Several distinct properties of the IgE repertoire determine effector cell degranulation in response to allergen challenge (1)
Total and allergen-specific IgE concentrations, IgE affinity, and IgE clonality are all distinct properties of an allergic patient’s IgE repertoire. Most commonly seen are patient-specific variations in total and allergen-specific IgE concentrations, but also more subtle and less accessible parameters, such as overall IgE affinity for allergens and the number of allergen epitopes recognized by a patient’s IgE repertoire (clonality), have been connected with different outcomes of skin prick tests or severity of allergic symptoms
Ten murine anti-Der p 2 IgG antibodies were converted into mouse/human chimeric IgE antibodies named H1 through H8, H10, and H12. Relative positions of Der p 2 epitopes recognized by the rIgE clones were determined by the ability of pairs of 2 rIgE clones to bind rDer p 2 simultaneously. Nine different binding patterns were observed. Mutation of Der p 2 in position K15, H74, or K82 each led to abolished antibody binding by 1 or more rIgE clones. Contrarily, mutation of Der p 2 in position K6, H30, or E62 only weakly affected binding of some rIgE clones. Based on these 2 mapping approaches, a 3- and a 2-dimensional map of epitopes recognized by the various rIgE clones was constructed.
By shuffling the light chains of the 10 hybridoma-derived rIgE clones, 12 additional Der p 2–specific rIgE clones were obtained.
Individual rIgE antibody affinities for rDer p 2 were determined by means of surface plasmon resonance experiments.
An irrelevant non–Der p 2–specific rIgE antibody (anti-tox, binding tetanus toxoid) was included in the IgE mixture as well to imitate a typical allergic IgE repertoire. when increasing the total rIgE concentration, basophils sensitized with increasing relative concentrations of Der p 2–specific rIgE led to both increased maximal degranulation response and sensitivity.
The highest level of basophil degranulation was seen when the 2 rIgE clones were present in equimolar amounts (ratio of 50:50). Maximal basophil degranulation decreased when the ratio between the 2 Der p 2–specific rIgE clones was 95:5 and even more pronounced with a ratio of 99:1. Basophils monosensitized with a single Der p 2–specific rIgE (ratios of 100:0/0:100) showed no degranulation, as expected, because of the requirement for the presence of IgEs having at least 2 different epitope specificities for productive cross-linking. Basophils sensitized with 2 low-affinity rIgE clones (LL combination) required a 500- to 1000-fold higher rDer p 2 concentration than basophils sensitized with 2 high-affinity rIgE clones (HH combination) to reach equivalent degranulation levels. A low-affinity rIgE clone was combined with a high-affinity rIgE clone (HL combination). This latter combination showed a basophil sensitivity that was only 2- to 5-fold lower than that of 2 high-affinity rIgE clones (HH combination).
Basophils sensitized with 3 different rIgE clones (brown curve) showed 5- to 20-fold higher sensitivity than basophils sensitized with only 2 different rIgE clones.
The presence of only a single antibody of high affinity in the allergen-specific IgE repertoire is required for the recruitment of an IgE of even very low affinity into productive FcεRI/IgE/allergen complexes. An allergen initially becomes captured by a high-affinity FcεRI-bound “anchor” IgE and is then dragged over the cell surface until this complex encounters other, even low-affinity, FcεRI-bound IgEs causing efficient cross-linking and effector cell degranulation.