Find the Linear Rangeįor western blot quantification, you must ensure your image was captured in a manner sensitive enough to detect a change in what we call the “linear range”. Four Simple Steps for Western Blot Quantification 1. So on to the four critical steps for ensuring your (semi) quantification is accurate. This is because they cannot tell you exactly how much of your protein of interest there is, only how much there is relative to another sample.Įven though western blot quantification is only semi-quantitative, you still need to be rigorous in how you quantify your blots to ensure your results are as accurate as possible. Western Blots Are Semi-Quantitativeīefore we take you through the necessary steps for quantifying your western blots, we need to clarify that western blots are only semi-quantitative. This means you can see how the levels of your protein of interest change between time points, conditions, and treatments. The value you obtain is directly proportional to the concentration of your protein of interest.Ī quantitative western blot will allow you to measure relative changes between different conditions, so rather than just saying a protein is present or absent in a sample, you can go further and say your target protein is present x-times higher in one sample compared to another for example. Why Should You Perform Western Blot Quantification?įirst, what do we actually mean by western blot quantification? Put simply, this refers to the measurement of the signal emitted by your target protein. While it can be used to simply show the presence or absence of a protein in a sample, it can tell you so much more if you take the next step and perform western blot quantification.Īs scientists, we love nothing more than quantitative data! Here, we have pulled together the 4 key steps for you to perform western blot quantification. It separates proteins based on size and then uses antibodies to detect specific proteins within the sample. Western blotting is a commonly used lab technique to identify proteins within a sample.
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