In Situ Cryo-Electron Tomography and Advanced Micromanipulator Techniques

In situ cryo-electron tomography is undergoing a revolution as more challenging samples are moving into the center of attention. The complexity of the targeted structures is rapidly expanding and requires that preparation techniques evolve to keep up with the growing demand for larger specimens. In particular, organoids and tissue, but also small animals that can be vitrified in whole by high-pressure freezing, are targets of this new branch of in situ cryo-electron microscopy, which seeks to provide a “biopsy at the nanoscale”. With its recent coming of age, cryo-lift out is responding to this call. Novel hardware now makes this preparation technique commercially available to a broad audience without the need for significant prior knowledge or experience. Thanks to this, cryo-lift out is becoming increasingly popular and, through the concerted effort of the research community, more versatile and higher throughout. The following chapter provides a topical overview of this new and rapidly developing technique. We introduce the basic principles of cryo-lift out and briefly summarize the technological and biological results so far.