PBM is essential for outgrowth of CTLL-2/Tax cells in the absence of IL-2. (A, B) CTLL-2 cells (107) were transfected either with the vector plasmid (pHβPr-1-neo) or with expression plasmids encoding Tax1, Tax2B or their mutants by electroporation. The cells were divided into two groups 24 h after transfection. From the first group, living cells were collected using Ficoll-Paque Plus (Amersham Biosciences) and used for Western blot analysis (A) using anti-Tax1 antibody (TAXY7) or anti-Tax2B polyclonal antibody . The second group (B) was seeded into 96 well plates and cultured in RPMI-FBS without IL-2 for 3 weeks, and the number of IL-2-independent colonies was counted under light microscopy. The percentage of positive wells indicates the proportion of the wells containing outgrowth of CTLL-2 cells. The data relates to two independent experiments with each duplicated transfection.