Immunoblotting of Protein Signaling Pathways

Cell extracts were prepared as described previously [43 (link)]. Briefly, lyophilized proteins were solubilized in 1X Laemmli sample buffer and boiled for 10 min. Protein concentrations were determined with the Bio-Rad Bradford Protein Assay Kit (Bio-Rad, Marnes-la-Coquette, France) before immunoblotting. The following antibodies were purchased from Cell Signaling Technologies (Ozyme, Montigny-le Bretonneux, France): AXL (8661), ATM (2873), phospho-ATM (Ser1981)–(4526), ATR (2790), phospho-ATR (Ser428)–(2853), Chk1 (2360), phospho-Chk1 (Ser345)–(2348), Chk2 (2662), phospho-Chk2 (Thr68)–(2197), DNA-PKc (4602), MERTK (4319), TYRO3 (5585). Antibody against phospho-DNA PKc (Ser2056) (ab18192) was purchased from Abcam (Paris, France).

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Silina L., Dufour F., Rapinat A., Reyes C., Gentien D., Maksut F., Radvanyi F., Verrelle P., Bernard-Pierrot I, & Mégnin-Chanet F. (2022). Tyro3 Targeting as a Radiosensitizing Strategy in Bladder Cancer through Cell Cycle Dysregulation. International Journal of Molecular Sciences, 23(15), 8671.

Publication 2022

Bio-Rad Bradford Protein Assay Kit

Antibodies Antibody Assay Cell extracts Laemmli buffer Mertk Protein

Corresponding Organization : Université Paris-Saclay

Other organizations : Université Paris Sciences et Lettres, Université Clermont Auvergne

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Variable analysis

independent variables

Lyophilized proteins were solubilized in 1X Laemmli sample buffer and boiled for 10 min.

dependent variables

Protein concentrations were determined with the Bio-Rad Bradford Protein Assay Kit.

control variables

Cell extracts were prepared as described previously [43].

controls

Positive control: Not explicitly mentioned.
Negative control: Not explicitly mentioned.

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