A Cryo-EM Grid Preparation Device for Time-Resolved Structural Studies

Abstract

Structural biology generally provides static snapshots of protein conformations that can inform on the functional mechanisms of biological systems. Time-resolved structural biology provides a means to visualise, at near-atomic resolution, the dynamic conformational changes that macromolecules undergo as they function. Recent advances in the resolution obtainable by electron microscopy (EM) and the broad range of samples that can be studied makes it ideally suited to time-resolved studies. Here we describe a cryo-electron microscopy grid preparation device that permits rapid mixing, voltage assisted spraying, and vitrification of samples. We show that the device produces grids of sufficient ice quality to enable data collection from single grids that results in a sub 4 [A] reconstruction. Rapid mixing can be achieved by blot and spray or mix and spray approaches with a delay of ~10 ms, providing greater temporal resolution than previously reported approaches.