Time-resolved cryotransmission electron microscopy.

Abstract

We describe a new technique, time-resolved cryotransmission electron microscopy (TRC-TEM), that can be used to study changes in microstructure occurring during dynamic processes such as phase transitions and chemical reactions. The sample is prepared on an electron microscope grid maintained at a fixed temperature in a controlled atmosphere. The dynamic process is induced on the grid by a change in pH, salt, or reactant concentration by rapid mixing with appropriate solutions. Alternatively, induction is by rapid change of specimen temperature, or by controlled evaporation of a volatile component. We call such procedures on-the-grid processing. The dynamic process is permitted to run for a defined time and then the thin-film specimen is thermally fixed by plunging into liquid ethane at its freezing point, producing a cryotransmission electron microscopy specimen. By repeating this procedure with varying delays between induction and sample fixation, we can observe transient microstructures. We demonstrate the use of TRC-TEM to study the intermediate structures that form during the transitions between L alpha, III, and HII liquid crystalline phases in phospholipid systems. We also identify several other possible applications of the technique.