I am trying to expand T cells, but looks clustering seriously, lots of black spheroids, tried to break them down by pipets, but not so helpful, this made cell counting not reliable, so I can not adjust density accurately, without accurate counting, also affect my next experiments. Thanks.
That's...unusual... They shouldn't...
If the clustered cells are black... they are probably dead, or it's contamination.
Maybe post a picture... and this is after isolation or after expansion?
T cell clustering is normal because T cells are suspend cells ( not adherent) and form clusters. shake or pipets your suspension.
Thanks guys. After monitoring days and adjusting microscope focusing, now I think it's normal clustering (instead of contamination, that's good news).
Based on focusing, it can be black or bright, when bright, it looks like
fig.1 from
https://www.stemcell.com/immunocult-human-cd3-cd28-t-cell-activator.html
Figure 1. Activated Morphology of Human T Cells Stimulated With ImmunoCult™ Human CD3/CD28 T Cell Activator
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