Antigen Valency as a Determinant of the Responsiveness of IgE-Sensitised Rat Basophil Leukemia Cells

Abstract

Rat basophil leukemia (RBL) cells were sensitised with varying proportions of monoclonal IgE anti-ovalbumin (OVA) and anti-DNP antibodies, and serotonin release was measured after challenge with aggregated OVA or dinitrophenylated human serum albumin (DNP-HSA). Highly aggregated OVA was shown to provoke the degranulation of RBL cells that had been sensitised with an IgE preparation containing 2% IgE anti-OVA antibodies. Highly substituted DNP₃₂-HSA induced degranulation of RBL cells sensitised with just 0.5% antigen-specific IgE. When cells were sensitised with high percentages of specific IgE, maximum degranulation was seen at concentrations of 2 μg/ml (aggregated OVA) and 50 ng/ml (DNP-HSA), while moderate degranulation was still seen at antigen concentrations as low as 50 and 2 ng/ml, respectively. Low-molecular weight aggregates of OVA and low-valency DNP₄-HSA only stimulated degranulation when high percentages of RBL Fcε receptor were occupied by antigen-specific IgE. The sensitising abilities of two anti-DNP monoclonal antibodies of differing affinities were compared. When challenged with low-valency antigen, only cells sensitised with the higher-affinity monoclonal antibody exhibited moderate levels of degranulation. Degranulation required exposure to high antigen challenge doses (5 μg/ml). Cells sensitised with either monoclonal antibody responded strongly when challenged with a wide range of concentrations (1–250 ng/ml) of high-valency DNP₃₂-HSA, although greater sensitivity was always seen with the higher-affinity antibody. These results suggest that antigen valency is a critical parameter for mast cell function, and that low-affinity antibody may be capable of sensitising mast cells to high-valency antigen.

Several distinct properties of the IgE repertoire determine effector cell degranulation in response to allergen challenge

“…A prominent role of low-affinity IgE antibodies for allergic symptoms could very well be in the context of allergenic cross-reactivity, where patients react with allergic symptoms when challenged with homologous allergen molecules from related species. 18 The requirements for degranulation of effector cells in vitro have previously been addressed by different means, including sensitization with complex sera, 7,19,20 cross-linking with polymerized antigens/allergens, [21][22][23][24] cross-linking with covalently polymerized IgE, 25,26 cross-linking with allergen extracts, 7 and use of rat effector cell lines, [21][22][23][24][25]27 in addition to a variety of other model systems. To our knowledge, the present study is the first in which factors governing human effector cell degranulation have been demonstrated in a model system consisting of an intact, clinically relevant allergen in combination with different mixtures of well-characterized, allergen-specific IgE antibodies directly mimicking allergic patientsÕ sera of different compositions.…”

Analysis of IgE Antibodies from a Patient with Atopic Dermatitis: Biased V Gene Usage and Evidence for Polyreactive IgE Heavy Chain Complementarity-Determining Region 3

“…The interaction between surface IgE and allergen is the key to unleashing the mast cell arsenal of proinflammatory mediators. It has been shown in vitro, using the rat basophil leukemia cell line, that degranulation is dependent upon the valency of the Ag (65,66). If polyreactive IgE Abs exist, Fc⑀RI aggregation of mast cell-and basophil-bound IgE could more easily occur.…”

A role for the hepatobiliary system in IgE‐mediated intestinal inflammation in the rat

“…Murine IgE antidinitrophenyl (DNP) antibodies were raised as ascites using the H‐1e‐DNP cell line, and IgE antiovalbumin (OVA) antibodies were raised as ascites using the OVA6B cell line. IgE antibody concentrations were quantified by ELISA assay and by rat basophil leukaemia cell degranulation assay as previously described [13]. Groups of rats were sensitized by i.v.…”