Cellular extracts were prepared using lysis buffer containing 50 mM Tris HCL (pH 7.5), 250 mM NaCl, 1% NP-40, 0.5% Na-deoxycholate, 0.1% SDS, and EDTA free protease inhibitor (Roche 11873580001). Extracts were run on a 4–12% Tris-Glycine gel (BioRad) and transferred onto PVDF membranes. Blots were blocked in 5% milk PBS-T for 1 h at room temperature followed by overnight incubation at 4 °C with primary antibodies at 1:1,000 (anti-HA Tag, Cell Signaling 3724S; anti-Flag Tag, Sigma F1804; anti-DDX5, Bethyl A300-523A; anti-DDX17, Bethyl A300-509A). Horseradish peroxidase (HRP) -conjugated secondary antibodies were used at 1:10,000 (anti-rabbit HRP, Santa Cruz sc-2030) or 1:1,000 (anti-mouse HRP, Cell Signaling 7076S).
Sc 2030
Manufactured by Cell Signaling Technology
Sc-2030 is a magnetic rack designed for use with Cell Signaling Technology's Sepharose beads. It is used to separate magnetic beads from liquid solutions during immunoprecipitation and other bead-based assays.
Automatically generated - may contain errors
2 protocols using sc 2030
1
Western Blot Analysis of Protein Expression
2
Protein Extraction and Western Blot Analysis
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Example 15
Cellular extracts were prepared using lysis buffer containing 50 mM Tris HCL (pH 7.5), 250 mM NaCl, 1% NP-40, 0.5% Na-deoxycholate, 0.1% SDS, and EDTA free protease inhibitor (Roche 11873580001). Extracts were run on a 4-12% Tris-Glycine gel (BioRad) and transferred onto PVDF membranes. Blots were blocked in 5% milk PBS-T for 1 hour at room temperature followed by overnight incubation at 4° C. with primary antibodies at 1:1000 (anti-HA Tag, Cell Signaling 3724S; anti-Flag Tag, Sigma F1804; anti-DDX5, Bethyl A300-523A; anti-DDX17, Bethyl A300-509A). HRP-conjugated secondary antibodies were used at 1:10,000 (anti-rabbit HRP, Santa Cruz sc-2030) or 1:1000 (anti-mouse HRP, Cell Signaling 7076S).
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