When you’re shopping for antibodies, there are so many factors to consider. For example, will it work in my cell or tissue model? Has it been tested in the application I want to use? Sometimes it’s a struggle to find what you need because your options are limited, but in other instances there may be several reagents that seem like they could work in your experiment.
It's time to check out another video from the CST Tech Tips playlist! In this edition of Tech Tips, we'll tackle a common protocol question customers ask our ChIP team: how much antibody to use for chromatin immunoprecipitaion (ChIP) experiments. Adding more antibody isn't always better - watch the video to learn why.
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The use of multiple antibodies in a single experiment can provide useful information to researchers. Co-staining with multiple antibodies and cellular dyes is a simple, low-content form of multiplex analysis. Techniques for performing multiplex analyses in cells and tissues are powerful research tools that are applicable to general cell biology studies as well as diagnostic purposes. These techniques allow researchers to detect multiple biomarkers to assess their samples. They also allow for easy colocalization studies to determine relationships between analytes. Here we describe two common techniques for fluorescent staining using multiple antibodies in the same assay.
Early exploration of unmapped biological signaling pathways were carried out using radiolabeled phospho-imaging. The development of phospho-specific antibodies to detect and quantify protein phosphorylation made life easier for researchers (less 32P waste to deal with), but the interpretation of data from these experiments comes with its own set of caveats.
well-validated antibody, the workhorse of immunofluorescence. If you are a seasoned pro at IF experiments, you are probably used to checking the antibody datasheet (or web page) for the recommended dilution. But have you ever wondered where those recommendations come from?
After months of hard work, your research has honed in on a hypothesis you can test with immunofluorescence (IF). You've chosen antibodies and performed pilot IF experiments (see The Importance of Validation), and the localization of the protein appears reasonable. But how can you be sure the IF data you've acquired represents real biological phenomena? We present two examples of experimental controls in this post.
After months of hard work, your research has zeroed in on a hypothesis you can test with immunofluorescence (IF). But now you have to make a choice. How do you decide which antibody to use to get reliable IF results? How do you know if the images are accurately reporting the target's localization? We explore some considerations in this post.
Part 1 gave an overview on mass spectrometry-based proteomics. Now it’s time to talk about how this strategy can be used to identify peptides with post-translational modifications (PTM) from a complex biological sample.
If you buy primary antibodies from different places, you may have noticed that many companies list the size of their products in micrograms per ml (i.e., by weight). You may have also noticed that CST does not - CST antibodies are listed by performance (e.g., 10 western blots) instead.
Specificity, consistency, and optimized assay conditions are three key elements that help ensure reliable immunofluorescence (IF) staining results each and every time. So, before starting an experiment you should ask yourself the following three questions:
- Is your antibody specific?
- Is your antibody supported by an optimized IF protocol?
- Is your antibody performing consistently?
Let's take them each in turn...