A precision cryostat design for manual and semi-automated cryo-plunge instruments

Here we describe a bench-top cryostat system to control the temperature of liquid ethane in a cryo-plunge apparatus designed for biological specimen preparation for electron cryomi-croscopy. It comprises a foam insulated Dewar containing a copper cryostat cup, whose temperature is controlled via an active feedback system to within 0.1 K. The device can easily be incorporated into existing manual and semi-automatic cryo-plunge instruments that are not equipped with cryogenic temperature control. Over the course of normal use, we find that using a cryostat is convenient, fast, and does not require special mixtures of cryogens like ethane/propane. This simple cryostat improves the reliability and reproducibility of biological specimen preparation for electron cryomicroscopy.