FPOP Tutorial Series: Reproducibility and Robustness in FPOP

Part of our video tutorial series on Fast Photochemical Oxidation of Proteins, a mass spectrometry-based structural biology tool. In this video, I discuss the phenomenon of hydroxyl radical scavenging. I introduce the concept of hydroxyl radical dosimetry, and its importance in ensuring robustness and reproducibility in your FPOP experiments. I discuss methods to correct for the addition of new radical-scavenging entities to your sample, including the concept of compensation. I show some of the experimental data demonstrating that compensation allows you to overcome the addition of radical scavengers to your solution, and introduce a new method for real-time monitoring of hydroxyl radical dosimetry and scavenging. I also discuss some practical pitfalls for the new FPOP experimenter when trying to compensate for scavengers. This video is made possible by a grant from the National Science Foundation (CHE 1608685). Support for the work this video is based upon comes from the National Institutes of General Medical Sciences, National Institutes of Health (R01 GM127267). Any distribution of this video should note the support of the National Science Foundation and the National Institutes of Health. Financial conflict of interest disclosures are made in the body of the video. Copyright Joshua S. Sharp. Work is available under a Creative Commons license with attribution and share-alike (CC BY-SA).