Western Blot Analysis of Protein Levels

Total cell lysates were prepared as previously described [24] (link). The protein concentration was determined using Bio-Rad protein assay reagent. A total of 70 µg of proteins was resolved on 10% Bis–Tris gels and transferred to nitrocellulose membranes. The protein levels of target genes were detected using specific primary antibodies and IRDye secondary antibodies on Odyssey Fluorescence Imaging System (LI-COR Biosciences, USA). The density of the immunoblots was analyzed using Odyssey V3.0 software and normalized with β-Actin.

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Zhang X., Wang G., Gurley E.C, & Zhou H. (2014). Flavonoid Apigenin Inhibits Lipopolysaccharide-Induced Inflammatory Response through Multiple Mechanisms in Macrophages. PLoS ONE, 9(9), e107072.

Publication 2014

protein assay reagent Odyssey Fluorescence Imaging System

A proteins Actin Antibodies Assay Bis tris Cell Gels Immunoblots Membranes Nitrocellulose Protein Protein genes Rad protein

Corresponding Organization : Wenzhou Medical University

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Protein levels of target genes

control variables

Protein concentration determined using Bio-Rad protein assay reagent
70 µg of proteins resolved on 10% Bis–Tris gels
Proteins transferred to nitrocellulose membranes
Protein levels detected using specific primary antibodies and IRDye secondary antibodies on Odyssey Fluorescence Imaging System
Immunoblot density analyzed using Odyssey V3.0 software
Immunoblot density normalized with β-Actin

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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