The interaction of immunoglobulin E (IgE) with its low affinity receptor (FceRII/CD23) plays a central role in the initiation and regulation of type I hypersensitivity responses. We have previously identified the importance of amino acid residues in the A-B loop of the Ce3 domain of human IgE and implicated a region close to the glycosylation site at Asparagine 371 as contributing to IgE-CD23 interaction. These residues were now targeted by site directed mutagenesis. The IgE-CD23 interaction was assessed by semi-quantitative flow cytometry. Replacement of the entire Ce3 A-B loop (residues 341-356) with the homologous rat IgE sequence resulted in complete loss of human CD23 recognition, as did replacement of residues 346-353, indicating that class specific effector residue(s) are contained within these eight amino acids. Lysine 352 within the A-B loop was identified as contributing directly to human CD23 interaction, mutation to the rodent homologue Glycine, or Glutamate, resulted in a significant reduction in binding compared to native IgE, while conservative substitution with Arginine effected a small, but statistically significant, enhancement of CD23 binding. Mutation of the Ce3 glycosylation site at Asparagine 371 to Threonine or Glutamine did not significantly affect CD23 recognition. Our results yield new insights into the structural basis of the hIgE-CD23 interaction, and hold promise for the rational design of drugs that can manipulate IgE mediated regulation of the allergic response.