A high-throughput screening strategy is described that involves the acquisition of two-dimensional 15N/1H correlation spectra in less than 10 min on 50 microM protein samples using cryogenic NMR probe technology. By screening at these concentrations, small organic molecules can be tested in mixtures of 100, which dramatically increases the throughput of the NMR-based assay. Using this strategy, libraries of more than 200 000 compounds can be tested in less than 1 month. There are many advantages of high-throughput NMR-based screening compared to conventional assays, such as the ability to identify high-affinity ligands for protein targets with no known function. This suggests that the method will be extremely useful for screening the large number of targets derived from genomics research.