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The development of miniaturized electronics has led to the design and construction of powerful experimental platforms capable of measuring electronic properties to the level of single molecules, along with new theoretical concepts to aid in the interpretation of the data. A new area of activity is now emerging concerned with repurposing the tools of molecular electronics for applications in chemical and biological analysis. Single-molecule junction techniques, such as the scanning tunnelling microscope break junction and related single-molecule circuit approaches have a remarkable capacity to transduce chemical information from individual molecules, sampled in real time, to electrical signals. In this Review, we discuss single-molecule junction approaches as emerging analytical tools for the chemical and biological sciences. We demonstrate how these analytical techniques are being extended to systems capable …