Gal-1 antagonized ERK signaling in thymocytes undergoing positive selection. (A) Histogram of phospho-ERK (pERK) staining in DP thymocytes from wild-type (thick line) and gal-1^−/− (thin line) OT-1 mice, and isotype control (shaded). Cells were immediately fixed and permeabilized after dissociation from thymic stroma to preserve positive selecting signals induced in vivo. (B) The average MFI (± SD) of pERK in DP thymocytes from 3 wild-type (□) and 4 gal-1^−/− (▧) OT-1 TCR mice. Results represent 2 independent experiments. (C) The average of fold over control (± SD) of pERK staining in wild-type (□) and gal-1^−/− (▧) DP thymocytes from C57Bl/6 (wild-type, n = 4; gal-1^−/−, n = 4) and OT-1 TCR transgenic mice (wild-type, n = 4; gal-1^−/−, n = 5). For panels B and C, 2-tailed unpaired Student t tests were performed to determine statistical significance. P values are listed in the panels.