Re: detecting apoptosis in one cell type in coculture

Posted by samuel connell on
URL: http://confocal-microscopy-list.275.s1.nabble.com/detecting-apoptosis-in-one-cell-type-in-coculture-tp7584201p7584205.html

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Hi Leo,

One idea would be to load the adherent cells with TMRE or TMRM. These dyes
bind to mitochondria with high mitochondrial transmembrane potential and
then are released from the mitochondria when this dissipates. One would
probably prefer to witness cytochrome C release from the mitochondria, but
this is a more time consuming assay design with GFP cytochrome C. Once you
have these cell lines expressing cleanly, those assays are relatively
straight forward.

There's some other related ideas in a Methods paper I worked on a while
back with some fantastic long term collaborators in Doug Green's lab.

Measuring apoptosis at the single cell level.

Free access is available now:

http://www.ncbi.nlm.nih.gov/pubmed/18314052

Cheers,
--
Sam

On Wed, Sep 9, 2015 at 9:31 AM, Leoncio Vergara <[hidden email]>
wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Good morning
>
> We are working on setting up an image screening assay in which we want to
> measure apoptosis in one cell type (adherent)  after exposure to T cells
> (non adherent). the adherent cells are grown on a monolayer and the T cells
> are added on top and incubated for a few hrs (~3-5 hrs). We want to study
> apoptosis in the adherent cells without interference from the T cells.
> CellEvent (Thermo Fisher) works great in other conditions and is ideal for
> a screening assay, but in this case it does not work because of the
> presence of the Tcells, We would like to know is there is an alternative
> indicator we can load in the adherent cells and wash prior to the T cells.
> We need to test multiple cell lines so fluorescent protein based indicators
> would not be ideal.
>
> Thanks in advance for your help.
>
> Leoncio Vergara MD
> Co-Director
>
> Center for Advanced Imaging (CAI) at the
>
> Center for Translational Cancer Research (CTCR),
>
> Institute for Biosciences and Technology,
>
> Texas A&M Health Sciences Center,
>
> Houston, Texas 77030
>