American Society of Hematology

Fig. 3.

$Fig. 3. Analysis of tyrosine phosphorylated proteins in thymocytes in response to MIP-1β. (A) Freshly-isolated unfractionated thymocytes were kept on ice before stimulation wth 10 nmol/L and 100 nmol/L MIP-1β, or anti-CD3 as a control, for 3 minutes at 37°C. After stimulation, the lysates were prepared as described in the Materials and Methods section. Electrophoresis under reducing conditions was performed using 12% tris-glycine gels and Western blots probed with antiphosphotyrosine (clone 4G10). (B) Thymocytes were purified and stimulated with anti-CD3 (10 mg) or 100 nmol/L MIP-1β as outlined in the Materials and Methods section. Electrophoresis under reducing condition was performed using 10% tris-glycine gels and Western blots probed with antiactive MAPK. Blots were stripped and reprobed with anti-ERK2 (lower panel) to show equivalent loading.$

Analysis of tyrosine phosphorylated proteins in thymocytes in response to MIP-1β. (A) Freshly-isolated unfractionated thymocytes were kept on ice before stimulation wth 10 nmol/L and 100 nmol/L MIP-1β, or anti-CD3 as a control, for 3 minutes at 37°C. After stimulation, the lysates were prepared as described in the Materials and Methods section. Electrophoresis under reducing conditions was performed using 12% tris-glycine gels and Western blots probed with antiphosphotyrosine (clone 4G10). (B) Thymocytes were purified and stimulated with anti-CD3 (10 mg) or 100 nmol/L MIP-1β as outlined in the Materials and Methods section. Electrophoresis under reducing condition was performed using 10% tris-glycine gels and Western blots probed with antiactive MAPK. Blots were stripped and reprobed with anti-ERK2 (lower panel) to show equivalent loading.

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