Hi everyone,
I am experiencing an issue with my THP-1 cells. After addition of 100 ng/ml PMA to differentiate into macrophage-like M1 cells, the THP-1s are not fully adhering down. There is a noticeable number of cells still in suspension/loosely-adhered multiple days after PMA addition, and the cells that are adhered down don't look quite as healthy and differentiated as they normally do.
I have screened for mycoplasma which was negative, and have changed to fresh media and fresh FBS. I have been working with these cells for over a year and haven't experienced this before - nor have any of my colleagues.
Does anyone have any idea what is happening?
Dear Richard:
I hope that you already solved your problem. Cells should be adhered at the bottom of the well 24 hours after the addition, even at low concentrations, although it´s never 100%. In my personal case, I had a lot of problems with the handling. Those cells are alive and if you mechanically stress them enough, they will not attach. If your problem persist, you should consider that someone else with more experience at the lab should handle your cells (at least until your cells adhere with PMA).
Also, I suggest you to read this little threat;
https://www.researchgate.net/post/PMA-treated_THP-1_cells_detach_after_washing_with_PBS
I forgot to mention. There is the possibility that one of your solutions or reagents is too old or maybe contaminated with "something".
- Did you decomplemented your FBS as usual?
- Did you change the lot of your medium and/or PMA?
- Are you sure that your medium and/or PMA did not get too old or it got oxidized by opening and closing to much the same vessels by not putting them in aliquots?
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