Triggering apoptosis by UV? - How to do that? (Aug/26/2008 )
I want to trigger intracellular apoptosis by UV treating the cells. Yesterday I put the cells under Lamiar Flow UV but they were still healthy after 30 min.
How can I induce apoptosis by UV?
Do I need 200 mJ/s? how to measure that?
Well, you can induce DNA damage by UV-irradiation of your cells as you describe, however the dose from the typical bulb in a laminar flow hood is rather low, mostly because the the intensity of the radiation is proportional to the distance from the source. Bringing your culture dish closer to the light will help somewhat. In addition, the typical polystyrene culture dish, and the media that the cells are in will both absorb significant amounts of the UV radiation, further lowering your cumulative dose.
Also, how are you determining apoptosis, and how long after irradiation? The reason I ask is that if you are administering sub-lethal doses tat do produce DNA damage, you should see a reduction in cell growth over a period of several days; during this time however, the sub-lethally irradiated cells may be able to rectify the damage and resume proliferation.
If your lab or institution has a UV cross linker it may be a better choice to use for UV irradiation, since you can control the intensity (=dose) and duration and will place a very intense UV source very close to the cells. I have heard of others using a standard UV trans-illuminator used for gels, but I'd be reluctant to go that way if you're trying to administer a controlled dose.
I'll keep that in mind and will try to find a cross linker.
Today I put the petri dish under the same laminar flow, but I tried to put it closer to the bulb, like 20cm away. I also removed the cap so UV could directly hit the cells. cells died after 45 minutes by apoptosis. I could clearly see membrane blebbing. and I think I put them so much under UV. next time I need to reduce the exposure and time. I need the apoptosis just before membrane blebbing.