In-gel Fluorescence

Contents

Fluorescent 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

Fluorescein-labeled-Ubiquitin

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 fluorescent signal can be imaged with any modern imager with the correct capabilities, such as the Azure 400, Azure 600, or new Azure Sapphire FL Biomolecular Imager. Learn more about how to get the best fluorescent imaging results by filling out this form for more information.

Read more: How to Normalize Western Blots to Total Protein

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Products that Detect In-gel Fluorescence

Azure 400

Flexible imaging system able to detect three fluorescent targets

Azure 600

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

Related Applications

Looking for a full list of applications?

Related blog posts about gel imaging…
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