Hi, I am trying to set up an antibody dependent cellular cytotoxicity assay using the well known calcein-AM. Cell loading is fine however I observe around 50% spontaneous release. 1.) Are there any parameters I have to change in order to obtain less spontaneous release? (for ex. Amount of cells, loading time, dye concentration) 2,) I am aware that lymphocytes can be separated from monocytes by plastic adherence. Could someone give me a nice protocoll for this? How many mononuclear cell should I put per cm^2 ? Or, where could I obtain more info on this protocol? I cannot find it because it is to "standard". Thanx joel
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