Hi all,
is there a way to analyze cell proliferation with CFSE with BD DIVA software? I activate murine splenocytes or purified CD4 and I use Cell Trace and follow the exact protocol but I only see the initial peak on Day 0 as expected. I harvest the cells each day from day 1 to 5 but instead of seeing multiple peaks, I rather see a "curve" each day, almost falling at the same place of intensity. Any ideas?? Should I only harevst the cells on day 4 or 5 (final)?
Hi Spyridoula,
You can try titrating down the CFSE to optimize the peaks. You should also gate on CD4+ live cells. Other advice I have is to:
1) look at D0 AND d4 or d5 only, not d1,2,3
2) change your collection size. For example, for D0 (or unstimulated), collect 5k CD4+ events but for d4 (or 5) collect 50-80K events so that the maxima of the 2 curves are about the same height. If you look closely, you will notice that there are humps starting to appear, you will see them resolve better with titrating down the cfse and by collecting many more events
Kind regards,
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