. We can also flatten the sections with a brush, but unless we are really quick, the sections roll up before we can get them on the slide. It makes it difficult to get serial sections. Any advice would be appreciated. Thanks again for all your help so far. Sincerely, Jenn Jennifer Dearolf, Ph.D. Associate Professor Biology Department Hendrix College 1600 Washington Ave. Conway, AR 72032 (501) 450-4530 (office) ___ 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] Rolling sections
Normally, we will use brush to hold the section after we have sectioned one third of the sample. After that, we will continue section the sample untill the end. If the sample is rolled up, we use the brush to smoothen it. Then quickly flip the glass slide to fish the sample. As the glass slide is keep in room temperature (warm), when put inside the cryostat machine (inside is cold), the section will just stick to the glass slide. This works quite well in our lab. Hope this can help... Regards, Ooi "TF" Sent by: histonet-boun...@lists.utsouthwestern.edu 04/29/09 11:00 AM Please respond to ti...@foxmail.com To "Dearolf, Jennifer" , "histonet@lists.utsouthwestern.edu" cc Subject Re: [Histonet] Rolling sections I also have problems in mounting brain sections...10um-30um in Leica Cryostathope anyone can share their experience? sometimes we use a brush to paint a mini drop of PB/water on to the slide, and then mount the sections. it is very flat then. 2009-04-29 TF 发件人： Dearolf, Jennifer 发送时间： 2009-04-29 06:56:50 收件人： histonet@lists.utsouthwestern.edu 抄送： 主题： [Histonet] Rolling sections Greetings, Histonetters! First, I wanted to thank all of you that responded to my e-mail a few years back about freezing small pieces of muscle tissue. We have found a method that works for us, and if anyone is interested, I would be happy to share. It still involves the wonderfully explosive isopentane, but it allows us to freeze fetal guinea pig muscle without artifact. I am writing today to ask a question about cutting frozen sections with a cryostat. We are having problems with the sections rolling once they come off the knife and before we can get them on a slide. We have a Microm 505E cryostat, and we cut our OCT mounted specimens at around -25 degrees C. We use Accuedge high profile blades, cut sections between 8 and 12 microns thick, and use a brush to pull the sections off. But, when we remove the brush, the sections roll up. Sometimes, they just arc up and other times they completely roll into a jellyroll. I have tried putting 70% EtOH in a beaker in the cryostat. This method was suggested to us by a vendor, but it doesn't seem to work consistently. We can also flatten the sections with a brush, but unless we are really quick, the sections roll up before we can get them on the slide. It makes it difficult to get serial sections. Any advice would be appreciated. Thanks again for all your help so far. Sincerely, Jenn Jennifer Dearolf, Ph.D. Associate Professor Biology Department Hendrix College 1600 Washington Ave. Conway, AR 72032 (501) 450-4530 (office) ___ 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 _ Confidential information. Unauthorized use or disclosure prohibited. Refer http://www.singhealth.com.sg/ContactUs/#Disclaimer ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Rolling sections
I also have problems in mounting brain sections...10um-30um in Leica Cryostathope anyone can share their experience? sometimes we use a brush to paint a mini drop of PB/water on to the slide, and then mount the sections. it is very flat then. 2009-04-29 TF 发件人： Dearolf, Jennifer 发送时间： 2009-04-29 06:56:50 收件人： histonet@lists.utsouthwestern.edu 抄送： 主题： [Histonet] Rolling sections Greetings, Histonetters! First, I wanted to thank all of you that responded to my e-mail a few years back about freezing small pieces of muscle tissue. We have found a method that works for us, and if anyone is interested, I would be happy to share. It still involves the wonderfully explosive isopentane, but it allows us to freeze fetal guinea pig muscle without artifact. I am writing today to ask a question about cutting frozen sections with a cryostat. We are having problems with the sections rolling once they come off the knife and before we can get them on a slide. We have a Microm 505E cryostat, and we cut our OCT mounted specimens at around -25 degrees C. We use Accuedge high profile blades, cut sections between 8 and 12 microns thick, and use a brush to pull the sections off. But, when we remove the brush, the sections roll up. Sometimes, they just arc up and other times they completely roll into a jellyroll. I have tried putting 70% EtOH in a beaker in the cryostat. This method was suggested to us by a vendor, but it doesn't seem to work consistently. We can also flatten the sections with a brush, but unless we are really quick, the sections roll up before we can get them on the slide. It makes it difficult to get serial sections. Any advice would be appreciated. Thanks again for all your help so far. Sincerely, Jenn Jennifer Dearolf, Ph.D. Associate Professor Biology Department Hendrix College 1600 Washington Ave. Conway, AR 72032 (501) 450-4530 (office) ___ 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] Rolling sections
Are you freezing muscle? If so, for our muscle biopsies we use OCT to adhere the base of the tissue to a piece of cork (which is frozen in the isopentane); the tissue is not surrounded by OCT. The tissue section is guided onto the blade holder with a brush and then picked up on the slide. All our other tissues are frozen surrounded by OCT. Becky Garrison Shands Jacksonville 904-244-6237 -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Dearolf, Jennifer Sent: Tuesday, April 28, 2009 6:52 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Rolling sections Greetings, Histonetters! First, I wanted to thank all of you that responded to my e-mail a few years back about freezing small pieces of muscle tissue. We have found a method that works for us, and if anyone is interested, I would be happy to share. It still involves the wonderfully explosive isopentane, but it allows us to freeze fetal guinea pig muscle without artifact. I am writing today to ask a question about cutting frozen sections with a cryostat. We are having problems with the sections rolling once they come off the knife and before we can get them on a slide. We have a Microm 505E cryostat, and we cut our OCT mounted specimens at around -25 degrees C. We use Accuedge high profile blades, cut sections between 8 and 12 microns thick, and use a brush to pull the sections off. But, when we remove the brush, the sections roll up. Sometimes, they just arc up and other times they completely roll into a jellyroll. I have tried putting 70% EtOH in a beaker in the cryostat. This method was suggested to us by a vendor, but it doesn't seem to work consistently. We can also flatten the sections with a brush, but unless we are really quick, the sections roll up before we can get them on the slide. It makes it difficult to get serial sections. Any advice would be appreciated. Thanks again for all your help so far. Sincerely, Jenn Jennifer Dearolf, Ph.D. Associate Professor Biology Department Hendrix College 1600 Washington Ave. Conway, AR 72032 (501) 450-4530 (office) ___ 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
