Defective T-cell activation in ΔSAM–SLP-76 knockin mice. (A) Splenocytes from SLP-76^m/+ and SLP-76^m/m littermates were either left untreated (gray area), stimulated with 3 μg/mL plate-bound anti-CD3 (solid line), or stimulated with 20 ng/mL PMA plus 0.5 μg/mL ionomycin (dotted line) for 6 hours and subsequently analyzed by flow cytometry. Surface expression of CD25 and CD69 on CD4⁺ gated cells is shown. (B) Purified SLP-76^m/+ and SLP-76^m/m CD4⁺CD44^low T cells were either stimulated with various concentrations of plate-bound anti-CD3 plus 1 μg/mL soluble anti-CD28 (left), or with PMA plus ionomycin (right). After 36 hours, cells were pulsed with [³H]thymidine for an additional 6 hours before being harvested for scintillation counting. Cell proliferation is represented by the counted radioactivity (CPM). (C) Purified SLP-76^m/+ and SLP-76^m/m CD4⁺ T cells were stimulated the same as in panel B. Supernatants were harvested after 8 hours, and the IL-2 concentrations were determined by ELISA.