Compared with batch and vapor diffusion methods, counter diffusion can generate larger and higher-quality protein crystals yielding improved diffraction data and higher-resolution structures. Typically, counter-diffusion experiments are conducted in elongated chambers, such as glass capillaries, and the crystals are either directly measured in the capillary or extracted and mounted at the X-ray beamline. Despite the advantages of counter-diffusion protein crystallization, there are few fixed-target devices that utilize counter diffusion for crystallization. In this article, different designs of user-friendly counter-diffusion chambers are presented which can be used to grow large protein crystals in a 2D polymer microfluidic fixed-target chip. Methods for rapid chip fabrication using commercially available thin-film materials such as Mylar, propylene and Kapton are also detailed. Rules of thumb are provided to tune the nucleation and crystal growth to meet users' needs while minimizing sample consumption. These designs provide a reliable approach to forming large crystals and maintaining their hydration for weeks and even months. This allows ample time to grow, select and preserve the best crystal batches before X-ray beam time. Importantly, the fixed-target microfluidic chip has a low background scatter and can be directly used at beamlines without any crystal handling, enabling crystal quality to be preserved. The approach is demonstrated with serial diffraction of photoactive yellow protein, yielding 1.32 Å resolution at room temperature. Fabrication of this standard microfluidic chip with commercially available thin films greatly simplifies fabrication and provides enhanced stability under vacuum. These advances will further broaden microfluidic fixed-target utilization by crystallographers.
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This content will become publicly available on November 1, 2025
Design of an atomic layer deposition system with in situ reflection high energy electron diffraction
We report the design, fabrication, and testing of an atomic layer deposition (ALD) system that is capable of reflection high energy electron diffraction (RHEED) in a single chamber. The details and specifications of the system are described and include capabilities of RHEED at varied accelerating voltages, sample rotation (azimuthal) control, sample height control, sample heating up to set temperatures of 1050 °C, and either single- or dual-differential pumping designs. Thermal and flow simulations were used to justify selected system dimensions as well as carrier gas/precursor mass flow rates. Temperature calibration was conducted to determine actual sample temperatures that are necessary for meaningful analysis of thermally induced transitions in ALD thin films. Several demonstrations of RHEED in the system are described. Calibration of the camera length was conducted using a gold thin film by analyzing RHEED images. Finally, RHEED conducted at a series of increasing temperatures was used to monitor the crystallization of an ALD HfO2 thin film. The crystallization temperature and the ring pattern were consistent with the monoclinic structure as determined by separate x-ray diffraction-based measurements.
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- Award ID(s):
- 1752956
- PAR ID:
- 10555734
- Publisher / Repository:
- AIP Publishing
- Date Published:
- Journal Name:
- Review of Scientific Instruments
- Volume:
- 95
- Issue:
- 11
- ISSN:
- 0034-6748
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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