Dear all,
I am using IL7Ra antibody (cllone A7R34) PEcy7 conjugated to stain for ILCs in mouse spleen but, I am not getting well resolved population of IL7Ra positive cells. I want to improve resoultion of this population. Do anybody have any Idea how to do this? Any suggestion about antibodies and flourochrome?
Hi Arshed, have you tried doing an enrichment for TCRb/CD3/CD5 negative, CD19 negative cells prior to staining?
Hi Susanna,
No i am not doing enrichment but, I think it would be nice idea to deplete (as much as possible) my samples with lineage positive cells. Thanks for your idea.
HI Arshed,
You're most welcome. We struggled a bit with column based enrichment as we were using the whole spleen, and removing bulk of the cells. So we are currently using a column free method.
Also, removing T and B cells using CD3, CD5, and CD19 was sufficient in our case. Notably, even when we use a bright fluorophore for IL7Ra (clone A7R34), the separation between positive and negative is not fantastic. But you can see enough of a distinction between negative and positive to gate.
Let me know if you'd like an example plot.
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