Currently, I'm trying to isolate a cells that produce a protein that localized near the cells membrane. This is mix cells from tissue. I had stained the cells with primary antibody+FITC conjugated secondary antibody. I detect the cells using BD accuri, with FL1 (FITC filter) and SSC. When the cells were cultured in different supplement, some showed growth, let say 10K to 40K after 3 days. Another treatment can go up to 1000K, mostly in area close to the the machine noise with no fluorescent signal. None detected with the control. How can I rationalize the results when compared to control? Adding PI did not differentiate between dead cells and debris.
Hi Shann Yu, thank you for the advice. We had tried with FDA to stain the cells, it did not show a very good signal. Similar to PI. We had use Presto blue instead but it is not consistent as time of incubation increase. We never tried with calcein AM, thus will take note on it. Nevertheless, having debris with the cells had caused the cells to be masked and due to mix cells, we had wide distribution of cells.
Thank you,
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