A new view on crystal harvesting

Luft, Joseph R.; Grant, Thomas D.; Wolfley, Jennifer R.; Snell, Edward H.

doi:10.1107/s1600576714008899

Cited by 4 publications

(4 citation statements)

References 11 publications

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“…These small drop sizes necessarily produce small crystals that cannot be fished by traditional looping. Methods have been developed to harvest from the 1,536 plates 37 ; additionally, the plates with crystals have been used directly at synchrotron sources for in situ data collection 38 . If a robust method for harvesting these crystals were developed, advances in synchrotron technology and micro-focused beams would further enable useful datasets to be obtained.…”

Section: Discussionmentioning

confidence: 99%

High-Throughput Screening to Obtain Crystal Hits for Protein Crystallography

Budziszewski

Snell

Wright

et al. 2023

JoVE

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X-ray crystallography is the most commonly employed technique to discern macromolecular structures, but the crucial step of crystallizing a protein into an ordered lattice amenable to diffraction remains challenging. The crystallization of biomolecules is largely experimentally defined, and this process can be labor-intensive and prohibitive to researchers at resource-limited institutions. At the National High-Throughput Crystallization (HTX) Center, highly reproducible methods have been implemented to facilitate crystal growth, including an automated high-throughput 1,536-well microbatch-under-oil plate setup designed to sample a wide breadth of crystallization parameters. Plates are monitored using state-of-the-art imaging modalities over the course of 6 weeks to provide insight into crystal growth, as well as to accurately distinguish valuable crystal hits. Furthermore, the implementation of a trained artificial intelligence scoring algorithm for identifying crystal hits, coupled with an open-source, user-friendly interface for viewing experimental images, streamlines the process of analyzing crystal growth images. Here, the key procedures and instrumentation are described for the preparation of the cocktails and crystallization plates, imaging the plates, and identifying hits in a way that ensures reproducibility and increases the likelihood of successful crystallization.

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Section: Discussionmentioning

confidence: 99%

High-Throughput Screening to Obtain Crystal Hits for Protein Crystallography

Budziszewski

Snell

Wright

et al. 2023

JoVE

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“…Experiments using plates from the HTX Center have also shown that the HT 1536-well microassay plates can be used for in situ X-ray diffraction data collection (Bruno et al, 2016). It is also possible to harvest crystals directly from the HT 1536-well microassay plates (Luft, Grant et al, 2014).…”

Section: History and Development Of High-throughput Crystallization S...mentioning

confidence: 99%

20 years of crystal hits: progress and promise in ultrahigh-throughput crystallization screening

Lynch¹,

Snell²,

Potter³

et al. 2023

Acta Crystallogr D Struct Biol

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Diffraction-based structural methods contribute a large fraction of the biomolecular structural models available, providing a critical understanding of macromolecular architecture. These methods require crystallization of the target molecule, which remains a primary bottleneck in crystal-based structure determination. The National High-Throughput Crystallization Center at Hauptman–Woodward Medical Research Institute has focused on overcoming obstacles to crystallization through a combination of robotics-enabled high-throughput screening and advanced imaging to increase the success of finding crystallization conditions. This paper will describe the lessons learned from over 20 years of operation of our high-throughput crystallization services. The current experimental pipelines, instrumentation, imaging capabilities and software for image viewing and crystal scoring are detailed. New developments in the field and opportunities for further improvements in biomolecular crystallization are reflected on.

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“…Contact-free crystal harvesting is possible using laser tweezers (Wagner et al, 2013) or magnetic convection (Tung et al, 2014) with the assistance of experienced human operators. Liquid-handling-assisted harvesting is a promising alternative for minimizing solvent background in cases where the crystals are sufficiently robust to endure the solvent-removal process (Luft et al, 2014;Kitago et al, 2010). In cases where room-temperature diffraction data are advantageous, microfluidic traps (Lyubimov et al, 2015) or silicon chips (Owen et al, 2017) can leverage one harvest step into many diffraction experiments.…”

Section: Introductionmentioning

confidence: 99%