Mass, charge, and subcellular localization of a unique secretory product identified by the basophil-specific antibody BB1

McEuen, Alan R., Calafat, Jero, Compton, Steven J., Easom, Nicholas J., Buckley, Mark G., Knol, Edward F. and Walls, Andrew F. (2001) Mass, charge, and subcellular localization of a unique secretory product identified by the basophil-specific antibody BB1. Journal of Allergy and Clinical Immunology, 107 (5), 842-848. (doi:10.1067/mai.2001.114650).

Abstract

Background: BB1 is a basophil-specific mAb (Lab Invest 1999;79:27-38). The identity of the corresponding antigen has not been determined, but it gives a granular appearance on staining and is secreted on activation of basophils.
Objective: We sought to further characterize the basophilspecific antigen identified by BB1.
Methods: Intracellular localization was determined by flow cytometry and by immunogold labeling and electron microscopy. Physical chemical properties were investigated by gel filtration chromatography and preparative isoelectric focusing.
Results: In flow cytometry, permeabilization of cells increased immunofluorescence 100-fold, confirming the predominantly intracellular localization of the antigen. It was further localized to the secretory granules by immunoelectron microscopy. Double labeling with a CD63-specific antibody demonstrated selective binding of BB1 to the granule matrix. Gel filtration chromatography indicated that the antigen is secreted as a complex of approximately 5 × 106 d, which was well resolved from the 210-kd supramolecular complex containing tryptase. The antigen was degraded by pronase. Isoelectric focusing indicated a highly basic protein with an isoelectric point of 9.6.
Conclusion: With its granule localization, release on cell activation, and unique properties, the antigen identified by BB1 could be a novel mediator of allergic disease. We propose the name basogranulin for this novel basophil-specific protein.

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Contributors

Author: Andrew F. Walls

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