Binding of Alexa633-labeled DNP-HSA to TNP-specific Ig-fLCs bound to mast cells.

Murine bone marrow-derived mast cells (BMMC) were incubated with TNP-specific Ig-fLC (6 µg) or IgE (1 µg). After incubation, Alexa633-labeled DNP-HSA (300 ng) was added. Administration of Alexa633-labeled DNP-HSA resulted in increased binding to BMMCs sensitized with TNP-specific Ig-fLC (B, middle) or IgE (B, right) as compared to non-sensitized BMMC (B, left panel). To evaluate whether the binding of A633-DNP-HSA was specific, a 30-fold excess of unlabeled DNP-HSA (10 µg) was administrated simultaneously with the Alexa633 labeled-DNP-HSA. This resulted in a detectable loss in binding to Ig-fLC- and IgE-sensitized BMMCs (C). Panel A depicts untreated BMMC. Results are representative for 3 independent experiments.