Protein Detection and Quantification Assay

The Western analysis assays for p53, Vinculin, and GAPDH detection were performed as previously described (18 (link)). Briefly, mixtures of 1 mg ml⁻¹ cell lysates and the fluorescent master mix were heated at 95 °C for 5 min. The samples and all other reagents were dispensed into the microplates and capillary electrophoresis Western analysis was carried out with the ProteinSimple WES instrument and analyzed using the inbuilt Compass software (ProteinSimple). DO-1 mouse anti-p53 antibody (1:400, Santa Cruz sc-126), mouse anti-vinculin antibody (1:600, R&D Systems MAB6896), and rabbit anti-GAPDH (1:800, Abcam ab128915) were used.

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Baradaran-Heravi A., Bauer C.C., Pickles I.B., Hosseini-Farahabadi S., Balgi A.D., Choi K., Linley D.M., Beech D.J., Roberge M, & Bon R.S. (2022). Nonselective TRPC channel inhibition and suppression of aminoglycoside-induced premature termination codon readthrough by the small molecule AC1903. The Journal of Biological Chemistry, 298(2), 101546.

Publication 2022

ab128915

Anti antibody Assays Capillary electrophoresis Cell Gapdh Mouse Rabbit Vinculin

Corresponding Organization : Institute of Structural and Molecular Biology

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Levels of p53, Vinculin, and GAPDH proteins

control variables

Cell lysate concentration (1 mg ml^-1)
Heating temperature (95 °C)
Heating duration (5 min)
Antibody dilutions (p53: 1:400, Vinculin: 1:600, GAPDH: 1:800)

controls

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

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