An exact polystyrene-graft-polyisoprene (PS-g-PI) synthesized by iterative anionic polymerization and graft reaction using diphenylethylene functional groups was characterized by liquid chromatography. As-prepared graft copolymer contains various byproducts other than the target PS-g-PI. After the fractionation by size exclusion chromatography (SEC), the PS-g-PI appears quite homogeneous by SEC analysis, but the temperature gradient interaction chromatography (TGIC) separation with respect to the PI branch number showed a significantly wide distribution in the number of PI grafts. The resolved TGIC peaks were fractionated, and the molecular weight of each block (backbone or branches) was estimated by liquid chromatography at critical condition (LCCC) analysis at the critical adsorption condition of the opposite block. The composition determined by LCCC analysis and H-1 NMR analysis yielded a reasonably self-consistent result. Through this study, we demonstrated that SEC analysis of this type of branched copolymers can lead to erroneous result and needs to be done with proper caution.