The Time-Resolved BioSAXS (TR-SAXS) sample environment at P12 provides several options for investigating rapid structural changes in biological macromolecules using SAXS.

These setups allow for precise time-resolved measurements to capture dynamic processes in solution under native-like conditions, allowing users to capture and analyze structural transitions that occur over short timescales.

Access to TR-SAXS is possible via a designated project proposal, and users are granted designated beamtime.

Users are strongly encouraged to contact the beamline staff to discuss the project.

Unless the user has pre-existing SAXS data of the system of interest in equilibrium, often users are asked to perform or mail-in their samples for Automated SAXS before TR-SAXS.

We provide the following modes for TR-SAXS:

Stopped-flow mixing SAXS measurements (SFM-SAXS) are performed at P12 using a Biologic mixing device coupled with an X-ray observation head.

This device operates in air and enables fast mixing of solutions, allowing for studying biomolecular interactions, conformational changes, and folding events over millisecond timescales.

The main SFM instrument is the µSFM (https://www.biologic.net/products/%c2%b5sfm/), which can mix ratios up to 1:9, and sample consumption can be as little as 2 µl.

For larger sample volumes and setups that require the use of multiple pistons, we also offer the SFM-400 mixing device (https://www.biologic.net/products/sfm2000-3000-4000-stopped-flow/). 

• Mixing and reaction times: The mixing ratio ranges from 1:1 to 1:9. The µSFM allows precise control over the reaction start time, with dead times as low as 1-2 milliseconds. 
• Sample exposure: The X-ray observation head observes the sample directly in the SAXS beam after mixing, enabling the acquisition of SAXS frames within microsecond to second-time ranges. Time points can be varied depending on the nature of the experiment.
• Volume and concentration: Reaction volumes typically range from 15 µL to 100 µL per sample, and sample concentrations of 1-10 mg/mL are recommended, depending on the biomolecule of interest.

• Temperature control: Experiments can be conducted at ambient room temperature or with temperature control upon request (4-55 °C)

Continuous (diode) and pulse lasers are available at P12. Both lasers could be connected to the sample exposure unit via an optical fiber, allowing for laser excitation of the sample in the SAXS beam path. 

A set of laser diodes with different wavelengths and powers is provided for continuous sample illumination. Please check with your local contact whether a diode with the required wavelength is available.

For pulse laser illumination,  an EKSPLA NT350 tunable laser is used: 

• Wavelength and pulse duration: The EKSPLA tunable laser offers wavelengths ranging 335-500 nm, 532 nm, 665-2600 nm, making it versatile for a wide range of photoactive molecules. Pulse durations are typically 5 ns, allowing for precise reaction initiation.
• Laser intensity: The laser intensity can be adjusted according to the specific experiment, with pulse energies ranging from 1 to 50 mJ depending on the wavelength.