(A and B) Western blot analysis of cultured HBMSCs with MoAb 103B2/9E10 and 105.A5. Membrane preparations from cultured HBMSCs were separated by 7.5% SDS-polyacrylamide gel electrophoresis under nonreducing and reducing conditions and transferred to nitrocellulose. The filters were successively incubated with either the 103B2/9E10 (A) or 105.A5 MoAb (B) supernatant (or isotype-matched, nonbinding controls) and antimouse-HRPO. The immunoreactive proteins were detected by ECL as described in Materials and Methods. Under nonreducing conditions, MoAbs 103B2/9E10 and 105.A5 identify two differentially migrating species with an apparent molecular weight of 80 and 160 kD (A and B, lane 1). However, upon reduction, the lower migrating 80-kD band is the major immunoreactive protein (A and B, lane 3). (A) Lane 1, nonreduced: 103B2/9E10 MoAb; lane 2, nonreduced: IgG₃negative control; lane 3, reduced: 103B2/9E10 MoAb; lane 4, IgG₃ negative control. (B) Lane 1, nonreduced: 105.A5 MoAb; lane 2, nonreduced: IgM negative control; lane 3, reduced: 105.A5 MoAb; lane 4, IgM negative control.