Updates on the surface antigens of basophils: CD16 on basophils of patients with respiratory or insect venom allergy and the rejection of CD203c and CD63 externalization decoupling by bisindolylmaleimides

Summary Background CD 16 was previously suggested to be a new marker of basophils that is subject to downregulation by Fcε RI crosslinking. Certain compounds, including supraoptimal concentrations of the PKC inhibitors, bisindolylmaleimides, decouple the release of granules containing CD 203c, CD 63 and histamine, and may thus help to identify the mechanisms related to the CD 16 externalization. Objective We hypothesized that CD 16 is differentially expressed on the surface of basophils in patients with birch pollen or insect venom allergy and is subject to a regulation in response to allergens. We also employed CD 203c and CD 63 externalization decoupling by bisindolylmaleimides. Methods We performed a basophil activation test coupled with CD 16 and histamine detection using cells isolated from patients with allergy to birch pollen or insect venom and negative controls. We employed two PKC inhibitors, bisindolylmaleimide II and Ro 31‐8220 at their supraoptimal concentrations and, after difficulties reproducing previously published data, we analyzed the fluorescence of these inhibitors alone. We identified the CD 16 isoforms by sequencing nested RT ‐ PCR amplicons from flow cytometry sorted basophils and by cleaving the CD 16b GPI anchor using a phospholipase C. Results We provide the first evidence that CD 16a is expressed as a surface antigen on a small subpopulation of human basophils in patients with respiratory and insect venom allergy, and this antigen shows increased surface expression following allergen challenge or Fcε RI crosslinking. We rejected the apparent decoupling of the surface expression of basophil activation markers following the administration of bisindolylmaleimides. Conclusions & Clinical Relevance The inclusion of α CD 16 in negative selection cocktails selects against a subset of basophils that are CD 16 + or CD 16 dim . Using CD 16 dim basophils and unstained leucocytes, we show that previous studies with supraoptimal concentrations of bisindolylmaleimides are likely flawed and are not associated with the differential expression of CD 203c and CD 63.