Fig. 5

Nck binds to human but not mouse CD28. a, b Human primary CD4⁺ T cells were stimulated for 5 min with B7-negative (−) or Dap3/B7 (B7) cells and anti-CD28 IP were performed on cellular extracts. Anti-Vav, anti-Nck, anti-p85 PI3K, and anti-CD28 western blottings were performed on anti-CD28 IP or total lysates (TL). Data represent one of three independent experiments. c, d CD4⁺ T cells isolated from murine spleen (n = 10) were pooled and stimulated for 5 min with with B7-negative (−) or B7.1-expressing RAW (B7) cells and anti-CD28 IP were performed on cellular extracts. Anti-Vav, anti-Nck, anti-p85 PI3K, and anti-CD28 western blottings were performed on anti-CD28 IP or total lysates (TL). The position of immunoprecipitated proteins as well as of immunoglobulin heavy chain (IgH) is indicated. e Equal amounts of total lysates from human or mouse CD4⁺ T cells were analysed by western blotting for Vav1 (upper panel) and Nck (lower panel) content. The position of molecular weight markers, expressed in kDa, is indicated on the right