An interesting result to share.
In one of our projects, cancer cells exposed to cell targeting phage show quite elevated alamar blue activity without any change in cell proliferation after 24 and 48hr culture. Alamar blue is used as a measure of mitochondrial activity so it may be that the exposure is stimulating mitochondrial production or at least overexpression of the relevant mitochondrial reductases..
Anyone have some other ideas?
Thanks,
Michael Firer
We are using alamarBlue quite a bit but this is certainly one of its shortcomings. How do you distinguish between 100% of cells having a metabolic depression of 50% from killing 50% of cells and the survivors are doing fine? Or as in your case when alamarBlue indicates that the surviving cells perform actually ‘better’. At the end of the day you need to do the more time consuming cell counts. However, I think you got an interesting finding by combining AB data with cell counts. In case you’re increasing the number of mitochondria per cell you should be able to test this using Western Blotting for one of the house keeper proteins (VDAC, cyt-c, etc.). In case you are increasing respiratory chain activity you probably would get some nice insights by doing respirometry on permeabilized cells or isolated mitochondria. However, since many cell lines (and cancers) are quite glycolytic poised you may actually are making them more ‘primary’-like. It would be quite interesting to know if your treatment shifts carbon flux from glycolysis to oxidative phosphorylation.
In addition to proposed experiments you may label the cells with JC-1 and 7-AAD to monitor the changes in live cell mitochondrial activity and apoptosis/necrosis events by Flow cytometry. Also, there are commercially available kits to measure the activity of mitochondrial complex I, IV etc. in cellular lysates.
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