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. 1997 Mar 3;185(5):825–832. doi: 10.1084/jem.185.5.825

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IL-12–dependent signaling in human Th1 and Th2 cell lines and clones. IL-12 induces Stat4 phosphorylation in human Th1 but not Th2 cells. Th1 and Th2 lines generated from cord blood lymphocytes as in Fig. 1 A were harvested on day 10 after priming. Th1 (CB13) and Th2 (CB21) clones were harvested on day 13 after restimulation. 5 × 10⁶ cells were washed and incubated 15 min at 37°C in medium with or without 2 ng/ml IL-12 followed by the preparation of whole cell extracts and immunoprecipitation with anti-Stat4 antiserum. Precipitated proteins were separated by SDS-PAGE (8%), transferred to nitrocellulose, and probed with anti-phosphotyrosine antibody 4G10 (anti-P-Y, top). As a control for Stat4 expression, blots were stripped and reprobed with anti-Stat4 antibodies (bottom).