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Therapeutic antibodies have predominantly been IgG‐based. However, the ongoing clinical trial of MOv18 IgE has highlighted the potential of using IgE antibodies in cancer therapy. While extensive studies targeting IgG glycosylation resulted in a rational basis for the development of enhanced biotherapeutics, IgE glycosylation remains an area with limited analyses. Previous studies on the role of IgE glycosylation present conflicting data with one study emphasizing the importance of N275 and T277 residues for FcεRI binding whereas another asserts the nonsignificance of IgE glycosylation in receptor interaction. While existing literature underscores the significance of glycans at the N275 position for binding to FcεR1 receptor and initiation of anaphylaxis, the role of other IgE glycosylation sites in folding or receptor binding remains elusive. This study systematically investigates the functional significance of N‐linked glycosylation sites in the heavy chain of IgE which validates the pivotal role of N275 residue in IgE secretion and stability. Replacement of this asparagine to non‐amine group moieties does not affect IgE function in vitro, yet substitution with aspartic acid compromises antibody yield. The deglycosylated IgE variant exhibits superior efficacy, challenging the conventional importance of glycosylation for effector function. In summary, our study unveils an intricate relationship between N‐glycosylation sites and the structural–functional dynamics of IgE antibodies. Furthermore, it offers novel insights into the IgE scaffold, paving the way for the development of more effective and stable IgE‐based therapeutics. [ABSTRACT FROM AUTHOR]

Recent years have seen the development of a variety of mammalian library approaches for display and secretion mode. Advantages include library approaches for engineering, preservation of precious immune repertoires and their repeated interrogation, as well as screening in final therapeutic format and host. Mammalian display approaches for antibody optimization exploit these advantages, necessitating the generation of large libraries but in turn enabling early screening for both manufacturability and target specificity. For suitable libraries, high antibody integration rates and resulting monoclonality need to be balanced - we present a solution for sufficient transmutability and acceptable monoclonality by applying an optimized ratio of coding to non-coding lentivirus. The recent advent of microfluidic- assisted hit discovery represents a perfect match to mammalian libraries in secretion mode, as the lower throughput fits well with the facile generation of libraries comprising a few million functional clones. In the presented work, Chinese Hamster Ovary cells were engineered to both express the target of interest and secrete antibodies in relevant formats, and specific clones were strongly enriched by high throughput screening for autocrine cellular binding. The powerful combination of mammalian secretion libraries and microfluidics-assisted hit discovery could reduce attrition rates and increase the probability to identify the best possible therapeutic antibody hits faster. [ABSTRACT FROM AUTHOR]