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Shinjo F , Hardy RR , Jongstra J
Monoclonal Anti-Lambda-5 Antibody Fs1 Identifies a 130 Kda Protein Associated with Lambda-5 and V(Pre-B) on the Surface of Early Pre-B-Cell Lines
International Immunology. 1994 Mar;6(3) :393-399
AbstractmAbs specific for mouse lambda5 protein were prepared by fusion of spleen cells from a hamster immunized with recombinant lambda5 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 A-1 and C-7 by phycoerythrin (PE)-conjugated FS1 (FS1 - PE) can be blocked by preincubation of these cells with unconjugated FS1 antibody or with affinity purified polyclonal lambda5 specific Ig but not with normal hamster or mouse IgG or with affinity purified polyclonal anti-Mb-1 Ig. From these experiments we concluded that FS1 specifically recognizes lambda5 protein. We used FS1 - PE to probe for surface (s) lambda5+ cells in normal BALB/c mouse bone marrow. Such cells were undetectable when total bone marrow or FACS sorted subpopulations were analyzed. However, when B220+, CD43+, slambda5- bone marrow cells were cultured for 4 days on the stromal cell line FLST2 in the presence of IL-7, slambda5 expression became apparent. Further expansion of these cells in IL7 alone augmented the slambda5 expression to readily detectable levels. This modulation may indicate that slambda5 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 lambda5 protein on the surface. Alternatively the binding of our FS1 mAb to the slambda5 molecules on normal bone marrow cells may be blocked by other proteins binding to the slambda5 complex in vivo and directly ex vivo. Previous analysis of surface lambda5 associated proteins on early mouse pre-B cell lines using a polyclonal anti-lambda5 rabbit antiserum had suggested that slambda5 protein was associated with a high molecular weight protein. Analysis of lambda5 and its associated proteins on early pre-B cell lines using our FS1 mAb confirmed our previous finding and showed that the early lambda5 receptor contains at least three proteins: lambda5, V(pre-B), and an as yet uncharacterized protein with a molecular weight of 130,000 designated p130.
NotesEnglish Article NC479 INT IMMUNOL