![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=1724231107&Signature=o~dD3kw~Sw2mo2OS9a2XLWDEpBcmG~8ZQlW8R8l-NcUbLfqbH90syWz0dvxfVlhDggaR1~F0a0gjvTBwPiHBfz5GdY46oTUQ64W2mQQMpcpNCvfPHziq5rhiCC9MzhODvYwSoWIGnzqhwudnWMhF~1CEg~u0qbzuSoYo958TugYoal0lB2L9sjgKr8dw9TYRhaM8pHh8pV5uQpu5i98YPaW-aD2BAmPV6ThVOv85HX-UrzKyokyJHpsPzR8p9ILtTzUiQAf9OZNWtyMkkHKypNp2hh0a82CJyGNgGI0NyKKPZo6VozjsHMPtdvvOZv~3RcvXxxfTdilO6gHZS1lU8w__&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.
