Simple Method of Confirming Wet Environment for Soft X-ray Microscopy to Observe Hydrated Biological Specimens

Observations of hydrated specimens were carried out by soft X-ray microscopy using a newly developed wet-environment sample holder. The sample holder consisted of two silicon nitride membranes, one of which was printed with a carbon-based spacer. A reverse-contrast image of the spacer was obtained at two different wavelengths on opposite sides of the carbon K absorption edge. By obtaining a reverse-contrast image of the spacer used in the sample holder, one can confirm the wet environment of the specimen. The general principles and efficiency of this novel method are discussed.