Dimethyl sulfoxide (DMSO), a well-known differentiation inducer in several myeloid cells, induces G1 phase arrest in many cell lines. In this study, we investigated the possibility of using DMSO to arrest H18 hybridoma cells to the G1 phase and monitor whether the arrest improves antibody production. We showed that DMSO in concentration ranging between 0.3% and 0.6% efficiently arrested H18 hybridoma cells in G1 phase. In our experiment, > 80% of cells grown for 36h in presence of the 0.6% DMSO were arrested in G1. Furthermore, expression levels of P27 were up-regulated tow fold during the G1 phase. Higher concentration of DMSO at 0.9% leads to cytotoxicity. Herein we show a simple way, a two-stage process for antibody production, which consists of a proliferation phase leading to the desired cell density, followed by an extended production phase during which the cells remain at G1 phase. Our observation that the addition of DMSO results in increase antibody production is of significance in further use of hybridoma cells in high density large scale cell culture.