I wanted to detect intracellular cytokines by flow cytometry, but found that the levels of CD107a and Granzyme B after resting overnight were significantly higher than when the cells were resuscated (the proportion of these two proteins in CD8 can reach 80-90%), and no stimulants were added during the period. Does anyone know why?
The medium I used was RPMI1640, 10%FBS, 100IU/ml IL-2, is it possible that IL-2 caused the elevated levels of these two proteins?
Dear Liwei Chang
I briefly wanted to reach out that I have (sometimes) observed a similar phenotype as yours, which was the reason, why I started skipping any resting periods for cocultures. For details on the protocol please see:
Article A role for the histone H2A deubiquitinase MYSM1 in maintenan...
The best explanation (and possibly a starting point for experiments) has been published by Karl Johan Malmberg's group in Oslo showing that IL2 family cytokines like IL15 and IL21 drive a reorganization of secretory lysosomes increasing the intracellular distribution of granzyme b/ CD107a in NK cells. Without knowing the details of your experiment and/or downstream application I would assume that something similar is happening to your CD8 cells given that all these cytokines signal through the common gamma chain or share certain subunits like the IL2Rb chain.
For details on their experiments, please check the following paper:
Article Remodeling of secretory lysosomes during education tunes fun...
I hope that helps.
All the best & kind regards,
Michael
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