Hey guys,
I am trying to stain eosinophily for intracellular proteins.
First step ist to stain them with fixable viability dye, after that fixate and permeabilise them to stain for intracellular proteins afterwards.
My problem is, that they become positive for all the stains, including the viability dye, eventhough I tested the viabilty with other stains, and the results showed good viability.
Has anybody encountered problems with that viability dye and eosinophils, or can recommend an alternative viability dye for intracellulare facsing of eosinophils?
Thank you very much!
Thanks for your quick reply.
Yes I do. I use the same protocoll for other cell types as well, and never had any problems
I use eBioscience™ Fixable Viability Dye eFluor™ 506 from Thermofischer.
Well, I never had problems with autofluorescence before, treating other cells. Thats why I am now in need of help =)
Kind of solved the problem now. The Permeabilization Buffer induced the autofluorescence. Thank you Tamara for your ideas!
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