Single-molecule techniques provide opportunities for molecularly precise imaging, manipulation, assembly and biophysical studies. Owing to the kinetics of bond rupture processes, rapid single-molecule measurements can reveal novel bond rupture mechanisms, probe single-molecule events with short lifetimes and enhance the interaction forces supplied by single molecules. Rapid measurements will also increase throughput necessary for technological use of single-molecule techniques. Here we report a nanomechanical sensor that allows single-molecule force spectroscopy on the previously unexplored microsecond timescale. We probed bond lifetimes around 5 μs and observed significant enhancements in molecular interaction forces. Our loading-rate-dependent measurements provide experimental evidence for an additional energy barrier in the biotin-streptavidin complex. We also demonstrate quantitative mapping of rapid single-molecule interactions with high spatial resolution. This nanomechanical interface may allow studies of molecular processes with short lifetimes and development of novel biological imaging, single-molecule manipulation and assembly technologies.