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. 2011 Nov 14;108(48):19323–19328. doi: 10.1073/pnas.1108052108

Fig. 3.

Fig. 3.

Mutation of BRS motifs abrogates early TCR signaling. TCRζ-deficient BW5δ (A) or 2B4 (B) cells expressing the indicated TCRζ constructs were analyzed before or after incubation with pervanadate (10 μM) at 37 oC. Immunoprecipitation was performed with anti-CD2 (OX34)–coated beads and blotted with anti-pY (4G10) antibody. Blots were stripped and reprobed with OX34. (A) Total cell lysate was used to measure pZap70 (pY319) and pLAT (pY119) and then was stripped and reprobed for total Zap70 and total LAT, respectively. (B) Total cell lysate was blotted with anti-pY (4G10) antibody and rabbit polyclonal anti-TCRζ. (C) BW5δ cells expressing matching levels of the indicated constructs were stimulated by plate-bound anti-CD2 (OX54) for 1 min. These cells then were fixed, permeabilized, and stained with a directly conjugated anti-pZap70 (pY319) antibody.