Identification of basophils by a mAb directed against pro-major basic protein 1

Background: Basophils possess characteristics of both mast cells and eosinophils, and all 3 cell types often are found together, particularly during allergic reactions. A mAb (J175-7D4) generated against the recombinant pro-form of human eosinophil granule major basic protein 1 (rproMBP1) appeared to stain only basophils in tissue specimens. Objective: We investigated J175-7D4 to characterize its specificity for basophils. Methods: Fluid-phase immunoprecipitation, Western blotting, and immunocytochemistry and immunohistochemistry were used to establish the specificity of J175-7D4. Results: First, J175-7D4 binds to various glycosylated and proteolytically processed forms of rproMBP1, but not to major basic protein. Second, cells transfected with the rproMBP1 gene and human placental tissue (known to express the pro-form of major basic protein 1 [proMBP1]) stain specifically with J175-7D4. In contrast, although mature eosinophils contain substantial major basic protein, they lack proMBP1 and do not stain. Neutrophils, lymphocytes, monocytes, and skin mast cells also are not stained. However, blood basophils are stained by J175-7D4, anti-IgE, Wright-Giemsa (metachromatically), and a previously characterized basophil-specific mAb, 2D7. Finally, formaldehyde-fixed, paraffin-embedded basophils are identically detected by J175-7D4 and 2D7, and J175-7D4 also recognizes putative basophils in formaldehyde-fixed., paraffin-embedded tissue specimens from inflammatory dermatoses, such as atopic dermatitis and delayed pressure urticaria. Conclusion: The J175-7D4 mAb recognizes proMBP1 as a novel marker for human basophils.J175-7D4 should prove useful for characterizing basophil involvement in human health and disease.