Decoupled and Real-Time Automation of the HX-MS Experiment

The HX experiment can be automated by following two distinct experimental workflows. In one paradigm, the experimental design decouples automated digestion and LC-MS analysis from the HX sample preparation step (i.e., decoupled HX-MS). To enable the LC-MS analysis to be decoupled from the HX expraiment, samples are flash frozen in liquid nitrogen afta- the quench and stored at -80°C until required. Samples can be stored for days/weeks at -80°C with minimal loss of deuterium label. When the sample is ready for LC-MS analysis, it is thawed immediately prior to analysis with LC-MS by the autosampler. 

The first semiautomated HX-MS system was described by Virgil Woods in 2001 and was based around the decoupled experimental paradigm [44], Woods outlined an end-to-end HX workflow, titled DXMS, that began with (manual) sample preparation and freezing of samples in quench solution. When ready, samples were placed in a modified HPLC autosampler able to perform the thawing, digestion, and LC-MS analysis. Although this automation was never commercialized, it was in use for over a decade and was demonstrated to be extremely productive. 

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