In situ spectroelectrochemical fluorescence microscopy for studying electrodes modified by molecular adsorbates

The complexity of the electrochemical interface when modified by molecular adsorbates can be significant, especially those that are used for biosensing (e.g., nucleic acids, proteins). Characterizing these sensor surfaces typically involves using many in situ and ex situ methods. Studying the biomolecule-modified interface in aqueous buffer is necessary and can be facilitated through the use of fluorophore labeling and microscopy. Fluorescence microscopy has been developed for studying biological samples with constantly improving spatial and temporal resolutions. Adapting these methods for the study of electrochemical interfaces provides unique opportunities to study the structure and dynamic behavior of the adsorbed molecules. Outlined are important considerations for implementing fluorescence microscopy in electrochemical environments along with a brief review of various microscopy techniques and their use for in situexamination of the interface. Far-field and confocal imaging as well as single molecule localization microscopy are briefly described for study of adsorbed structures on the surface resulting in resolutions that span many orders of magnitude (mm to nm). Characterizing the local chemical environment of the adsorbed layers using Förster energy transfer and fluorescence lifetime imaging are discussed. These advanced approaches to imaging and characterizing electrochemical interfaces reveal more than the average behavior, providing the ability to question the influence of the variety of possible adsorbed structures on the performance of the interface.