Hi, I have some problems to measure calcium influx in sorted ILC2s from gut compartment (BL6 mice). Here our protocol :

- Deprivation of the cells overnight

- wash the cells with HBSS media without calcium

- staining with Fluo4AM 37°C 45 min (5uM)

- wash with HBSS with calcium and analyze

I did two different experiments on sorted ILC2s and I was not able to see staining by micrsocpy or FACS. The cells are alive but can not be stainined. In the same experiment we tested splenocytes and ILC3s sorted in the same time and the staining works perfectly (by this way, they can answer to ionomycin). Does anyone have experiences with that ?

Thanks a lot for your help

Best

Julie