I am performing microgravity experiments on subpopulations of purified T cells. My control gorup will sit in the incubator for 24 hours and I continue to find small aggregates at the bottom of my eppendorf tubes. I am curious as to whether or not these are cells that were not completely purified. I performed my purification step by negative selection.
Does anyone have any experience with this and should I use another medium other than RPMI?
Purified T cell subpopulations (exp with CD4 T cells) form aggregates. You could try PBS.
Yes, T cells will form aggregates, especially when stimulated. It doesn't matter which medium you use, it has to do with the activity of the T cells. Cell to cell contact is just one of the many variables that microgravity culture conditions (I'm assuming you are using the vertical rotator, I forgot what those were called) disrupt.
Now, if you really want to see what would happen with the aggreated cells in the microgravity culture. You would have to let the cells aggregate and then resuspend them in the rotattion esselv. However, the aggregated cells will require you to readjust the rotation speed... just a little, probably not by much, to keep them in suspension.
I missed those experiments... Very frustrating sometimes.
Purified T cells will also form aggregates. And negative selection is a better way for T cell purification compared to positive selection (FACS based purification is much better). This aggregation will probably form in all kinds of media. You may re-suspend them to avoid aggregation.