Abstract
Crystal harvesting has proven to be difficult to automate and remains the ratelimiting step for many structure-determination and high-throughput screening projects. This has resulted in crystals being prepared more rapidly than they can be harvested for X-ray data collection. Fourth-generation synchrotrons will support extraordinarily rapid rates of data acquisition, putting further pressure on the crystal-harvesting bottleneck. Here, a simple solution is reported in which crystals can be acoustically harvested from slightly modified MiTeGen In Situ-1 crystallization plates. This technique uses an acoustic pulse to eject each crystal out of its crystallization well, through a short air column and onto a micro-mesh (improving on previous work, which required separately grown crystals to be transferred before harvesting). Crystals can be individually harvested or can be serially combined with a chemical library such as a fragment library.
| Original language | English |
|---|---|
| Pages (from-to) | 986-999 |
| Number of pages | 14 |
| Journal | Acta Crystallographica Section D: Structural Biology |
| Volume | 74 |
| DOIs | |
| State | Published - 2018 |
| Externally published | Yes |
Funding
Major ongoing financial support for acoustic droplet ejection applications was through the Life Science Biomedical Technology Research resource, supported by the National Institutes of Health, National Institute of General Medical Sciences (NIGMS) through a Biomedical Technology Research Resource P41 grant (P41GM111244) and by the DOE Office of Biological and Environmental Research (KP1605010). Additional funding was granted by the National Science Foundation and the National Institutes of Health/ National Institute of General Medical Sciences under NSF award DMR-0936384 awarded to the MacCHESS facility, which is supported by award GM-103485 from the National Institute of General Medical Sciences, and by the SSRL Structural Molecular Biology Program, supported by the DOE Office of Biological and Environmental Research and by the National Institutes of Health, National Institute of General Medical Sciences (including P41GM103393). Personnel for this study were recruited largely through the Science Undergraduate Laboratory Internships Program (SULI) in the summers of 2013, 2014 and 2017, supported through the US Department of Energy, Office of Science, Office of Workforce Development for Teachers and Scientists (WDTS). Data for this study were measured on beamline X25 of the National Synchrotron Light Source (NSLS), on beam-line A1 of the Cornell High Energy Synchrotron Source (CHESS), on beamline BL14-1 of the Stanford Synchrotron Radiation Lightsource (SSRL) and on beamline 17-ID1 (AMX) at the National Synchrotron Light Source II (NSLS II). We thank Labcyte Inc., and especially Joe Olechno, Richard Ellson and Richard Stearns, for their technical support and guidance. Author contributions are as follows. LL and ASS designed the experiment. ASS wrote the paper with input from all authors. YNS, HMB, LM, MTB, DL, SLC, RN, ASS and LL grew crystals, obtained data and analyzed data. JMW and KG performed three-dimensional visualization. ASS and SM trained and supervised student interns.
Keywords
- Acoustic droplet ejection
- Automation
- Crystal harvesting
- Crystal mounting
- Crystallography
- Drug discovery
- High-throughput screening
- Microcrystals