Conjugates were formed between DO11.10 naive and memory CD4 T cells, and APCs pulsed with SEB. At the indicated times, the conjugates were assessed for the presence of the TCR, ZAP-70 and pZAP-70. The two far-right columns in both panels show fluorescence overlays and the DIC images of the conjugates, respectively. Beneath the x-y images is a three-dimensional reconstruction along the x-z axis of a mature immune synapse. Data are representative of 3 separate experiments, examining a minimum of 50 conjugates per experiment.