A small pool of SLP-2 is associated with the plasma membrane.

<p>A) Intact parental Jurkat T cells, as well as Jurkat T cells stably transfected with full length SLP-2-gfp or ΔN-SLP-2-gfp were biotinylated. Next, cells were lysed and surface proteins were immunoprecipitated with an antibody against biotin. The immunoprecipitate samples (Biotin ip) were immunoblotted for SLP-2 to detect the pool of SLP-2 associated with surface molecules. Whole cell lysates (WCL) from the transfectants were blotted to show expression levels of the transgenes. Samples were also blotted for caspase 3 as a negative control for surface protein pull-down. B) Increasing levels of SLP-2-gfp or ΔN-SLP-2-gfp were induced with increasing concentrations of doxycycline and biotin immunoprecipitation was performed. Whole cell lysates (WCL) and biotin immunoprecipitates (biotin ip) were blotted as in A. C) T cells isolated from wild type and SLP-2 T-K/O mice were analyzed by biotin immunoprecipitation and blotted as in A. Samples were also blotted for actin as a loading control for SLP-2 T-K/O samples and also for GAPDH as a negative control for surface protein pull-down. These results are representative of at least 3 independent experiments.