CrystalDirect Automatic Crystal Harvester

This video presents CrystalDirect®, a fully-automated crystal harvester robot working in a high throughput crystallography environment.
CrystalDirect® is a co-development of the European Molecular Biology Laboratory (EMBL) and Arinax Scientific Instrumentation.

High throughput harvesting process

CrystalDirect® uses EMBL well plates which are commercially available from MiTeGen (CrystalDirectTM plates). These plates are fully compatible with standard 96-well plates. They are composed of a plastic frame and a thin membrane on which the samples are deposited (“hanging drop” type). The CrystalDirect® robot cuts the membrane around one or more than one selected crystals by means of laser photoablation. (Cipriani et al.,2012)

Thereafter the crystal support is fixed on a SPINE sample pin and immediately cryo-cooled.

Thanks to this laser photoablation technology, CrystalDirect® gives also the possibility to separate nested crystals by laser cutting from crystal agglomerates (crystal surgery). Protein crystals harvested by this technique do not get deteriorated, neither does their diffraction power as could be demonstrated by numerous experiments. (Zander et al.,2016)

CrystalDirect® is a fully-automated system designed for high throughput crystal harvesting that fits perfectly to modern crystallization platforms: CrystalDirect® needs about 1 minute per harvest and reaches this performance with a success rate which is superior to 99 %.

Intended for highly automated workflow, CrystalDirect® is compatible with a large variety of sample changer robots and especially with the Flex ED 6-axis Robot, developed by the EMBL.

Multiple selectable harvesting methods

CrystalDirect® offers a large amount of cut shapes for the laser photoablation process.  The different shapes adapt to different protein crystal distributions and orientations on the native membrane. Consequently, CrystalDirect® can select and harvest single crystals, batches of crystals, micro-crystals and single needles from clusters.

From the left to the right: single crystal selection, multiple crystal selection, single needle crystal selection

Examples of cut shapes for automated crystal harvesting which are integrated in the software menu of CrystalDirect

Integrated cryo-cooling process

CrystalDirect® features an integrated cryo-cooling process. Just after each harvest, the system automatically moves the protein crystal into a gaseous stream of 100 K cold nitrogen.  The cryo-cooled sample can be transferred into a Dewar either manually or by means of a sample changer robot.

Cryo-protection by liquid removal

CrystalDirect® enables the removal of the mother liquor surrounding the protein crystals. In this way, the risk of mechanical stress to the crystal during the cryo-cooling process is strongly reduced. As a result, in most cases it isn’t necessary to add cryo-protectant (Pellegrini et al., 2011).

This automatic liquid removal process has a high level of quality (Zander, U., et al., 2016). In addition, the operating time and the repeatability are highly optimized compared to the manual approach. 

In situ soaking

The combination of the CrystalDirect® harvesting robot with CrystalDirect™ plates allows liquids to be added to the protein crystal with reduced risk of mechanical or chemical stress: a micro aperture is cut in the plate membrane by laser photoablation. Then, a drop of liquid is placed on the aperture and, in this way, the added liquid gradually diffuses into the mother liquor. This gentle method of in situ crystal soaking reduces the risk of chemical stress and eliminates any risk of mechanical stress which comes with crystal manipulation.

Web based selection

A Crystallization management software system that uses a sample tracking tool, e.g. CRIMS, gives the user the possibility to select protein crystals which are meant to be harvested. During this step the user also selects a cut shape and its orientation; all of this from their home laboratory. This information is then transferred to and used by CrystalDirect® to automatically harvest the chosen protein crystal(s).

CRIMS is a development of the EMBL Grenoble – HTX laboratory (Márquez Team)

Technical specifications

 Performances
Harvesting time 1 crystal/min
Success rate >99 %
Laser cutting No heat transfer
Micrometric precision
Multiple cut profiles
(pre- & user-defined)
Crystal soaking process 500 µL reservoirs
film piercing: photo ablation
 Software
User interface Windows / JAVA
Web based crystal selection CRIMS, customizable
Multi-device servers TINE, TANGO, EPICS
Sample changer robot Robot hardware & software integration
Power
230 VAC; 50 Hz <3680 W
110 VAC; 60 Hz <3680 W
Dimensions (X,Y,Z)
1.0 x 1.0 x  2.3 m3