Differential Protein Profiling by 2D-Electrophoresis

Two-dimensionl electrophoresis is a powerful tool to analyze differential protein expression levels with or without 250 μg/mL SBE treatment according to previously described procedures [50 (link)] which have been both high sensitivity and efficacy. The cell lysate (180 μg) was applied to 18-cm IPG, pH 4–7 linear strip (GE Healthcare, Uppsala, Sweden) and separated on the IPGphor III System for the first dimension. The 2-DE was carried out on 10% acrylamide gels (Bio-Rad, Hercules, CA, USA) and the gels were visualized by silver staining. Protein spots were quantified using the Prodigy SameSpots software (Nonlinear Dynamics version 1, Newcastle, UK). More than 1.5-fold alterations at 95% confidence interval (p < 0.05) were considered statistically significant. All experiments were repeated three times to confirm reproducibility.

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Wu T.H., Lin T.Y., Yang P.M., Li W.T., Yeh C.T, & Pan T.L. (2024). Scutellaria baicalensis Induces Cell Apoptosis and Elicits Mesenchymal–Epithelial Transition to Alleviate Metastatic Hepatocellular Carcinoma via Modulating HSP90β. International Journal of Molecular Sciences, 25(5), 3073.

Publication 2024

IPGphor III System 10% acrylamide gels

Corresponding Organization :

Other organizations : Kaohsiung Veterans General Hospital, Keelung Chang Gung Memorial Hospital, Taipei Medical University, National Research Institute of Chinese Medicine, Ministry of Health and Welfare, Chang Gung Memorial Hospital, Chang Gung University of Science and Technology, Chang Gung University

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Variable analysis

independent variables

250 μg/mL SBE treatment

dependent variables

Differential protein expression levels

control variables

Cell lysate (180 μg) applied to 18-cm IPG, pH 4–7 linear strip
Separation on the IPGphor III System for the first dimension
2-DE carried out on 10% acrylamide gels
Gels visualized by silver staining
Protein spots quantified using the Prodigy SameSpots software
More than 1.5-fold alterations at 95% confidence interval (p < 0.05) considered statistically significant
Experiments repeated three times to confirm reproducibility

controls

No positive or negative controls were explicitly mentioned in the input protocol.

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