Well-defined amphiphilic block copolymers, poly[poly(ethylene glycol) methyl ether methacrylate]-block-poly(2,5-dibromo-3-vinylthiophene) poly(PEGMA)-b-poly-(DB3VT), were synthesized by reversible addition-fragmentation chain transfer polymerization. A facile bottom-up modification was developed for preparing cross-linked core-shell nanoparticles from the amphiphilic block copolymers, in which an in situ Suzuki coupling reaction was carried out between 2,5-dibromide groups in the block copolymers and various diboronic acid compounds in the presence of a palladium catalyst after micelle formation in THF/H2O solution. The stable and uniform core shell structure was confirmed by scanning force microscopy and dynamic light scattering observations. Furthermore, the size of the cross-linked core shell nanoparticles could be controlled by adjusting the compositions of each segment in the block copolymers and Suzuki coupling reaction conditions. The relationship between the optical properties and chemical structures of pi-conjugated segments in the particles was clearly observed by changing the coupling agents (diboronic acid compounds). These results demonstrate that stable and uniform cross-linked core shell nanoparticles were successfully prepared from well-defined amphiphilic block copolymers, and tuning of their optical properties could be accomplished as well.