![Fig. 4. Immune complex kinase activity of TM4SF-associated and total c-kit fractions in the absence or the presence of SLF. / MO7e cells were stimulated with or without 50 ng/mL SLF at 37°C for 10 minutes. Cells were solubilized with lysis buffer containing 1% CHAPS. The cell lysates were immunoprecipitated with indicated first antibodies. Immunoprecipitated materials were eluted with 1% NP-40 lysis buffer and supernatants were immunoprecipitated with anti–c-kit mAb (cloneYB5.B8). In the case of anti–c-kit mAb as a first immunoprecipitation antibody, immunoprecipitated proteins were treated with NP-40 in the same way. The immune complexes were incubated with 10 μCi (0.37 MBq) [32P-γ] adenosine triphosphate for 10 minutes at room temperature. Immune complexes were separated by SDS-PAGE and transferred onto Immobilon-P membrane. [32P] incorporation into c-kit protein was visualized by autoradiography (upper panel). The same membrane was immunoblotted with anti–c-kit polyclonal antibody (lower panel). These are the representative results from 2 separate experiments.](https://ash.silverchair-cdn.com/ash/content_public/journal/blood/99/12/10.1182_blood.v99.12.4413/4/h81222688004.jpeg?Expires=1735550883&Signature=2REhFyZrMbtF7oB-Ohpc2O2maDuyItkDCW6mnNpHjQq3x7cVlo7mTq1jxELzUqijwNNlVyqrcNq5k7hMsbed41oBBr~yjXEP1~WXMw~sO3JYnlq2BHetyDOLTwS06hB88cJjZDXGKNjMhPsBz6QCDt9eFPKNaSQRX5AOSCsFpCDDC~TPfwSb4Qa8XmWtbMeFt5XrMoeBiRqSkv~SCVR5kwy~lewfrITZmKvnK8PthqQpz5Cr1NtD4YzLwbFwMYl6AJXgDQYoWI7OwpM-yTEX2pt6ESJLAMydBbaJla3fp9~f0uUq6UsoI3q7CIAZCziqiMz9geXGDixmmi-2FvlZqw__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Immune complex kinase activity of TM4SF-associated and total c-kit fractions in the absence or the presence of SLF.
MO7e cells were stimulated with or without 50 ng/mL SLF at 37°C for 10 minutes. Cells were solubilized with lysis buffer containing 1% CHAPS. The cell lysates were immunoprecipitated with indicated first antibodies. Immunoprecipitated materials were eluted with 1% NP-40 lysis buffer and supernatants were immunoprecipitated with anti–c-kit mAb (cloneYB5.B8). In the case of anti–c-kit mAb as a first immunoprecipitation antibody, immunoprecipitated proteins were treated with NP-40 in the same way. The immune complexes were incubated with 10 μCi (0.37 MBq) [32P-γ] adenosine triphosphate for 10 minutes at room temperature. Immune complexes were separated by SDS-PAGE and transferred onto Immobilon-P membrane. [32P] incorporation into c-kit protein was visualized by autoradiography (upper panel). The same membrane was immunoblotted with anti–c-kit polyclonal antibody (lower panel). These are the representative results from 2 separate experiments.
