X-CHIP: an integrated platform for high-throughput protein crystallization and on-the-chip X-ray diffraction data collection

Abstract

The X-CHIP (X-ray Crystallization High-throughput Integrated Platform) is a novel microchip that has been developed to combine multiple steps of the crystallographic pipeline from crystallization to diffraction data collection on a single device to streamline the entire process. The system has been designed for crystallization condition screening, visual crystal inspection, initial X-ray screening and data collection in a high-throughput fashion. X-ray diffraction data acquisition can be performed directly on-the-chip at room temperature using an in situ approach. The capabilities of the chip eliminate the necessity for manual crystal handling and cryoprotection of crystal samples, while allowing data collection from multiple crystals in the same drop. This technology would be especially beneficial for projects with large volumes of data, such as protein-complex studies and fragment-based screening. The platform employs hydrophilic and hydrophobic concentric ring surfaces on a miniature plate transparent to visible light and X-rays to create a well defined and stable microbatch crystallization environment. The results of crystallization and data-collection experiments demonstrate that high-quality well diffracting crystals can be grown and highresolution diffraction data sets can be collected using this technology. Furthermore, the quality of a single-wavelength anomalous dispersion data set collected with the X-CHIP at room temperature was sufficient to generate interpretable electron-density maps. This technology is highly resourceefficient owing to the use of nanolitre-scale drop volumes. It does not require any modification for most in-house and synchrotron beamline systems and offers a promising opportunity for full automation of the X-ray structuredetermination process.

Polarization-resolved second-harmonic generation microscopy as a method to visualize protein-crystal domains

“…Crystalline samples with multiple domains are not always easily identifiable by bright-field imaging, especially for the specific case of twinning. Consequently, rapid and nondestructive identification of crystalline domains could significantly improve the productive throughput of synchrotron facilities (Chayen & Saridakis, 2008;Santarsiero et al, 2002;Walter et al, 2003;Chayen, 2003;Bergfors, 2003;Stojanoff et al, 2011;Kisselman et al, 2011).…”

Microfluidics: From crystallization to serial time-resolved crystallography

“…169 It is also possible to mount crystals grown directly on the mount, as in the case of the X-CHIP, where pinned droplets are protected against dehydration by a thin coating of oil (Figure 8(c)) 186,187 or the aforementioned graphene-based microfluidic devices. 123 Such samples can be cryocooled, protected by a capillary sheath or other materials, or analyzed directly using a traditional goniometer setup for sample manipulation.…”

A Graphene-Based Microfluidic Platform for Electrocrystallization and In Situ X-ray Diffraction

“…Such platforms have been increasingly harnessed to facilitate the diffraction analysis of challenging targets for both static and dynamic structure determination. Various platforms have been developed to improve the growth and subsequent mounting of tiny and fragile crystals for X-ray diffraction analysis [24][25][26][27][28], including dense array-style devices [29][30][31][32][33][34][35][36][37][38][39][40][41], platforms for the lipidic cubic phase crystallization of membrane proteins [42,43], and thin-film sandwich devices [44]. In the meantime, the challenges of such platforms lie in the need to maintain a protected sample environment, as well as minimize the interference of device materials with the subsequent X-ray analysis.…”