ResourceSpectroscopy
Imaging Three-color Western Blots with the Azure c600
Imaging Three-color Western Blots with the Azure c600
10 Sept 2015This application note demonstrates a procedure to obtain three-color fluorescent blots using WesternDot® reagents. Multicolor detection is a powerful application of fluorescent Western blotting. Two or more proteins can be detected on one blot using antibodies labeled with different fluorophores. The ability to simultaneously assay multiple proteins on a Western blot represents a significant advantage of fluorescent detection over chemiluminescent detection, because chemiluminescent detection requires stripping and re-probing a blot or using duplicate blots to detect two proteins, approaches that can introduce errors and variation.
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Fluorescence SpectroscopyFluorometers and spectrofluorometers (also called fluorescence spectrometers) are used to measure the intensity and wavelength of fluorescent light emitted from a sample after excitation by illumination. Spectrofluorometers utilize monochromators to select the desired wavelengths, whereas filter fluorometers employ a set of filters. Spectrofluorometers for measuring steady-state fluorescence and lifetime fluorescence (or time-resolved fluorescence) are available, as well as fluorescence microscopes and microplate readers. Find the best fluorescence spectroscopy products in our peer-reviewed product directory: compare products, check customer reviews and receive pricing direct from manufacturers.AntibodiesAntibodies are used in techniques such as confocal and fluorescence microscopy, flow cytometry, ELISA, ELISPOT, immunohistochemistry, western blotting and immunopreciptation. Select specific antigen reactivity, high specific affinity, low non-specific binding, monoclonal or polyclonal, primary or secondary antibodies and associated conjugates such as an enzyme or dye for visualization.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.FluorescenceThe emission of fluorescence occurs when a photon of energy is supplied to a fluorescent chemical compound by an external source, causing it to become excited. Fluorescence can be detected and measured for different purposes using microplate readers, fluorescence microscopes, fluorescence scanners, and flow cytometers.ChemiluminescenceAntibioticsAntibiotics are substances that kill or inhibit the growth of microorganisms, such as bacteria and fungi. They are essential in the treatment of infections and in preventing disease spread. Explore antibiotics research and diagnostic tools in our peer-reviewed product directory; compare products, check reviews, and get pricing directly from manufacturers.Blotting