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Fluorescence detection enables quantitative, multiplex analysis of western blots on your bench top—without the need for ECL optimization, film or a darkroom. While the detection limits are still not as low as chemiluminescent detection, fluorescent detection has the unique advantage of allowing multiple targets to be assayed for on the same blot at the same time without the need to strip and reprobed. We offer Invitrogen™ Alexa Fluor™ dyes and Invitrogen™ WesternDot™ probes for Western blotting using fluorescence detection.

Alexa Fluor dyes are trusted, high performance fluorescent dyes backed by our experienced, problem-solving technical support team. Long-wavelength reporters like WesternDot probe antibodies are detected on membranes with high sensitivity and minimal background signal or scatter.

Protein method: Fluorescent Western Blotting

Featured product

iBind-210x210

The Invitrogen™ iBind™ Western System is an automated western blot processing platform that requires less primary antibody and delivers sensitive, reproducible western results. All blocking, antibody incubation, and washing steps are hands-free, allowing you to load your solutions and walk away. There is no electricity or battery required. You can also use your existing chemiluminescent, chromogenic, or fluorescent western detection protocols, including HRP-, AP-, Alexa Fluor dye–, and IRDye™ (LI-COR™)–conjugated secondary antibodies.

WesternDot conjugates enable multiplex beyond two colors when other fluorescent detection reagents cannot. WesternDots probes have fluorescent properties that allow them to be excited by UV but detected at much longer wavelengths. Other kinds of fluorescent detection reagents that are added in two color experiments need to be excited by wavelengths of light that also excite the inherent fluorescence of membranes which leads to high background and lower signal to noise ratios. Detect both strong and weak signals at the same time with a 4,000-fold linear dynamic range. With an appropriate reader, you can multiplex up to three probes on the same blot—providing an extra level of precision and biological context for your measurements.

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flourescence

Alexa Fluor dyes are trusted, high-performance fluorescent dyes that offer intense fluorescence, and increased photostability. Power your Western blotting with Alexa Fluor dye–labeled antibodies for the consistency and high quality you need. Using Alexa Fluor 680 and Alexa Fluor 790 secondary antibodies, you can generate multicolored western blots that can be imaged on standard near-IR fluorescence scanners such as the LI-COR™ Odyssey™ Imaging System and the Kodak™ image station.