Acoustofluidic protein crystalligraphy

Serial crystallography (SX) has become one of the standard techniques to obtain time-resolved high-resolution structural information from microcrystals of various proteins and enzymes. Nevertheless, reliable sample delivery and fast mixing are often the primary limiting factors in time-resolved SX.

We propose to make use of acoustofluidics in combination with state-of-the-art 3D etching of fused silica and 3D printing of injection nozzles to enable mix-and-inject with sub-millisecond resolution to greatly improve the reliability, efficiency, and capability of SX.

We will use a piezoelectric actuator to create a 2D ultrasonic standing wave perpendicular to the crystal flow, focusing the crystals into a single line and enabling the crystals to pass through the fluid interface when two laminar flows are used.

The acoustofluidic cell can then be integrated in a 3D-printed design for dual flow-focusing nozzles or directly connected to a liquid flow cell.

Additional functionality can be built into the acoustofluidic chip, such as concentration enrichment, size selection and crystal bunching.

We would integrate the setup at several beamlines (MicroMAX, MAX IV; P11, PETRA III; SPB/SFX, EuXFEL) to perform time-resolved SX on HEW lysozyme, proteinase K, and NSP3 mac1, thus benchmarking the acoustofluidic device.

This would allow for the use of acoustofluidics beyond expert users and lead to the ability to reveal enzymatic reactions in real time without photoactivation.