[Cytometry] CD4 purification

Jayaraman, Sundararajan anue2468 at uic.edu
Tue Feb 9 21:42:44 EST 2010


Hello,

There are kits available to purify human CD4+ T cells deleted of monocytes
and dendritic cells that express the CD4 determinant weakly. "Untouched"
CD4+ T helper cell kit allows one to purify the CD4+ T cells by deleting
other CD4+ cells. These are available from Miltenyi Biotec and Dynabeads
(Invitrogen). THIS IS NOT A PRODUCT ENDORSEMENT, HOWEVER.

Purified CD4+ T cells are not activated and useful for gene expression
studies. We have experience in gene expression studies using mouse
"untouched" CD4+ T cell kit from Miltenyi. There are many reports using
human CD4+ T cell enrichment kits to polarize distinct T helper subsets
from enriched CD4+ T cells and for subsequent gene analysis. Check it on
the PubMed.

Check the data sheet for these products from both of the companies
mentioned above and choose what is convenient for you.

 Regards.
-- 
S. Jayaraman, Ph.D.
Associate Professor of Surgery
University of Illinois at Chicago

On Tue, February 9, 2010 3:59 pm, Howard Shapiro wrote:
> Is it your intention to study mRNA in CD4+ T cells? If so, you will need
> to separate them from the CD4+ monocytes/macrophages also present in PBMC.
> If you did this by positively selecting with CD3, you would stimulate the
> CD4+ T cells, so getting rid of the monocytes with CD14 and/or other
> antibodies that they express and CD4+ T cells do not would be a better
> bet.
>
> -Howard
>
>
>
> -----Original Message-----
>>From: Mous Kim <kim.mous at ua.ac.be>
>>Sent: Feb 9, 2010 3:48 AM
>>To:
>>Cc: cytometry at lists.purdue.edu
>>Subject: [Cytometry]  CD4 purification
>>
>>Dear all,
>>
>>Could anyone comment on the following please?
>>
>>I intend to perform mRNA expression analyses through real-time qPCR on
>> CD4 cells purified with magnetic beads starting from frozen PBMC. I'd
>> prefer purifying through positive selection since cell populations tend
>> to be purer compared to negatively selected cells. I do not intend to
>> stimulate the purified CD4 cells prior to putting them into trizol. Has
>> anyone of you observed that cells became stimulated / that proteins/mRNA
>> levels increased due to the CD4 positive selection?
>>
>>Thank you in advance for your comments/suggestions !!
>>
>>Best regards,
>>Kim
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