Re: [Histonet] Katushka (Far-red fluorescent protein TurboFP63) ex-vivo

to keep the signal in the same channel as Katushka itself Anatoli Gleiberman, PhD Director of Histopathology Buffalo Biolabs LLC, 73 High Street Buffalo, NY, 14203 Phone:716-849-6810x354 e-mail:agleiber...@buffalobiolab.com -Original Message- From: Monica Aguilera via Histonet

Re: [Histonet] Davidson's fixative and IHC question

in regular NBF. Anatoli Gleiberman, PhD Director of Histopathology Buffalo Biolabs LLC, 73 High Street Buffalo, NY, 14203 Phone:716-849-6810x354 e-mail:agleiber...@buffalobiolab.com -Original Message- From: Jay Lundgren via Histonet [mailto:histonet@lists.utsouthwestern.edu] Sent

Re: [Histonet] anti p53

DO-1 mouse monoclonal anti-human p53 either from Santa Cruz or from Abcam are the best. Anatoli Gleiberman, PhD Director of Histopathology Buffalo Biolabs LLC, 73 High Street Buffalo, NY, 14203 Phone:716-849-6810x354 e-mail:agleiber...@buffalobiolab.com -Original Message- From: Hughes

RE: [Histonet] HA-tag antibody

for immunofluorescence. Anatoli Gleiberman, Ph.D. Director of Histopathology Buffalo Biolabs LLC 73 High Street Buffalo, NY 14203 Phone: 716-849-6810x354 e-mail: agleiber...@buffalobiolabs.com -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun

[Histonet] RE: Oil Red O staining question

I used successfully Oil Red both on flash frozen and formalin-fixed sucrose cryoprotected samples. Don't see any reason why not use formalin fixation. Anatoli Gleiberman, Ph.D. Director of Histopathology Buffalo Biolabs LLC 73 High Street Buffalo, NY 14203 Phone: 716-849-6810x354 e-mail

[Histonet] RE: replacement for Dako's anti-mouse Ki67

Michele, BD Pharmingen, cat.# 556003, mouse monoclonal. I have used extensively the same clone B56 labeled with AlexaFluor488 and 555 - all they work very well on mouse and human tissues. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203

[Histonet] RE: Anti-human nuclear antibody

Elizabeth, We are able to detect human cells in mice organs using MAB1273 anti-mitochondria antibody (1:50) from Millipore on FFPE sections after standard HIER retrieval. As a detection reagent we use MaxFluor 594 Mouse on Mouse IF detection kit (MaxVision, cat.No MF03), Anatoli Gleiberman

RE: [Histonet] Smooth muscle cell actin

frozen and formaldehyde fixed samples, paraffin after formalin etc). I work with it last 8 years without any problem. They are relatively expensive, but last for long due to high working dilution. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo

RE: [Histonet] Permanent Fluorescent Mounting Media

I tried it - got a free sample, liked how it looks (and especially price - two-times less than ProLong Gold)and ordered couple of bottles. Unfortunately they polymerized very fast and it was impossible to revive them. So, we switched back to ProLong. Anatoli Gleiberman, PhD Director

RE: [Histonet] Regarding a problem in manual paraffin embedding

(Alternative fixation - 6 parts of absolute ethanol, 3 parts of distilled water, 1 part of 37% formaldehyde - 4-6 h, transfer in 70% ethanol overnight) 4 changes of isopropanol, 1h each 3 changes of xylene - 45 min each 3 changes of paraffin 1h each. Anatoli Gleiberman, PhD Director of Histopathology

RE: [Histonet] Albumin and Epcam to stain mouse cell pellets

on cryo-sections after fixation by perfusion with 4% paraformaldehyde in PBS. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail: agleiber...@cbiolabs.com -Original Message- From

RE: [Histonet] cell membrane

Liz, It is possible to play with different catenins (gamma etc.) to get rid of nuclear staining. But if the target is epithelium I would try to use EpCAM first. And in case of different epithelial tumors EpCAM usually is up-regulated. Anatoli Gleiberman, PhD Director of Histopathology Cleveland

RE: [Histonet] Fluorescence-filters

Filters are usually very stable. What is burning out are fiber optic cables that are present in some illumination systems such as X-Cite in Zeiss microscopes. These optical fiber cables should be replaced after 2000 hours. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs

[Histonet] RE: Monoclonal antibodies from different companies

Agree with Tresa. It could happen because some companies purchased original clones (not Ig concentrate for further distribution) and screw them. I had such problems occasionally, in particular with now extinct Chemicon. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc

[Histonet] GI fix

Hi histoneters, Does anyone have experience with IHC on samples fixed with GI Fix from BBC? Any differences from routine NBF? Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail

[Histonet] blocking endogenous peroxidase

% peroxide is also effective, but in some cases it causes extensive bubbling that can damage fragile sections. So, I am using methanol-peroxide for paraffin sections and stable peroxide buffer (buffer provided in DAB developer kits, such as Pierce metal-enhanced DAB kit) for cryo- sections. Anatoli

RE: [Histonet] Stain and component shelf life

I am still using some stains from late 1800 to yearly 1900 (azocarmine, pyronin, indigocarmine) from IG Furberindustrie - and they are better than contemporary from Sigma. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849

[Histonet] RE: Using methyl benzoate on a VIP 1000

- and smells much better, for sure. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail: agleiber...@cbiolabs.com -Original Message- From: histonet-boun...@lists.utsouthwestern.edu

[Histonet] RE: Abs for detecting immune cells in mouse FFPE sections

CD3 Rabbit monoclonal from Fisher (MA1-39552) works fine in IF on mouse paraffin sections, B220 rat monoclonal from eBioscience (14-0452-85) works fine in IF on mouse paraffin sections. Used both in 1:100 dilution. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73

[Histonet] Bright M3500 microtome

Hello everyone, Looking for opinion about Bright M3500 microtome - any advantages in comparison to Microm HM325 or Leica RM2235? Thanks Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail

RE: [Histonet] (no subject)

You can freeze samples (either fresh or formalin fixed, but in later case with sucrose cryoprotection) in 7% gelatin in PBS instead of OCT. Sometimes it gives much better conditions for cutting as well. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street

[Histonet] RE: Frozen eye problems

Try Neg-50 (Fisher, cat#22-110-617) instead of OCT, we don't have any problem with fresh-frozen mouse eyes. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail: agleiber...@cbiolabs.com

[Histonet] RE: Ki-67/PHH3 double staining

there is a little masking of Ki67 rather than PHH3. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail: agleiber...@cbiolabs.com -Original Message- From: histonet-boun

RE: [Histonet] BRDU

with all range of antigens(for multi-color fluorescence). Unfortunately, there are no enzymatic reaction for EdU - only fluorescence. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail

RE: [Histonet] Neuronal staining mouse limb sections

Hi Hendrik, Try rabbit neuro-specific beta-III tubulin antibody (Abcam ab18207). Works fine on frozen and paraffin sections. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail: agleiber

RE: [Histonet] re: triple immunofluorescence for apoptosis (bluefluorophore?)

this case, you will have four color fluorescence. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail: agleiber...@cbiolabs.com -Original Message- From: histonet-boun

RE: [Histonet] EGFP expression in mouse testis cryosections

case. For brain, intestine and even liver I usually use fluorescence. Good luck. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail: agleiber...@cbiolabs.com -Original Message- From

[Histonet] RE: Short term tissue storage PBS???

Jo-Ann, Nothing is changed, but storage in PBS does not affect neither morphology nor antigens. I used to keep some neutral formalin- or formaldehyde-PBS fixed specimens up to 1 year in sterile PBS at +4 C without any visible changes. So - don't worry. It will be fine. Anatoli Gleiberman, PhD

[Histonet] RE: 53BP1 and gamma-H2AX antibodies

Rabbit anti-Phospho-Histone H2A.X (Ser139) Antibody #2577 from Cell Signaling works very well in FFPE(dilution 1:100 in immunofluorescence). Don't know good anti 53BP1. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810

RE: [Histonet] fixation question for IHC

is quite sensitive to coagulation induced by the presence of small amount of alcohol. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail: agleiber...@cbiolabs.com -Original Message

[Histonet] RE: Beta Galactosidase antibody

Hi Margaert, Try this one from Aves lab: http://www.aveslab.com/products/epitope-tag-and-gfp-antibodies/b-gal-b-galactosidase-lacz-antibody/ I did not try them on paraffin, but it worked fine on formaldehyde-fixed cells and cryo-sections from formaldehyde-fixed embryonic tissue. Anatoli

[Histonet] RE: dual fluorescent staining

Hi Ross, Use sequential staining with monovalent anti-mouse Fab fragments (donkey or goat) from Jackson ImmunoResearch labeled with different fluorochromes - DyLight488 and DyLight594, for example. As blocking step I use 5% normal donkey serum. Usually it works fine. Anatoli Gleiberman, PhD

RE: [Histonet] hi-profile blades

We use AccuEdge low and high profile from Sakura and several low and high profile blades from Sturkey (Silver, Gold and Extremus) and found that Sturkey knifes are as good as AccuEdge (both for paraffin and cryo sections) and significantly cheaper if you buy it directly from the company. Anatoli

RE: [Histonet] Immunofluorescence double labeling with two rabbitantibodies

mouse monoclonals. Worked for me. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail: agleiber...@cbiolabs.com -Original Message- From: histonet-boun...@lists.utsouthwestern.edu

RE: [Histonet] post-fixation question

Hi Inan, 2h post-fixation for immunohistochemistry on mouse brain is usually more than enough. We use 1h post-fixation at room temp for mouse brain following storage (sometimes, for many weeks) at +4 C in PBS before sucrose cryo-protection, embedding and sectioning. Anatoli Gleiberman, PhD

RE: [Histonet] Necrotic and apoptotic cells

are usually impregnated with plasma proteins - and bright staining for endogenous IgG is easy and reliable way to visualize such cells. Works well on FFPE sections after citrate boiling retrieval. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203

RE: [Histonet] NFkB p65

FFPE after retrieval by citrate boiling. Anatoli Gleiberman, PhD Director of Histopathology Cleveland Biolabs, Inc 73 High Street Buffalo, NY 14203 phone:716-849-6810 ext.354 fax:716-849-6817 e-mail: agleiber...@cbiolabs.com -Original Message- From: histonet-boun