[Histonet] Human nuclear antibody MAB1281

[Eva Permaul]

Liz,
Any chance you could find out what protocol was used by the ones that have
used it? or help me find some of the literature references? Everywhere I
look I just see people who have not gotten it to work asking for help.
Thanks,
Eva

On Thu, Feb 25, 2016 at 10:39 AM, Elizabeth Chlipala <liz at premierlab.com>
wrote:

> Eva
>
> We have worked with various antibodies in order to detect human cells in a
> rodent background.  I know that this antibody has been referenced in the
> literature and I also have known of other individuals who have had success
> with it, but in our hands we have never been able to make it work on FFPE
> tissues but I believe it works in frozen sections.  We have opted to try
> some different antibodies.
>
> NuMA -we researched this protein and determined that it may work -here is
> a snippet of what we found from a quick look into the Human Protein Atlas
> entry for NUMA1. Though it is functionally involved in the structural
> rearrangement of DNA during mitosis and apoptosis, it is a constitutively
> expressed structural protein. From the images on HPA (
> http://www.proteinatlas.org/ENSG00000137497-NUMA1/tissue/primary+data)
> it looks like it is almost universally expressed, with the one exception of
> hepatocytes. The vast majority of neurons, which are overwhelmingly
> post-mitotic, show fairly strong expression, so that's encouraging.   We
> initially looked at the cell signaling antibody (our target was human cells
> in a rat background) but found that it does cross react with rat, its fine
> with mouse but this antibody is our hands was found to be sensitive to time
> in fixation and retrieval time and temp.  We opted to try another source
> for this antibody - abcam.   They had several different antibodies for that
> target.  We work with their antibodies a lot and they normally do not list
> the sequence of the antigen so we could not check the sequence homology to
> other species ourselves but you can call them and they will provide that
> information.  Here is what they provided to us -   "We have not tested
> ab86129 for cross reactivity with rat and we have not received any
> researcher feedback for using the antibody with this species. The immunogen
> sequence has 56% identity with the rat protein, so it is unlikely that it
> will cross react. However we have not confirmed this experimentally and we
> do not know if it would show non-specific staining with rat. I do think
> ab86129 would be the best antibody to try since it has the lowest homology
> with the rat protein. The other antibodies that have not been tested with
> rat have 69% (ab55767) or 88% (ab84680) homology."  Ab86129 worked very
> nicely in our hands.
>
> HLA-1 this will also work nicely but the protein is expressed in the
> cytoplasm and expression levels can vary but it does work nicely in some
> instances.
>
> So for us its dependent upon the project as to which antibody we will use
> but the NuMA is expressed in the nuclei so it will work quite nicely for
> dual staining of another protein that is expressed in the cytoplasm or cell
> membrane.
>
> Good Luck
>
> 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
>
> Ship to Address:
>
> Premier Laboratory, LLC
> 1567 Skyway Drive, Unit E
> Longmont, CO 80504
>
> -----Original Message-----
> From: Eva Permaul via Histonet [mailto:histonet at lists.utsouthwestern.edu]
> Sent: Thursday, February 25, 2016 7:07 AM
> To: histonet
> Subject: [Histonet] Human nuclear antibody MAB1281
>
> Good morning,
> We recently bought MAB1281 with the hope of being able to determine if the
> cells in a mouse model was human or mouse.
> We want to use it to stain FFPE tissues. I have tried it with Citrate per
> the companies recommendation. I ran it on a human tonsil but saw no
> staining at all.
> Is there anyone who has been able to get this antibody to work? And if so
> would you please share your protocol?
> Thank you,
> Eva
> _______________________________________________
> Histonet mailing list
> Histonet at lists.utsouthwestern.edu
> http://lists.utsouthwestern.edu/mailman/listinfo/histonet
>