Updated project metadata. New cysteine probes, N-acryloylindole-alkynes (NAIAs), have been developed. We have applied NAIA-5 for cysteine profiling in HepG2 cell lysates, and the results have been compared to the conventional cysteine probe iodoacetamide-alkyne (IAA). NAIA-5 allows identification of more ligandable Cys than IAA in our experiment, and more interesting, a unique pool of Cys which have not been profiled previously by any reported cysteine probe. We have further utilized NAIA-5 for covalent ligand screening experiment. This allows us to discover hit compounds targeting the newly identified cysteines and proteins. One of the hit compounds, CL1, demonstrated good bioactivity by arresting cell cycle at G1 phase through targeting proteins associated with cell cycle and regulations. In addition, NAIA-5 has been successfully coupled with NAI compound for studying cysteine oxidative modifications in HepG2 cells by MS experiments.