Cryofixation during live-imaging enables millisecond time-correlated light and electron microscopy.

Researchers seek to link cellular functions to their smallest structural components. Currently this requires correlation of two imaging techniques, live imaging and electron microscopy. Current correlative methods, however, have limited time resolution due to the sample preparation procedures for electron microscopy. Following live imaging, samples are transferred from the light microscope to a cryofixation, or ultra-fast freezing, instrument. The biological process progresses until the sample freezes, 1 second or more after the last live image. In this work, samples are cryofixed directly within the light microscope field of view. By eliminating the transfer step, time correlation between light and electron microscopy images of our samples is limited only by the freezing rate to the order of milliseconds rather than seconds.