Hybridoma that produces rat anti-mouse interleukin 6 receptor (IL-6R) antibody, MR16-1, was established by the fusion of mouse P3U1 myeloma cells and spleen cells from mouse soluble IL-6R (sIL-6R)-immunized Wistar rat. In the present study, we examined the characteristics of MR16-1 in vitro and in vivo. MR16-1 bound to mouse sIL-6R dose-dependently. MR16-1 suppressed IL-6-induced proliferation of 7TD1 cells in a dose-dependent manner and this inhibitory effect was reversed by the addition of a higher concentration of IL-6. Cross-reactivity study using T cells from mouse, rat, and human revealed that MR16-1 did not cross-react with human and rat IL-6R. Binding region analysis using several human-mouse chimeric IL-6Rs showed that half of the fibronectin domain II of mouse IL-6R (amino acids 214-285) was required for MR16-1 binding. Furthermore, MR16-1 completely suppressed IL-6-induced antibody production in DNP-KLH immunized mice. These lines of evidence demonstrate that MR16-1 is useful to investigate the physiological and pathological roles of IL-6 and sIL-6R in mice.