Selected article for: "bromophenol blue and room temperature"
Author: Kummer, Susann; Avinoam, Ori; Kräusslich, Hans-Georg
Title: IFITM3 Clusters on Virus Containing Endosomes and Lysosomes Early in the Influenza A Infection of Human Airway Epithelial Cells
  • Document date: 2019_6_12
    • ID: 1345qct4_14
    Snippet: Whole-cell lysates were prepared using a 6 × lysis buffer (10% sucrose, 0.1% bromophenol blue, 5 mM EDTA pH 8.0, 200 mM Tris pH 8.8, 10% SDS, and 2% beta-mercaptoethanol). Protein samples were resolved by 12.5% SDS-PAGE at 160 Volt for 1.5 h. Then, the proteins were transferred to Immobilon-FL membranes (Merck, Darmstadt, Germany). The membranes were blocked in Li-Cor blocking buffer for 30 min at room temperature and probed with first antibodie.....
    Document: Whole-cell lysates were prepared using a 6 × lysis buffer (10% sucrose, 0.1% bromophenol blue, 5 mM EDTA pH 8.0, 200 mM Tris pH 8.8, 10% SDS, and 2% beta-mercaptoethanol). Protein samples were resolved by 12.5% SDS-PAGE at 160 Volt for 1.5 h. Then, the proteins were transferred to Immobilon-FL membranes (Merck, Darmstadt, Germany). The membranes were blocked in Li-Cor blocking buffer for 30 min at room temperature and probed with first antibodies (rabbit anti-IFITM3, catalogue no. 11714-1-AP, Proteintech, Manchester, United Kingdom, and rabbit anti-actin, catalogue no. A2066, sigma-aldrich, Steinheim, Germany, mouse anti-tubulin no. T5168, sigma-aldrich, Steinheim, Germany, or mouse anti-GAPDH, catalogue no. sc365062, Santa Cruz, Dallas, Texas, USA) in a blocking solution diluted 1:5000 at 8 • C overnight. After three repeated washings in 1 × TBST for 15 min, the membranes were incubated with a secondary donkey anti-mouse 800 antibody (catalogue no. 926-32212, Li-Cor) or secondary donkey anti-mouse 700 antibody (catalogue no. 925-68072, Li-Cor, Lincoln, Nebraska USA), for 1 h at room temperature in the dark. The protein bands were visualized using the Li-Cor Odyssey scanner (Li-Cor, Lincoln, Nebraska USA).

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