# Spitrobot-2 advances time-resolved cryo-trapping crystallography to under 25 ms

## TL;DR

Spitrobot-2 is a faster, more user-friendly system for capturing protein crystal structures in real-time, enabling detailed studies of molecular processes.

## Contribution

Spitrobot-2 reduces cryo-trapping delay to 23 ms and improves automation for easier sample handling.

## Key findings

- Spitrobot-2 achieves a cryo-trapping delay time of 23 ms, doubling the speed of the previous version.
- 12 crystal structures of three model systems show successful cryo-trapping with conformational changes and ligand binding observed within 25 ms.

## Abstract

We previously introduced the spitrobot, a protein crystal plunging system that enables reaction quenching via cryo-trapping with a time resolution in the millisecond range. Here we present the next generation, spitrobot-2, as an integrated benchtop device. User-friendliness has been improved by semi-automatic sample exchange. Moreover, a fully automated shutter shields the liquid nitrogen from the humidified environment, improving sample integrity. Most importantly, the cryo-trapping delay time has been reduced to 23 ms, making spitrobot-2 twice as fast as the previous generation. This further expands the number of target systems that can be addressed by cryo-trapping time-resolved crystallography. Using 12 crystal structures of three independent model systems, we demonstrate successful cryo-trapping via observation of conformational changes and ligand binding within 25 ms. These improvements increase the convenient access to cryo-trapping, time-resolved X-ray crystallography empowering the MX community with efficient tools to advance research in structural biology.

Spitrobot is a protein crystal plunging system that enables reaction quenching through cryo-trapping with millisecond time resolution. Here, the authors present Spitrobot 2 as an integrated benchtop device with improved net sample preparation time and cryo-trapping delay time, increasing the number of target systems that can be studied using cryo-trapping time-resolved crystallography.