Analyzing Protein Expression in HEK-293T Cells
Corresponding Organization : Cleveland Clinic
Other organizations : Amity University, National Cancer Institute, University of Massachusetts Chan Medical School
Variable analysis
- Transfection duration: 24 h or 36 h
- Protein expression/abundance of AKT, phospho-AKT (S-473), Myc, HA, K48, and GST
- HEK-293T cell line
- Radioimmunoprecipitation assay lysis buffer composition (1% NP-40, 20 mM Tris-Cl, pH 7.5, 150 mM NaCl, 1 mM Na2EDTA, 1 mM EGTA, 1% sodium deoxycholate, and 1 mM Na3VO4)
- Protein estimation using BCA Protein Assay Kit
- SDS-PAGE and Western blotting procedure
- Blocking with 5% nonfat dry milk or 5% bovine serum albumin
- Primary and secondary antibodies used
- Positive control: Not explicitly mentioned
- Negative control: Not explicitly mentioned
Annotations
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