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Soft Microscopy of Negative Stained Soft Materials: Balancing Dose Rate and Sample Damage

Electron energy loss spectroscopy (EELS) and energy-dispersive x-ray spectroscopy (EDX) in a scanning electron microscope (STEM) are promising methods for the elemental and electronic structural analysis of soft biological materials [1]. When preparing these soft materials for STEM, the sample preparation and imaging method impacts the information one can gain from the EELS/EDX spectrum. Here we ponder the advantages and disadvantages of using a stained sample for EELS/EDX analysis. Negative staining can increase the background in an EELS/EDX spectrum making an unstained sample preferable. However, staining provides morphological information that can give a clearer picture of the structure when paired with an EELS/EDX map. By imaging stained and unstained samples at room and cryogenic temperatures, we aim to tune the critical dose, impacting the quality of the EELS/EDX spectrum. This work analyzes the role that sample fixation may play in preparing soft biological samples for analytical microscopy techniques. The sample used in this work is a