In-gel Fluorescence

Fluorescence detection in-gel electrophoresis

In-gel fluorescence has become popular for validating the generation of fusion proteins, analyzing protein-protein interactions, and electrophoretic mobility shift assays. Fluorescent fusion proteins can easily be visualized without having to transfer to a membrane for traditional Western analysis. In the image below, fluorescent proteins were detected in a native gel and imaged with an Azure Imaging System.

Fluorescent protein in native gel.
Fluorescent protein in native gel imaged with an Azure imager

Visualizing proteins in-gel without Western Blotting

Another application of in-gel fluorescence is staining the total protein in a gel before transfer for Western blottingAzureRed protein stain is compatible with downstream Western blotting. It can also provide fluorescent staining of total protein in your gel post-electrophoresis, for relative quantitative comparisons. AzureRed can be imaged with an Azure Imager, or another CCD- or laser-based fluorescent imaging system. By including known concentration standards and imaging the stained gel, you can get quantitative information about your gel bands.

Read more: Accurate Western Blot Normalization with AzureRed Fluorescent Protein Stain

Imaging fluorescent proteins in-gel for a gel-shift assay


The image above shows the interaction of Fluorescein-labeled Ubiquitin (FL-Ub) with its binding partners, E1 and E2 Ubiquitin-activating enzymes, over a treatment time course. The upper bands represent ubiquitin in complex with E1 or E2. Fluorescence was detected with the Azure c600 using excitation at 470 nm and emission filter at 497 nm.

What to use for the best in-gel fluorescence results

After fluorescent proteins are detected, the fluorescence can be imaged with the Azure 400, Azure 600, new Azure Sapphire FL Biomolecular Imager, or any other digital imager. Learn more about how to get the best results by filling out this form for more information.

Read more: How to Normalize Western Blots to Total Protein


New to Western blotting? Need to troubleshoot your Western blot?​ Want to brush up on Western blotting best practices? Claim your free Western Blotting eBook!
Related Products
Azure 400 Visible fluorescent imaging system

Flexible imaging system able to detect three fluorescent targets

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The ultimate Western blot imaging system, capable of in-gel fluorescence, and more

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