U937 attachment on flask/well - (May/23/2007 )

i induce my U937 with 10ng/ml PMA for 48 hrs. after differentiation, my cells become attached. i proceed with my infection (with bact). during the washing step (using 1x PBS), i realised some of my cells become detached.
does U937 attach strongly or can be easily detach? what can we do to reduce detachment during washing and pipetting?
or is there any other way to wash away unadhered extracellular bacteria after infection which is milder to the cells?
Thank you!!!

I encountered the same situation, where the differentiated U937 were detached during washing step when i was doing virus infection. The U937 differentiation was done using PMA at concentration of 10nM for 48 hours at 37C, and the infection was performed once the PMA-containing media was removed. According to the literatures, the differentiated U937 should be able to attach well. While for my case, after differentiation, though the U937 cells were still attached in clumps but did not show spindle shape, and the cells are easily detached after washing. Is this due to the incomplete differentiation? I've tried increase PMA concentration and extend the differentiation time, but i get more dead cells instead. I found people replaced fresh media and let the cells grow for another few days after the differentiation, then only they proceed to their experiment. Does this help?


singsin

After your bacterial infection, do you leave the cells in culture or proceed to analyzed the cells.
If you leave them in culture it is difficult to wash the cells with PBS because they do detach fairly easily.
But if you are anaylzing the cells remove media/bacteria then collect the cells. The cells will spin down much more quickly under low rpm than bacteria and the cells can be separated from the bacteria in this manner.