Some of you may be doing intracellular staining for cytokines. I am particularly interested in staining mouse lymphocytes under conditions as close to ex vivo as possible. I am comparing cytokine responses of normal and infected mice. My experience with PMA and ionomycin plus brefeldin A was that relatively higher frequencies of cytokine-producing cells could be obtained, but the difference between normal and infected mice seemed obscured. I tried Ag and anti-CD28 plus brefeldin A as in vitro activation, the frequency of positively stained cells was below 1% and such a number was hard to reproduce. Wish to hear from you, and we can discuss more details. Thanks in advance. Hai Qi Dept. of Pathology, UTMB
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