A high-quality crystalline sample is critical for performing an experiment at LCLS. The requirements for preparing such a sample may be different from growing crystals for an experiment at the synchrotron.
From sequence to structure
In order to determine the three-dimensional structure of a macromolecule, the molecule of interest is first produced and purified using molecular biology techniques. When the macromolecule is of sufficient purity, it can be used for crystallization experiments.
Many different crystallization screens are available to quickly scan for conditions under which the molecule of interest starts to nucleate and grow into a crystal. Any crystals can then be fished from the crystallization solution and used to measure the X-ray diffraction, for example at a synchrotron light source.
Preparing for a serial crystallography experiment
Serial crystallography experiments at X-ray free electron lasers (XFELs) such as LCLS have unique requirements. Because of the highly intense X-ray pulse the XFEL delivers to the sample, crystals are destroyed after taking a single measurement, and the sample must be replenished before the next X-ray pulse arrives. Instead of growing a single large crystal, relatively large volumes of sample are used which contain many small-sized crystals.
Several crystallization methods are available, many of them adapted from the tested and proven protocols for growing the large crystals studied at the synchrotron light source.
A description of the following methods can be found on the crystal growth page:
- Vapor diffusion crystallization
- Simple batch crystallization
- Rapid-mixing batch crystallization
- Free interface diffusion crystallization
- Crystallization by concentration
- Lipidic cubic phase crystallization
It can be challenging to properly characterize serial crystallography samples because the crystals are often only a few micrometers to hundreds of nanometers in size.
Several methods used for crystalline sample characterization are described in detail on the characterization page, including:
- Light microscopy
- Fluorescence microscopy
- Second-order non-linear imaging of chiral crystals
- Reversibility of crystallization
- Dynamic light scattering
Lastly, an appropriate sample delivery method should be selected. Many different methods are available at LCLS, which are described in the sample delivery section.