The cause of MS is uncertain, but an autoimmune disorder of the CNS is likely, and myelin basic protein (MBP) is a candidate antigen. MBP exists in different isoforms, generated by differential splicing of exons, and in charge isomers, generated by posttranslational modifications. Different isoforms and charge isomers presumably subserve different functions, and they vary in abundance in immature myelin found during myelinogenesis and remyelination compared with mature myelin. The 18.5-kd isomer is most abundant in normal human adults and consequently has been used almost exclusively for immunologic studies in MS. In the present study, we examined a different but abundant charge isomer of MBP, termed MBP-C8, to determine whether it could be recognized by MBP-specific cytotoxic and proliferative T-cell lines (TCL) and whether a T-cell response directed exclusively against citrulline-containing residues of MBP-C8 exists in MS patients and healthy controls. We showed that citrulline affects antigen recognition by some TCL that are specific for areas of MBP that contain the citrulline residues. Following stimulation with MBP-C8, MBP-C8-specific TCL could be generated from both MS patients and controls. T-cell responses against antigens that appear during myelinogenesis and during remyelination may be important in inducing and perpetuating an autoimmune response involved in the pathogenesis of MS.