I wish to image cells during 72h. I wish to stain them with a nuclear dye in order to facilitate the cell tracking process (I will use CFSE in parallel). I was thinking to Hoescht 33342 which is quite bright but I'm not sure about how toxic it could be for my cells after 72h.
I wish to avoid to have to transfect them with a NLS-fluorophores.
What should I use ?
Hi,
I've successfully used Hoechst 33342 for live imaging of random cell migration in different types of breast cancer cell lines. Staining with a low concentration (100 ng/ml) for 45 min. was enough for live imaging. Colleagues use the same set up, and image either confocal or epi-fluorescent, between 12h live and 48h live, and in a few studies even 72h live. So it's definitely possible.
Make sure you set your exposure low (ie low laser power/low epi exposure). Depending on your cell line and it's default motility, find an optimal imaging interval for tracking. You can always enhance the Hoechst signal after the experiment if you only use it for tracking.
good luck!
Take a look at this article. It may answer your question.
http://www.ncbi.nlm.nih.gov/pubmed/20931137
Thanks Andre for your quick reply. I saw this article last night after posting my question. I will take a look and try what they advised.
Thanks
Hi,
I've successfully used Hoechst 33342 for live imaging of random cell migration in different types of breast cancer cell lines. Staining with a low concentration (100 ng/ml) for 45 min. was enough for live imaging. Colleagues use the same set up, and image either confocal or epi-fluorescent, between 12h live and 48h live, and in a few studies even 72h live. So it's definitely possible.
Make sure you set your exposure low (ie low laser power/low epi exposure). Depending on your cell line and it's default motility, find an optimal imaging interval for tracking. You can always enhance the Hoechst signal after the experiment if you only use it for tracking.
good luck!
Thanks Michiel for your feedback, I will try low concentration / low exposure / bigger intervals. Hopefully, it should be ok.
The main aspect is that I don't want it impairs my cell proliferation.
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