Drop on fixed target reaction initiation approach for serial and time resolved crystallography

Kamps, J. J. A. G.; Hinchliffe, P.; Glerup, J.; Freeman, E. I.; Lang, P. A.; Tooke, C. L.; Beer, M.; Parkinson, L.; Gu, D.-H.; Park, S.; Devenish, N.; Zhou, T.; Shilova, A.; Rabe, P.; Schofield, C. J.; Spencer, J.; Park, J.; Owen, R.; Orville, A. M.; Aller, P.

doi:10.64898/2025.12.17.694847

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bioRxiv · 10.64898/2025.12.17.694847

Drop on fixed target reaction initiation approach for serial and time resolved crystallography

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Abstract

We describe the design and implementation of a drop on fixed target method for time-resolved serial crystallography at both synchrotron and XFEL facilities. A piezoelectric droplet dispensing pipette is employed for addition of picolitre volume (40 - 90 pL) aqueous droplets, containing (co-)substrate(s) or ligand(s), onto enzyme microcrystals immobilised on a solid support. The system was tested with various enzyme systems, including lysozyme and two -lactamases, CTX-M-15 and AmpCEC. Mitigation strategies for cross-well contamination, including the implementation of interleaved controls, are described; the overall performance of the system at synchrotron and X-ray free electron laser facilities was evaluated. This drop on fixed target method is a reliable framework for time-resolved crystallography and will improve the consistency of measurements across facilities.

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Kamps, J. J. A. G., Hinchliffe, P., Glerup, J., Freeman, E. I., Lang, P. A., Tooke, C. L., Beer, M., Parkinson, L., Gu, D.-H., Park, S., Devenish, N., Zhou, T., Shilova, A., Rabe, P., Schofield, C. J., Spencer, J., Park, J., Owen, R., Orville, A. M., Aller, P.. 2025-12-19. Drop on fixed target reaction initiation approach for serial and time resolved crystallography. https://doi.org/10.64898/2025.12.17.694847

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