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High viscosity extruder can be used to collect serial data on membrane or soluble protein crystals that are embedded in viscous media (for example LCP). We are using the high viscosity extruder developed at the Arizona State University (Weierstall et al., Nature Commun. 2014).
Figure from Weierstall et al., Nature Commun. 2014. Middle section through the LCP injector
The high viscosity extruder can easliy be installed on I24 beamline upon request. Installation takes about 30min-1h. Reservoir capacity is 20 µl. This is enough for ~ 2h of data collection on I24 and, given that crystal concentration is high enough, is also enough to collect a full dataset.
High viscosity extruder installed on I24 beamline
High viscosity extruder: from crystals to structure
Video of LCP going through 3 different capillary sizes (offline tests)
Ideally you will need membrane protein microcrystals in LCP (grown in syringes, not plates) with a concentration of at least 106 crystals/ml or a slurry of soluble protein microcrystals at 107-108 crystals/ml. High crystal concentration ensures high hit rates during data collection so it is crucial for reducing the time that is necessary to collect the data. Crystals grown in LCP can be directly loaded into the reservoir (20 µl) from the syringe. For soluble protein microcrystals, we will need to mix the crystal pellet with viscous media matrix. This is not an exhaustive list, but we typically use LCP, agarose, grease, or cellulose-based media. Each of media types will have their advantages and disadvantages. Importantly, not every matrix can be used with each sample. Ideally, different media types are tested with the sample of interest in the offline setup at Diamond prior to data collection. We provide full support on the experiment including sample preparation, running the experiment, real-time data processing and postprocessing.
Lysozyme crystals mixed with LCP.
Loading the sample into the reservoir.
If you have any more quest please contact us.
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