mAbs specific for mouse λ5 protein were prepared by fusion of spleen cells from a hamster Immunized with recomblnant λ5 protein synthesized in bacteria and the mouse myeloma cell line SP2/0-Ag14. Here we report the characteristics of the antibodies produced by the FS1 hybridoma. FS1 antibody stains a variety of mouse pre-B cell lines but not B cell lines or T cell lines. The staining of the pre-B cell lines Awl and C-7 by phycoerythrin (PE)-con]ugated FS1 (FS1 - PE) can be blocked by prelncubatlon of these cells with unconjugated FS1 antibody or with affinity purified polyclonal λ5 specific Ig but not with normal hamster or mouse IgG or with affinity purified polyclonal antl-Mb-1 Ig. From these experiments we concluded that FS1 specifically recognizes λ5 protein. We used FS1 -PE to probe for surface (s) λ5+ cells in normal BALB/c mouse bone marrow. Such cells were undetectable when total bone marrow or FACS sorted subpopulatlons were analyzed. However, when B220plus;, CD43+, sλ5+ bone marrow cells were cultured for 4 days on the stromal cell line FLST2 in the presence of IL-7, sλ5 expression became apparent. Further expansion of these cells in IL7 alone augmented the sλ5 expression to readily detectable levels. This modulation may indicate that sλ5 expression on normal bone marrow cells in vivo is transient and that at any given moment only a small fraction of bone marrow cells expresses low levels of λ5 protein on the surface. Alternatively the binding of our FS1 mAb to the sλ5 molecules on normal bone marrow cells may be blocked by other proteins binding to the 8X5 complex in vivo and directly ex vivo. Previous analysis of surface λ5 associated proteins on early mouse pre-B cell lines using a polyclonal anti-λ5 rabbit antlserum had suggested that sλ5 protein was associated with a high molecular weight protein. Analysis of λ5 and Its associated proteins on early pre-B cell lines using our FS1 mAb confirmed our previous finding and showed that the early λ5 receptor contains at least three proteins: λ5, Vpre-B, and an as yet uncharacteiized protein with a molecular weight of 130,000 designated p130.
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Shinjo et al. (1994) studied this question.