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A quick overview of the steps you can take to ensure your Western blots are quantitative. This free guide includes a troubleshooting section and tear-out quantitative Western blotting checklist.
Getting quantitative data from your Western blot takes more than just a fluorescent secondary and digital imager. You need to consider your sample prep, your normalization method, and the linearity of your signal. To help keep your quantitative Western blot workflows on track, we’ve created a checklist of steps that can be taken to ensure your Western blot quantitation is as accurate as it can be.
Check out this publication from Boys Town National Research Hospital that imaged chemiluminescent Western blots with an Radiance HRP Substrate and an Azure imager.
Have you treated your samples consistently, ensuring cells and tissues are thoroughly lysed, immediately placed on ice, and/or been treated with appropriate reagents such as protease inhibitors?
Have you treated your samples consistently, ensuring cells and tissues are thoroughly lysed, immediately placed on ice, and/or been treated with appropriate reagents such as protease inhibitors?
Are you using a robust and consistent western blotting protocol (remember that you can reduce variability by using the same reagents for each blot or implementing lot-to-lot controls, and keeping your transfer conditions and incubation and wash times the same)?
Are you normalizing to total protein?
Have you validated the specificity of your antibodies?
Have you verified that your system is linear?
With products that offer sensitive detection, wide dynamic range, easy-to-use features, and lot-to-lot consistency Azure Biosystems expertly streamlines and supports your quantitative fluorescent Western blot workflows.
