[Histonet] CD34 for primate tissue

[Elizabeth Chlipala]

Cathy

Before doing that I would check the sequence homology of the immunogen between human and rhesus.  I would do this for all sources of the antibody, if the company does not want to provide you with the sequence then technical support should be able to blast the sequence for you and get back to you the sequence homology, greater than 70% is worth a shot it may not work or it may not.  You also need to check if it's the same clone it might be the same antibody just distributed by different vendors, check the protein concentration, purification method, isotype, and if there are matching images you are likely looking at the same antibody so you don't want to waste your time with ordering the same antibody but from a different vendor.    Remember information on specification sheets can be inaccurate - this depends upon the vendor, some vendors actually test internally while other vendors rely of user for cross reactivity information so you need to take that information with a grain of salt.  My suggestion is to take a look at the antibody on CiteAb - this website is a bit better than biocompare since it rates antibodies by number of citations, check to see if the other antibodies have publications with your use case.  The link for CiteAb is https://www.citeab.com/ We find with some antibodies that work across species that the primary antibody concentration may fluctuate so a concentration that works in human may not work in the particular species you are working with.  There are many other parameters such as time in fixation between your human tissue and rhesus tissue.  Are you getting any hint of staining or is it just completely negative.

It's really important to spend some significant time upfront when researching antibodies for a new target - you want to biases yourself towards success.  If you are an NSH member I posted a worksheet for target development on the BLOCK you can access it there.

Liz

Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC
Premier Laboratory, LLC
PO Box 18592
Boulder, CO 80308
(303) 682-3949 office
(303) 682-9060 fax
(303) 881-0763 cell
liz at premierlab.com
www.premierlab.com

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Premier Laboratory, LLC
1567 Skyway Drive, Unit E
Longmont, CO 80504

-----Original Message-----
From: Cathy M. Mathis/Comparative Medicine via Histonet [mailto:histonet at lists.utsouthwestern.edu] 
Sent: Thursday, June 30, 2016 8:07 AM
To: histonet at lists.utsouthwestern.edu
Cc: Eddie Martin
Subject: Re: [Histonet] CD34 for primate tissue

Rhesus is a species of old world monkey.  There are a couple of other companies that have this same clone and on their datasheets they say it cross reacts with rhesus monkey.  The CD34s (2) that I have do not state on their data sheets that they cross react with anything except human and yet are the same clone as the companies that say it does.  I guess I will just break down and purchase a different clone that states it works in rhesus monkey.
Cathy

-----Original Message-----
From: Eddie Martin [mailto:edmartin26 at gmail.com]
Sent: Wednesday, June 29, 2016 8:22 PM
To: Cathy M. Mathis/Comparative Medicine
Cc: histonet at lists.utsouthwestern.edu
Subject: Re: [Histonet] CD34 for primate tissue

What species is the kidney you are trying to stain with IHC. Because you mentioned that you used normal human kidney and it worked , but Rhesus kidney didn't stain. I'm wondering if the Rhesus kidney is human or another animal species? The Novocastra CD34 (QBEND10) is intended for human tissue. 

Best,
Eddie Martin, HTL,QIHC

Sent from my iPhone

> On Jun 29, 2016, at 4:43 PM, Cathy M. Mathis/Comparative Medicine <cmmathis at wakehealth.edu> wrote:
> 
> 2 more bits of information about my dilemma; I did try staining 
> without any retrieval - no signal I am using rhesus kidney as a 
> control (getting no signal), but I also ran some human kidney and got beautiful staining with a 20 minute HIER using a pH 8 EDTA buffer.
> So I know the antibodies and my protocols work, just not on my rhesus.  Does anyone know of a CD34 that will work in this species?
> More suggestions please?  Thank you all for your time.
> Cathy
> 
> -----Original Message-----
> From: Eddie Martin [mailto:edmartin26 at gmail.com]
> Sent: Wednesday, June 29, 2016 5:34 PM
> To: Cathy M. Mathis/Comparative Medicine
> Cc: histonet at lists.utsouthwestern.edu
> Subject: Re: [Histonet] CD34 for primate tissue
> 
> Hi Cathy,
> 
> EDTA (Retrieval #2) for 20 minutes on the Bond-Rx should be sufficient. And all that is necessary. Please contact me if you need additional help. 
> 
> Best,
> Eddie Martin, HTL, QIHC
> Edmartin26 at gmail.com
> 
> Sent from my iPhone
> 
>> On Jun 29, 2016, at 8:16 AM, Cathy M. Mathis/Comparative Medicine <cmmathis at wakehealth.edu> wrote:
>> 
>> Good morning my fellow Histo-netters, Does anyone have experience 
>> with any CD34 antibody that would work in FFPE rhesus tissues?  A few companies have one with clone QEBend 10 and say that it works but I have had no luck.  I am using the Bond RX polymer system and I've tried all the epitope retrievals from Leica.  Now I am try epitope retrieval offline; pronase, trypsin, pressure cooker with high and low pH solutions.  Still no signal.  Any help would be greatly appreciated.
>> Cathy
> 


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