Cells were seeded in 6-well plates, collected using a cell scraper and suspended in RIPA buffer (Thermo Scientific, Pittsburgh, PA) supplemented with protease inhibitor cocktail and phosphatase inhibitor (Sigma Aldrich). Cells were homogenized over ice by sonication. After purification of the sample by centrifugation, protein concentration was determined by protein assay (Bio-Rad, Hercules, CA). The proteins were separated by 4–12% SDS–polyacrylamide gel electrophoresis (SDS–PAGE) and electrically transferred to a polyvinylidene difluoride membrane (Santa Cruz Biotechnology). After blocking the membrane using 5% non-fat milk, target proteins were detected using anti-IFITM1, anti-PARP, anti-ERα, anti-phospho-STAT1 (ser701), anti-STAT1, anti-p21, anti-p53 or anti-laminin B (Santa Cruz Biotechnology) antibodies. Membranes were stripped and re-probed for β-actin (Cell Signaling). The appropriate horseradish peroxidase (HRP)-conjugated secondary antibody was applied and the positive bands were detected using Amersham ECL Plus Western blotting detection reagents (GE Health care, Piscataway, NJ) and exposed to autoradiography film (Midwest Scientific).
Anti laminin b
Manufactured by Santa Cruz Biotechnology
Anti-laminin B is a lab equipment product offered by Santa Cruz Biotechnology. It is a reagent designed to detect and quantify the presence of laminin B, a key structural protein found in the extracellular matrix. The product can be utilized in various research applications that require the identification and analysis of laminin B.
Automatically generated - may contain errors
Lab products found in correlation
Ripa buffer, by Thermo Fisher Scientific (2 mentions)
Protease inhibitor and phosphatase inhibitor cocktail, by Merck Group (1 mentions)
Autoradiography film, by GE Healthcare (1 mentions)
Protein assay, by Bio-Rad (1 mentions)
Amersham ecl plus western blotting detection reagent, by GE Healthcare (1 mentions)
Anti ifitm1, by Santa Cruz Biotechnology (1 mentions)
Anti parp, by Santa Cruz Biotechnology (1 mentions)
Anti p21, by Santa Cruz Biotechnology (1 mentions)
Anti erα, by Santa Cruz Biotechnology (1 mentions)
Anti p53, by Santa Cruz Biotechnology (1 mentions)
β actin, by Cell Signaling Technology (1 mentions)
Anti lamin a c, by Cell Signaling Technology (1 mentions)
Anti ha, by Abcam (1 mentions)
Anti α tubulin, by Abcam (1 mentions)
Anti ctbp1, by BD (1 mentions)
Anti flag, by Cell Signaling Technology (1 mentions)
Anti flag m2, by Merck Group (1 mentions)
Imagej software, by Bio-Rad (1 mentions)
Nuclear extract kit, by Active Motif (1 mentions)
Anti ctbp2, by BD (1 mentions)
2 protocols using anti laminin b
1
Western Blot Analysis of Protein Expression
2
Protein Extraction and Western Blot Analysis
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Total protein extracts were prepared as previously described (Fedele et al., 2017 (link)). For lysates preparation from frozen GBM samples, small pieces in the range of 0.02 to 0.05 grams were selected, washed with cold PBS Buffer and then treated with RIPA buffer, containing Phosphatase/Protease Inhibitors and PMSF. The tissues were disrupted using pestles and then processed according to our standard protocol (Fedele et al., 2017 (link)). Cytoplasmic and nuclear extracts were prepared using a Nuclear Extract Kit (Active Motif) according to the manufacturer’s instructions. Western blots were performed using the following antibodies: anti-ZBTB18 (Abcam # ab118471); anti-ZNF238 (Sigma # SAB1406998); anti-FLAG M2 (Sigma #F1804); anti-FLAG (Cell Signaling Technologies #14793), anti-HA (Abcam ##18181), anti-CTBP1 (BD Biosciences #612042); anti-CTBP2 (BD Biosciences #612044); anti-CAPN2 (Cell Signaling Technologies #2539S); anti-CAST (Cell Signaling #4146S) anti-alpha Tubulin (Abcam #ab7291) and anti-Laminin B (Santa Cruz #sc-6216), anti Lamin A/C (Cell Signaling Technologies ##4777). The quantification of western blot bands was performed using the ImageJ software (Biorad) and normalized to the level of αTubulin.
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