RE: [Histonet] OT: Stay Safe during Sandy
Why, thank you so much, Pam! High winds/heavy rain due to hit Buffalo area starting around 5PM. Regards, Merced -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Pam Barker Sent: Monday, October 29, 2012 11:18 AM To: Histonet Subject: [Histonet] OT: Stay Safe during Sandy Hi Histonetters!! Stay safe and warm during this storm. I am praying for you and all of my friends and family up north. I hope Sandy doesn't cause damage or disruption to you and yours. Thank You! Pam Barker President/Senior Recruiting Specialist-Histology RELIA Solutions Specialists in Allied Healthcare Recruiting 5703 Red Bug Lake Road #330 Winter Springs, FL 32708-4969 Phone: (407)657-2027 Cell: (407)353-5070 FAX: (407)678-2788 E-mail: rel...@earthlink.net www.facebook.com http://www.facebook.com/PamBarkerRELIA /PamBarkerRELIA www.linkedin.com/in/reliasolutions www.twitter.com/pamatrelia ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet - No virus found in this message. Checked by AVG - www.avg.com Version: 2013.0.2742 / Virus Database: 2617/5861 - Release Date: 10/29/12 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Why people are having issues unsubscribing
There are 3 reasons why it is not immediately obvious to people that one of the links at the bottom of Histonet email helps you unsubscribe: 1. In this day and age of internet and email hypertext clutter people simply do not have the time nor the desire to click and explore every link they come across to see which one is going to lead them to the desired page. If the hypertext of a link does not jump out at a person with the info they want, they disregard it and keep looking or freak out and ask the whole list. Neither link at the bottom of Histonet email explicitly states the info most people are probably looking for - that is, to unsubscribe. 2. Most email (junk or subscription) that people receive contain a link that states, To unsubscribe CLICK HERE, or words to that effect. I believe this is what most people are looking for who are trying to unsubscribe from Histonet. 3. Most people trying to unsubscribe are not even reading well-intentioned email sent from Histonetters trying to help them. They see [Histonet] in the subject line and disregard it because they're trying to unsubscribe, not read more Histonet email. Just trying to facilitate some understanding here of the problem. That said, my suggestion to the list moderator would be to include a link at the bottom of Histonet email that explicitly states To unsubscribe CLICK HERE. This should reduce some of the clutter of unwanted and unnecessary off-topic (i.e., unsubscribe me please!!!) Histonet email. Regards, Merced Merced M Leiker Cardiovascular Medicine Biomedical Research Building Rm 348 State University of New York at Buffalo 3435 Main St., Buffalo, NY 14214 (Ph) 716-829-6118 (Fx) 716-829-2665 __ 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] Cardiac Myocytes
We use Cardiac Troponin I or Cardiac Troponin T all the time (pigs). These are also in mice. Regards, Merced From: histonet-boun...@lists.utsouthwestern.edu [histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Amos Brooks [amosbro...@gmail.com] Sent: Tuesday, February 28, 2012 3:18 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Cardiac Myocytes Hi, What would one use to identify cardiac myocytes in mice. I know there are specific antibodies, but I was kinda hoping for something a bit more mundane like desmin. Any ideas? Amos ___ 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] autofluorescence and sudan black
It should not interfere. I use Sudan Black a lot. What concentration do you use? Regards, Merced From: histonet-boun...@lists.utsouthwestern.edu [histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Carmen Maria Garcia Pascual [carstj2...@hotmail.com] Sent: Wednesday, February 29, 2012 7:50 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] autofluorescence and sudan black Good morning!I am Carmen from the University of Valencia. Now I am performing immunofluorescence of mouse decidua. I was having problems with the autofluorescence so I found a sudan black protocol and I probed it, I incubated the tissue 30 min with sudan black after the staining, but I lostes the red signal (alexa 594) so I repited the experiment but incubating before the immunohistochemestry, and want to know if you know if this migth interfere with the antibody attachment to the tissue.thank you very much!kind regards!Carmen ___ 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] Immunofluorescence
Paraffin does indeed give a lot of autofluorescence. We use a solution of 0.3% Sudan Black in 70% Ethanol for 10 minutes on the tissues to help with that after the staining. It doesn't mask the autofluorescence completely but reduces it enough to give us good contrast of signal against the background. Mostly we look at cardiac tissue. Maybe someone will have a better idea for brain... Regards, Merced -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Natalia Fernandez Sent: Tuesday, February 07, 2012 10:35 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Immunofluorescence Hi everybody! I have a problem with my experiment. I try to do immunofuorescences in old human brains (parafine sections). First I thought that the too much colocalisation was due to my antibodies (primary or secondary). After several tests (Only 1st antibody, only 2nd one, simple staining, inverse simple staining,..) I saw that the tissue itself shows a lot of fluorescence (without antibodies, and even after a treatment against lipofuscine (solution of Potassium permanganate). So my question is: Do you have the same problem? Do you think that parafine could be the reason of this autofluorescence?! What can I do? Thank you so much for your answers :) Natalia ___ 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] Nail Polish sealant
Interesting. I've never had an issue. Regards, Merced -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Andrea Hooper Sent: Thursday, January 12, 2012 10:04 PM To: gayle callis Cc: Histonet; kas...@mail.nih.gov Subject: Re: [Histonet] Nail Polish sealant Amen Gayle, as always you hit the nail on the head. I never use nail polish sealant because of being burned so many times due to the isopropanol leaching into - and ruining - the sample. In addition if it gets onto pricey fluorescence lenses it's difficult to clean. It's not worth the risk, especially given the great hard set mountants available. I also detest the mountants with DAPI mixed in. Nothing better tan doing your own even counter stain with Hoechst 33342 for 10 minutes before mounting for crisp beautiful nuclei. Andrea On Jan 5, 2012, at 12:35 PM, gayle callis gayle.cal...@bresnan.net wrote: You wrote: We are currently starting up some IHC on frozen tissue sections. After staining with different fluorescent antibodies, we end with applying DAPI w/Prolong gold and then coverslipping. We would like to seal the coverslip so that we can keep the slides longer. Any suggestions on where and how best to apply the nail polish for a permanent fix on the coverslips? * Prolong Gold antifade reagent is a hard seal to begin with. We only seal ends but not the sides of cover glass. Our coverslips go right to edge of slides e.g. 25 X 30, 25 x 40, or 25 X 50.We don't like having nail polish slop over the edges onto back of slides but one could seal all sides of coverslip if careful. We buy the thinner top or base coat nail polish, but in general, prefer to use thinned mounting media rather than nail polish. There can be some issues here.If you are trying to view GFP or RFP (red fluorescence protein) labeled cells or tissue components, you should not seal the coverslip with nail polish since the alcohol in nail polish leaches under the coverslip and causes GFP/RFP to fade. This fact was published in Science. We found that dumping out cheap clear nail polish from bottle, rinsing away the residue with acetone, and then adding permanent mounting media and thinning that with toluene to the consistency of top coat nail polish works best. Toluene or xylene based sealants cannot leach under the cover glass since these solvents are NOT miscible with water in the PBS.Thinned mounting media is better sealant for GFP purposes (no fading) and also works for IF stained sections (perfect seal). We love the little brush in the nail polish bottle for application. Thicker clear nail polish (for non GFP studies) or IF stained sections is messy during application so we buy the cheapest top coat polish we can find at Walmart. DAPI in the Prolong Gold will cause an uneven staining gradient so that some of the nuclei in the center of a section are not stained as brightly as the nuclei on the outer edges of a stained section. The cause is not getting enough thicker Prolong Gold/DAPI over the section or not having just the right amount of buffer on the section to permit a good flow of this wonderful mounting media over the section.We now complete all IF staining then stain with a DAPI solution before cover slipping with Prolong Gold. You can buy ready to use DAPI solutions from Pierce or Biogenex, or make up the solution in house. You can find the recipe at IHCworld website or simply Google. We do NOT store our IF stained slides in the cold, but in a folder at RT in a dark drawer before viewing on the day after staining to reduce any movement/flow under the coverslip. Fluorophores can and will eventually fade. I do not recall any studies saying storing IF stained slides in the cold reduces fading but we never have space to do cold storage anyway and store slides at RT.The new fluorophores (Alexas and Dylights) remain stable over a longer time even for several weeks compared to fluorescein derivatives e.g. FITC TRITC. Gayle M. Callis HTL/HT/MT(ASCP) Bozeman MT ___ 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
[Histonet] stripping antibodies
Anyone know a buffer or method (not using heat) to strip antibodies off cells used in an immunocytostaining procedure? The cells are in a multi-well tissue culture dish and have only been labeled with primary (unconjugated) antibody. Thanks. Merced M Leiker Cardiovascular Medicine Biomedical Research Building Rm 348 State University of New York at Buffalo 3435 Main St., Buffalo, NY 14214 (Ph) 716-829-6118 (Fx) 716-829-2665 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] RE: Using Nail Polish on coverslip?
I'll seal the coverslip around all the edges and let it dry in the hood for up to half an hour (because of the smell). I use a clear color that doesn't fluoresce under the miscroscope and also allows the slide to be stored at low temp (-20) without the nail polish seal cracking. Also be sure the polish is toluene- and benzene-free. We've discovered Revlon #771 Clear fits these parameters and works very well. Cheaper nail polishes seem to crack or flake off at cold temps. Regards, Merced -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Kasai, Miki (NIH/NCI) [E] Sent: Wednesday, January 04, 2012 11:14 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Using Nail Polish on coverslip? We are currently starting up some IHC on frozen tissue sections. After staining with different fluorescent antibodies, we end with applying DAPI w/Prolong gold and then coverslipping. We would like to seal the coverslip so that we can keep the slides longer. Any suggestions on where and how best to apply the nail polish for a permanent fix on the coverslips? Thanks, ___ 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] microtomes
Do you have access to a confocal microscope or one that does Z-stack imaging? You can find out the thickness of your tissue sections that way (after you've mounted them on a slide of course). Regards, Merced -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Amos Brooks Sent: Friday, November 18, 2011 8:55 AM To: histonet@lists.utsouthwestern.edu; sseg...@slu.edu Subject: [Histonet] microtomes Hi, If you are concerned about the thickness of the sections being accurate to the setting, I would suggest picking up a cheap micrometer from a hardware store. (OK perhaps not cheap as you will want a quality one, but the cost of these isn't terrible.) You can't really measure 5 microns (or whatever you cut at) well, so you will need to take 10-50 sections of a blank block. Remember to keep a constant rhythm that you would use on a normal block. Measure the thickness before and after then divide these thicknesses by the number of sections taken. Remember, also, to take the measurements at the same temperature that you are cutting the sections at. If your difference isn't exactly right how far off is it? You would then know if you should use a higher or lower setting on the microtome. Amos On Thu, Nov 17, 2011 at 10:33 PM, histonet-requ...@lists.utsouthwestern.edu wrote: Message: 13 Date: Thu, 17 Nov 2011 16:02:50 -0600 From: Salomao Segal sseg...@slu.edu Subject: [Histonet] microtomes To: histonet@lists.utsouthwestern.edu Message-ID: cacdnk8huzek7dgaept--kjzy219ynwgmepiwq5dkorpjtn8...@mail.gmail.com Content-Type: text/plain; charset=ISO-8859-1 The settings in a rotary microtome may indicate the thickness of sections but... how do you know that it indeed cuts at the indicated thickness, particularly if it is say an old device that you inherit from somebody else's lab junk? Is there a way of measuring the magnitude of advances after each rotation? Thanks Solomon Segal ___ 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] cochineal mentioned in comic strip
Thanks for forwarding these links. I found the comic strip and Anatech article fascinating (in their own rights). Regards, Merced -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Mark Ray Sent: Monday, October 24, 2011 8:32 AM To: histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] cochineal mentioned in comic strip The history of logwood, the source of hematoxylin, was similar. People have always wanted colors and dyestuffs were very precious in the millennia before the advent of practical organic chemistry. It's also interesting to consider that Carmine made from Cochineal is a popular red food coloring. It's likely that we've all eaten this bug byproduct. On 10/23/2011 6:04 PM, Lee Peggy Wenk wrote: Just for fun: Check to see if your Sunday comic section carries Jump Start. Check out today’s 10/23/11 strip. Jump Start, a comic strip about a couple (policeman and nurse) and their kids, has the oldest girl wanting to be a cochineal insect for Halloween – which is where histology gets carmine dye for the mucicarmine stain. http://www.gocomics.com/jumpstart/2011/10/23 If you want to read a fascinating book about the the role of carmine in the exploration of the America’s, enslaving the people of Central and South America, pirates stealing ships loaded with the dye, spying, politics, government and religion, and the “unions” of the dyeing industry back then – find or buy a copy of “A Perfect Red: Empire, Espionage, and the Quest for the Color of Desire” by Amy Butler Greenfield, 2006. If you want a more abbreviated version, Anatech’s newsletter ”The Innovator” had an article about carmine in their Winter 2007 issue – all about the history, and about why the quality of Mucicarmine has gone downhill in the past few years. (And also towards the end of the newsletter - what Anatech has done to try to improve the quality of the carmine. The article includes promoting their version of mucicarmine, so just a head’s up – this is their newsletter to promote their products. But they do a great job at educating in general, too. So I enjoy reading and learning from their newsletters. Great photos of what stains SHOULD look like.) http://www.anatechltdusa.com/Innovators/Innovator12_06.pdf No – I don’t get any money talking about the comic strip, the book or Anatech. I just think it’s neat to read about the history of dyes. And really great to to read about cochineal in a Sunday comic strip! Peggy A. Wenk, HTL(ASCP)SLS Beaumont Hospital Royal Oak, MI 48073 (The opinions expressed are my own.) ___ 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
[Histonet] RE: pouring of liquid nitrogen into a cryostat
I would definitely think it was cause components of the chamber as well as the chamber itself to crack. Liquid nitrogen is MUCH MUCH colder than any temperature your chamber would be, even at its coldest. The temp difference would be too extreme. Just my opinion having a bit of experience playing around with the stuff. Regards, Merced -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Kaye Ryan Sent: Friday, October 14, 2011 12:36 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] pouring of liquid nitrogen into a cryostat Hi Everyone, Has anyone ever heard of pouring liquid nitrogen into the chamber of a cryostat that has warmed up to help cool it down more quickly? Does this do harm to the chamber or mechanisms of the cryostat? Any information would be appreciated. Thanks ___ 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] Antigen retrieval survey
96 degree water bath for half an hour in either citrate buffer (pH 6) or Tris-HCl (pH 9), depending on which gives better retrieval for that antibody. -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Andrea T. Hooper Sent: Wednesday, July 13, 2011 11:10 PM To: Histonet Subject: [Histonet] Antigen retrieval survey Hi All, I am doing a survey and will be happy to compile results and share if folks will respond! What is your favorite antigen retrieval method and/or panel? Buffer Source/composition Temperature Device Thanks, Andrea ___ 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] Vasculature in tumors
I agree... I've used CD31 to stain tumor vasculature. Some tumor vasculature isn't even lined with endothelium; there's just blood channels. But most should be lined. Regards, Merced -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of JR R Sent: Friday, July 01, 2011 1:55 PM To: histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Vasculature in tumors You are probably looking for capillaries maybe arterioles so there wont be any elastic lamina for the pentachrome stain to detect. Instead try IHC with an endothelial marker like VE Cadherin (best) or in a pinch, CD31 or PECAM. Jerry Ricks Research Scientist University of Washington Department of Pathology Date: Fri, 1 Jul 2011 12:48:33 -0500 From: sgoe...@mirnarx.com To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Vasculature in tumors Hello all and happy long weekend (yes our boss gave us the rest of the day off at 2!!!) Looking to stain vasculature in tumors. I did trichrome, pentachrome, and PAS (with digestion) and am being told that these are not optimal for what they are looking for. Any other ideas to stain blood vessels would be awesome!! Thanks ya'll! Sarah Goebel-Dysart, BA, HT(ASCP) Histotechnologist Mirna Therapeutics 2150 Woodward Street Suite 100 Austin, Texas 78744 (512)901-0900 ext. 6912 ___ 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
