Hello everyone,
I'm currently analyzing expression of Granzyme B by T cells (CD4, CD8 and gamma delta) and NK cells and I found out that CD8, gamma delta T cells and NK cells express granzyme B without being stimulated. Does it seem correct to you? I feel like I shouldn't have as much as granzyme B positive cells.
As an example, I get about 30% of Granzyme B+ CD8 T cells without stimulation (percentages are greatly increased for gamma delta T cells and NK cells).
Here is my protocol:
Human PBMCs were thawed and rested 4 hours at 37C. Then PBMCs were stimulated (or not as negative control) for 20 hours at 37C and PBMCs were stained and analyzed by flow cytometry.
I tried with/without brefeldin A, I analyzed the cells just after thawing, after 4 hours of resting and after 20 hours of stimulation and I always get the same data.
Has anyone ever got similar data? Am I missing anything in my protocol? Does anyone has any tips?
Hi Emilie Picard!
Yes, that's totally normal. I usually get around 30% in T cells, and in NK cells above 80%. Actually, upon stimulation, both T and NK cells degranulate and the intracellular levels of granzyme B should be lower than in resting cells.
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