Two monoclonal rat anti-mouse IgM antibodies, Bet 1 and Bet 2, are described in this paper. Bet 1 defines a new allotypic determinant, Igh-6.5, expressed on IgM molecules in serum and on B lymphocytes, whereas Bet 2 recognizes a determinant on IgM molecules of all mouse strains tested. Both reagents bind to the IgM myeloma protein MOPC104E, but not to IgG myeloma proteins, including FLOPC21, MOPC21, and UPC10. Using serum from various mouse strains to inhibit the binding of Bet 1 to MOPC104E, 3 distinct inhibition patterns were found. BALB/c, DBA/2, and CBA sera inhibited strongly, C56BL/6 (B6), SJL, AKR, and NZB sera inhibited weakly, and A and AL sera showed no inhibition of binding of BET 1 to MOPC104E. All sera tested were equivalent in their inhibition of the binding of Bet 2 to MOPC104E. When spleen cells from different mouse strains were reacted with fluorescein-conjugated Bet 1 (F-Bet 1) and subjected to flow microfluorometry analysis, 3 types of staining patterns, corresponding to those obtained with the serum inhibition assay, were also found. The determinant recognized by Bet 1 is controlled by a gene linked to the Igh-C gene complex. C.AL20 behaved like AL, and C.B20 and BAB/14 behaved like B6, both in the serum inhibition assay and in flow microfluorometry analysis of spleen cells stained with F-Bet 1. In addition, the capacity of serum from individual (BALB/c X B6) X B6 backcross progeny to inhibit the binding of Bet 1 to MOPC104E was linked to the expression of the Ig-1a marker.