Mark and Kristen,
Hi, sorry, I didn't realize the target was T-cells expressing no CD3 although I'm not sure what that cell even is unless a T-cell precursor. Thymocytes are derived from a pool of CD3-/CD4-/CD8- T-cell precursors that have transited to the thymus. I didn't realize she wanted to see these also. But soon after, and in the vast majority of lymphocytes in thymus, they do express CD3 (and TCR). Otherwise they can not possibly go through positive and negative selection or anergy to become card carrying T-cells. So yes there are CD3- Tcell precursors in thymus (I guess) and so do see them by SATB1. I guess I do have to not be afraid to try something new and look into this research immunology and histology thing some day. Not exactly sure how diagnostics on transformed cells fits into the topic but I guess I misunderstood what Kristen was looking for. Thanks for the new information.
Ray Koelling
PhenoPath Labs
Seattle, WA
-------------- Original message --------------
From: "Tarango, Mark"
> Kristen & Ray,
>
> I still say SATB1 is the way to go. Kristen wants to stain T-cells that
> don't express CD3 yet. She'll need a T-cell marker that is expressed at
> the earliest point possible (to include all T-cells, even CD3-negative
> T-cells).
>
> Nearly every marker has some overlap with another lineage. She probably
> has already tried CD3, that being the most obvious and probably the
> first thing anyone would think of, when trying to stain a T-cell (but it
> ain't working out and she wants to know of other options). I really
> doubt she's working in a clinical rat lab. She's doing research; but
> even clinical labs end up using research antibodies.
>
> I've used SATB1 on human and mouse and it works great once your protocol
> is optimized. Take it from someone who's actually used it.
>
> How would you tell the difference between something like ALK-negative
> anaplastic large T-cell lymphoma and Hodgkin's lymphoma if the
> morphology is similar and the tumor cells don't express mature B or
> T-cell antigens? You have to go for something that is expressed at an
> earlier stage. I'd chose PAX5 (B-cells) and SATB1 (T-cells) to be able
> to figure it out. CD3 is useless in such a situation.
>
> This is research; don't be afraid to try something new to get you closer
> to your answer.
>
> Mark Adam Tarango HT(ASCP)
> Histology & IHC Supervisor
> Nevada Cancer Institute
> One Breakthrough Way
> Las Vegas, NV 89135
> Direct Line (702) 822-5112
> Treo (702) 759-9229
> Fax (702) 939-7663
>
>
> -----Original Message-----
> From: histonet-bounces@lists.utsouthwestern.edu
> [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of
> koellingr@comcast.net
> Sent: Wednesday, December 19, 2007 10:45 PM
> To: Kristen Broomall; Histonet
> Subject: Re: [Histonet] Rat T Cell Marker
>
> Kristen,
> To me something like CD3 is the way to go if I understand your project
> as you described it. Something like SATB1, which I've never used by IHC
> but know something about, seems like overkill or using a good (but
> incorrect) tool for the job. SATB1 (special AT-rich binding protein 1)
> I'm sure works well in Marks hands and others. But it is a not well
> characterized molecule being a loop-folding domain transcriptional
> regulator and while true found "mainly" in thymocytes, there are
> certainly indications of its presence in myeloid and erythroid
> precursors as well as developing mouse CNS. And it can be upregulated
> and down-regulated. And probably Marks statement that there might be
> anti-rat SATB1 (and SATB2) antibodies around is true, but might not be
> so easy to find or use. Compare that to the absolutely, precisely
> characterized and described gamma/delta/epsilon/zeta/eta CD3 TCR
> complex. It has been studied in minute detail all over the world, its
> presence or absence in cell types
> is documented and redocumented countless times. The number I've heard
> is about 10,000 copies per T-cell and literally defines that cell. If
> you wanted to "mark" T-cells in flow, frozen or paraffin I think 99% of
> labs would use CD3 at some point whether in periphery or thymus. Well
> characterized and protocoled anti-rat CD3 IHC antibodies exist in
> multiple clones and from multiple vendors for rat IHC. I like BD
> Pharmingen but just my opinion. Without ever having stained for SATB1 in
> rat, I believe what was said and guess yes you could do it, the question
> is, is that the easiest and most efficient way to get to your answer?
>
> Ray Koelling
> PhenoPath Labs
> Seattle, WA
>
> -------------- Original message --------------
> From: "Kristen Broomall"
>
> > Good Morning all, I was wondering if anyone had suggestions on an
> antibody
> > to detect T cells (only, no B cells) in formalin fixed, paraffin
> embedded
> > rat thymus and spleen. I have a Pan T Cell marker that is working well
> in
> > the spleen, but so-so in the thymus, so I think that it is only
> staining the
> > mature T cells, and not immature cells (which of course, is what we
> also
> > need to see). I've been reading up alot on T cells and trying to
> figure this
> > out (CD3, CD4, CD8, TCR, lions, tigers and bears - Oh My!), but I fear
> our
> > deadline for getting staining done is shorter than the time I have to
> > educate myself properly!
> >
> > Any suggestions would be greatly appreciated!
> >
> >
> > Happy Holidays,
> > Kristen Broomall, HT (ASCP)
> > histotechkb@gmail.com
> > _______________________________________________
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> > Histonet@lists.utsouthwestern.edu
> > http://lists.utsouthwestern.edu/mailman/listinfo/histonet
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