Trichloroacetic acid

Protein isolation Mammalian cells - Human gingival epithelial cells

Experiment
Protein isolation Mammalian cells - Human gingival epithelial cells
Product
Trichloroacetic acid from Sigma-Aldrich
Manufacturer
Sigma-Aldrich

Protocol tips

Protocol tips
- The supernatants are precipitated with 10% trichloroacetic acid for concentration by desalinization, washed with acetone, and then dried at room temperature. The dried pellets and the remaining cells are suspended in a reducing sample buffer containing 2-mercaptoethanol.

Publication protocol

The supernatants were precipitated with 10% trichloroacetic acid (Sigma) for concentration by desalinization, washed with acetone, and then dried at room temperature. The dried pellets and the remaining cells were suspended in a reducing sample buffer containing 2-mercaptoethanol. Samples were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Separated proteins were electrotransferred to polyvinylidene difluoride membranes (Millipore, Bedford, MA) with a semidry electroblotter (Bio-Rad, Richmond, CA). Membranes were blocked with 1% (wt/vol) skimmed milk in Tris-buffered saline (TBS, pH 7.5) containing 0.05% Tween 20 (TBS-T) and incubated with mouse anti-IL-1β antibody (Cell Signaling Technology, Beverly, MA) or mouse anti-α-tubulin antibody (Cedarlane, Ontario, Canada) in Immuno-enhancer reagent A (Wako Pure Chemical Industries) overnight at 4°C. After being washed with TBS-T, membranes were incubated with a horseradish peroxidase-conjugated goat anti-mouse IgG antibody (Sigma) in Immuno-enhancer reagent B for 1 h at room temperature. Labeled proteins were visualized with ImmunoStar LD Western blotting substrate (Wako Pure Chemical Industries) or Western blotting substrate (Thermo Scientific, Waltham, MA).

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Manufacturer protocol

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