RACK1 antibody | knockout validation | Cell Signaling 5432

This is a knockout-validated antibody summary, based on the publication "Trans-kingdom mimicry underlies ribosome customization by a poxvirus kinase", as cited below [1]. Labome curates formal publications to compile a list of antibodies with unambiguous specificity within Validated Antibody Database (VAD).

Antibody information

Rabbit monoclonal IgG

Company: Cell Signaling

Antibody: RACK1

Catalog number: 5432

Summary: Rabbit monoclonal IgG against a synthetic peptide corresponding to central residues of human RACK1 protein. Reacts with human, mouse, rat, monkey, and D. melanogaster. Suitable for western blot and immunoprecipitation.

Validation Method

Western blot

Sample

Wild type or RACK1-knockouts (KO1, KO2). Whole-cell lysates were prepared in lysis buffer (62.5mM Tris-HCl at pH 6.8, 2% SDS, 10% glycerol, 0.7M β-mercaptoethanol).

Blocking agent

5% non-fat dry milk, 0.1% Tween in TBS for 1h at room temperature.

Primary incubation

Diluted in 3% BSA, 0.1% Tween in TBS overnight at 4°C.

Secondary incubation

1:3,000 dilution HRP-conjugated secondary antibody (GE Healthcare Life Sciences) in 5% non-fat dry milk, 0.1% tween in TBS for 1h at room temperature.

Detection

Pierce ECL Western Blotting Substrate (Thermo Fisher Scientific) for 1min before exposure to X-ray film.

Figure

Western blot analysis of samples from wild type or RACK1-knockouts (KO1, KO2). Please see Figure 1b in the article [1].

References
  1. Jha S, Rollins M, Fuchs G, Procter D, HALL E, Cozzolino K, et al. Trans-kingdom mimicry underlies ribosome customization by a poxvirus kinase. Nature. 2017;546:651-655 pubmed publisher