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Western Blot Normalization with Azure Biosystems
Western Blot Normalization with Azure Biosystems
15 Sept 2015Normalization is critical for obtaining accurate, quantitative data from Western blots. Normalization accounts for unequal loading of samples across the lanes on a gel, and for differences in transfer efficiency across a blot. Traditionally, protein normalization for Western blots involves comparing the relative abundance of a protein of interest to that of an unrelated protein, the normalization or loading control. This application note explains a total protein normalization (TPN) method, in which the relative abundance of the protein of interest is compared to the total protein content of the sample.
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Western BlottingWestern blotting equipment is used to transfer and identify specific proteins within a sample, reveal protein modifications, as well as give a semi-quantitative estimation of their concentration. Western blotting equipment includes all apparatus necessary to transfer proteins from gel to membrane and subsequent processing steps. Protein transfer can be performed by electroblotting with wet, semi-dry and dry transfer systems onto nitrocellulose and PVDF membranes. Blocking, washing and labeling of membranes follows, involving buffers, blocking reagents, blotting / incubation trays, labeling reagents, immunoblotting assays, antibodies and conjugates. Automated equipment for these steps is available to accelerate your lab workflow. Finally, detection and imaging of proteins can be conducted using gel documentation and imaging systems. Find the best western blotting equipment in our peer-reviewed product directory: compare products, check customer reviews and receive pricing direct from manufacturers.Gel Doc / Image AnalysisGel documentation (gel doc) or gel imaging systems are used for the analysis of proteins, antibodies and nucleic acid immobilized in polyacrylamide or agarose gels, membranes or microarrays. Explore a range of a gel imaging systems, densitometers, scanners, transilluminators or UV lamp + CCD cameras for your image analysis solutions. Colorimetric, fluorescent and/or radioisotopic samples can be visualized and documented for further analysis. See gel doc / Image analysis software for quantitative 1D and 2D analysis of your samples. Find the best gel doc / image analysis products in our peer-reviewed product directory: compare products, check customer reviews and receive pricing direct from manufacturers.Protein PurificationProtein purification is a vital step in drug discovery, therapeutics, biotech and life science research. The purification process typically involves subcellular or membrane protein extraction with cell lysis kits, separation of proteins from cell debris by filtration or spin columns, and the isolation of proteins of interest from other proteins and impurities with affinity purification (including fusion protein tags and antibody binding proteins A, G and L), immunoprecipitation or chromatographic methods, such as ion exchange, size exclusion and immobilized metal affinity chromatography. All purification methods come in multiple formats for your laboratory needs, including agarose or magnetic beads, resins, columns and filter plates. Find the best protein purification equipment in our peer-reviewed product directory: compare products, check customer reviews and receive pricing direct from manufacturers.Protein QuantificationThe detection and quantification of proteins in a sample is vital across life sciences, pharmaceutical research and clinical diagnostics, and a variety of equipment is available to scientists to simplify the workflow. Proteins of interest can be easily labeled and detected on light-based detection instruments. Immunoassay kits allow you to identify a specific protein of interest and protein detection beads or antibody microarrays allow you to identify multiple specific proteins at once. Protein interactions and enzyme activity can also be monitored with protein-protein interaction assays. Additionally, the biophysical characterization of proteins is made easy with biokinetic analyzers. Find the best protein detection and quantification products in our peer-reviewed product directory: compare products, check customer reviews and receive pricing direct from manufacturers.Gel ImagingGel imaging captures high-resolution images of electrophoresis gels to analyze DNA, RNA, and proteins. Essential in molecular biology, this technique helps visualize results for accurate quantification and documentation. Discover top imaging systems and compare user-reviewed options in our directory.Protein BiologyThe analysis of protein expression, identity and function is vital for many areas of life science research and drug discovery. Some of the most commonly used techniques in protein analysis include Western blotting, electrophoresis and mass spectrometry.Blotting