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Automated Crystal Harvesting

 

 

Automated crystal harvesting & cryo-cooling plus remote in-plate x-ray characterization are now available at the HTX lab and BM14.

 

Automated Crystal harvesting and Cryo-cooling are now available thanks to the CrystalDirect technology(1,2) developed by the HTX lab and the EMBL instrumentation team. This system is based on a new crystallization plate that allows growing crystals on very thin films that can then be excised with a LASER beam to recover the crystalline material. The harvested film pieces are attached to a SPINE standard pin and stored in liquid nitrogen for synchrotron data collection. In addition to the full automation of the crystal harvesting and cryo-cooling process, the CrystalDirect system offers some advantages, like a reduced mechanical stress for the crystals (as no tools enter the crystallization drop) and a simplified and efficient cryo-cooling method that does not require the addition of cryo-protectants(3). Specific Crystal Pointing tools have been developed in CRIMS to enable remote operation.  

 

CrystalDirect plates

 

 

The CrystalDirect Harvester

For the harvesting process, crystal position are marked using the Xtal pointing tool provided in the CRIMS image viewer. Positionning information is then transferred from CRIMS to the harvester (See tutorial). Crystals are recovered by cutting the film around them with a LASER working in photo-ablation mode.

This system is not restricted by crystal size, reduces mechanical stress and permits reproducible and operator-error free harvestings. Customized cut shapes allow to match the size and shape of a single crystal, but also to perform crystal surgery or alternatively to recover full drop containing multiple crystals in a single x-ray data collection pin.

 

harvesting shapes

 

 

In-situ characterization

Due to their design, CrystalDirect plates allow to collect diffraction data in-situ with very low background. In collaboration with the BM14 beam and Instrumentation teams, we have developed a plate goniometer and crystal pointing interfaces in CRIMS As well as for the CrystalDirect harvester, the Xtal pointing tool allows to mark a crystal position through CRIMS. The marked position is then transferred to the BM14 beam line and used to align the crystal with the x-ray beam. A dedicated goniometer head for in-situ experiments has been developed by the EMBL instrumentation team that is now installed at BM14.

 in-situ config at BM14

 

 

 


References

1. Marquez & Cipriani. Methods Mol Biol. 2014, 1091: 189-95. PMID:23695565

2. Cipriani et al., Acta Crystallogr D Biol Crystallogr. 2012 Oct;68:1393-9. Epub 2012 Sep 18. PMID: 22993093

3. Pellegrini et al, Acta Crystallogr D Biol Crystallogr. 2011 Oct;67:902-6. Epub 2011 Sep 8. PMID: 21931222