As previously described [14 (link)], WB analysis was used to detect protein expression. RIPA Lysis Buffer (Beyotime) was used to extract protein. The proteins were resolved by SDS-PAGE gel and transferred to PVDF membrane. The membrane was incubated with anti-PDK1 (1:2,000, Sigma-Aldrich) or anti-β-actin (1:1,000, Sigma-Aldrich), and then incubated with goat anti-mouse IgG (1:20,000, Biovision). Using the ECL luminescent solution (Meilunbio), the protein signals were visualized.
Protocol detail
Western Blot Protein Detection
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Corresponding Organization : Guangxi Medical University
Other organizations : Shandong First Medical University, Jinan City People's Hospital, Yunnan Provincial Hospital of Traditional Chinese Medicine, Liaocheng People's Hospital
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Variable analysis
independent variables
- Anti-PDK1 antibody concentration (1:2,000)
- Anti-β-actin antibody concentration (1:1,000)
- Goat anti-mouse IgG antibody concentration (1:20,000)
- Protein expression
- RIPA Lysis Buffer (Beyotime) for protein extraction
- SDS-PAGE gel for protein resolution
- PVDF membrane for protein transfer
- ECL luminescent solution (Meilunbio) for protein signal visualization
- Positive control: Not explicitly mentioned
- Negative control: Not explicitly mentioned
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