Are you blocking with Monensin or Brefeldin A during the last few hours of the stimulation? You might find the following protocol for intracellular staining of mouse & human cytokines useful: http://www.ebioscience.com/ebioscience/appls/FCI.htm. Best regards, Lily eBioscience http://www.ebioscience.com ----- Original Message ----- From: Martha Moreno-Lafont To: Cytometry Mailing List Sent: Tuesday, June 19, 2001 5:05 PM Subject: IFN-g detection Dear flowers, we are trying to detect intracellular IFN-g in murine spleen cells, but we have problems with positive control. We activated 8 x 10E6 spleen cells with 10 and 20 ug PHA, and with 6 ug PMA + 4 ug Ionomicin for 4 hours but results were no IFN-g. We used 0.2 ug anti IFN-g-PE from Pharmingen. In our lab., we activated human whole blood with 10 ug PHA for 4 h and intracellular IFN-g were detected. Any of you have experience in murine cells activation and IFN-g detection? Thanks in advance, Martha Dra. Martha Moreno-Lafont Depto. de Inmunología Esc. Nal. de Ciencias Biológicas - IPN México D.F., México Tel: (525) 729 6300 ext. 62368 Fax: (525) 729 6300 ext. 62489
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