Cyber-ostracism threatens adolescents' socialization and the reverse may also matter. However, little attention has been paid to the bidirectional relationship between cyber-ostracism and prosocial behaviors in adolescents, along with the underlying mechanisms. Therefore, this study examined the longitudinal association between cyber-ostracism and prosocial behaviors, as well as the role of rumination. A total of 716 Chinese adolescents completed the questionnaires at three time points during six months. Adolescents reported on cyber-ostracism, prosocial behaviors, rumination and demographic characteristics at each time point. The random-intercept cross-lagged panel model (RI-CLPM) was carried out. Results from RI-CLPM analysis suggested that cyber-ostracism predicted adolescents' prosocial behaviors negatively across time at the within-person level. Moreover, the RI-CLPM analysis further suggested that rumination mediated the longitudinal association between cyber-ostracism and prosocial behaviors at the within-person level. Specifically, the findings suggest that cyber-ostracism contributes to increased rumination among adolescents, subsequently leading to reduced engagement in prosocial behaviors. Our findings uncover the effect of cyber-ostracism on adolescents' prosocial behaviors and the mediating role of rumination at the within-person level.