Samples were homogenized in chilled radioimmunoprecipitation assay (RIPA) buffer (MilliporeSigma, St. Louis, MO, USA, cat# R0278) with Pierce protease-inhibitor cocktail (ThermoFisher Scientific, Waltham MA, USA cat# A32963) and 1 mM sodium vanadate (MilliporeSigma, St. Louis, MO, USA, cat# S6508). After brief sonication (Dismembrator Model F60, ThermoFisher Scientific, Waltham, MA, USA), the samples were prepped in Laemmli buffer, after which 15 to 30 µg of the protein extracts were loaded for SDS-PAGE. Antibodies and the dilutions used in the study are provided in Supplementary Table S1 . Proper loading was confirmed by probing with GAPDH antibodies for primary heart tissue and β-actin for cell lines. Immunoblot images were captured using an Odyssey® Fc Imaging System, and densitometry of blots was performed using Image Studio software version 5.2 (LI-COR Biosciences, Lincoln, NE, USA) normalized to loading controls. Uncropped blot images are provided in Supplementary Figure S6 . TG content of myocardial tissue and serum was assessed using a Triglyceride Colorimetric Assay Kit (Cayman Chemical, Ann Arbor, MI, USA, cat# 10010303) according to the manufacturer’s instructions.
Dismembrator model f60
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Dismembrator Model F60 is a laboratory equipment designed for homogenization and cell disruption. It utilizes a high-speed motor to drive a pestle that pulverizes and disperses samples in a grinding chamber. The Dismembrator is capable of processing a variety of materials, including tissues, cells, and powders, to obtain a homogeneous suspension or fine powder.
Automatically generated - may contain errors
Lab products found in correlation
Odyssey fc imaging system, by LI COR (2 mentions)
Pierce protease inhibitor cocktail, by Thermo Fisher Scientific (2 mentions)
Triglyceride colorimetric assay kit, by Cayman Chemical (1 mentions)
Ripa buffer, by Merck Group (1 mentions)
Image studio software version 5, by LI COR (1 mentions)
Ne per, by Thermo Fisher Scientific (1 mentions)
Image studio software, by LI COR (1 mentions)
2 protocols using dismembrator model f60
1
Protein Extraction and Western Blot Analysis of Myocardial Tissue
2
Analyzing CREB3 Cleavage in Heart Tissues
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Heart tissues were homogenized in ice-cold radioimmunoprecipitation assay buffer (MilliporeSigma, catalog no. R0278) with Pierce Protease Inhibitor cocktail (Thermo Fisher Scientific, catalog no. A32963) and 1 mM sodium vanadate (MilliporeSigma, catalog no. S6508). Following brief sonication (Dismembrator Model F60, Thermo Fisher Scientific), the samples were prepped in Laemmli buffer after which 15 to 30 μg of protein extract were loaded for SDS–polyacrylamide gel electrophoresis (SDS-PAGE). For immunoblots assessing CREB3 cleavage, cells were directly lysed in 2x Laemmli buffer and boiled for 5 min before loading in SDS-PAGE. Nuclear fractionation was done using NE-PER (Thermo Fisher Scientific, catalog no. 78833). Antibodies and dilutions used are provided in the table S4. Proper loading was initially confirmed by Ponceau S and then by probing with glyceraldehyde-3-phosphate dehydrogenase antibodies or otherwise indicated. Image capture was performed by Odyssey Fc Imaging System, and blot quantification was performed using Image Studio software (LI-COR Biosciences).
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