Sevrin_CellRep_2024

Published: 13 August 2024| Version 1 | DOI: 10.17632/6nmytyss4n.1
Contributor:
Thomas Sevrin

Description

Original Western Blot images from all figures of Sevrin et al. 2024 Cell Report paper.

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Steps to reproduce

Protein concentration of cell lysates was determined using the BCA assay according to the manufacturer’s instructions (PierceTM BCA Protein Assay Kit). Loading solutions were prepared at 1 µg protein/µL using 4X Laemmeli buffer and DTT. Ten micrograms of protein were resolved by SDS PAGE using 10% acrylamide gels in Mini-PROTEAN Tetra cell (Biorad) systems. Proteins were then transferred into PVDF membranes using the XCell IITM wet transfer system (Invitrogen). Membranes were blocked for 1h at room temperature using 4% BSA in TBS-Tween 0.1% (TBS-T), then incubated with primary antibody (diluted 1:1000 in 1% BSA in TBS-T) from 1h at room temperature to overnight at 4°C. Membranes were then incubated for 1h at room temperature with HRP-coupled secondary antibodies diluted at 1:5000 in 1%BSA in TBS-T. Finally, blots were developed using homemade ECL reagent or SuperSignal™ West Femto reagent (Thermo Scientific) with the iBright™ CL750 Imaging System (Thermo Scientific).

Institutions

Categories

Drug Resistance in Cancer, Pancreatic Cancer, Western Blot, Targeted Therapy

Funders

  • National Cancer Institute
    Maryland, United States
    Grant ID: R01CA244660
  • Horizons Foundation
    California, United States
    Grant ID: 101136926
  • Science Foundation Ireland
    Ireland
    Grant ID: 22/PATH-S/10875

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