Fluorescence imaging of [Ca²⁺]_i responses from individual cells after CD3, CD148, and CD3+CD148 crosslinking. Cells previously loaded with fura-2 (2 μmol/L), were incubated with the corresponding antibody (CD3, 10 μg/mL; 14341, 40 μg/mL; CD45, 40 μg/mL) for 30 minutes at 4°C. After washing, cells were plated on Cell-Tak–coated glass coverslips. After establishing baseline values, saturating amount of polyclonal GAM was added to prewarmed samples. [Ca²⁺]_i was measured every 5 seconds for 15 minutes in every cell. Calcium changes are expressed as the percentile change of basal. A total of 20 cells representative of each condition are shown. Each line represents an individual cell.