Rapid Antibody Glycoengineering in Chinese Hamster Ovary Cells

Recombinant monoclonal antibodies bind specific molecular targets and, subsequently, induce an immune response or inhibit the binding of other ligands. However, antibody functionality and half-life may be reduced by type distribution host-specific glycosylation. Attempts to produce superior have inspired development genetically modified producer cells that synthesize glyco-optimized antibodies. Glycoengineering typically requires generation a stable knockout knockin cell line using methods such as clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9. Monoclonal produced engineered are then characterized mass spectrometric determine if desired glycoprofile has been obtained. This strategy is time-consuming, technically challenging, specialists. Therefore, alternative utilizes streamlined protocols for genetic glycoengineering glycan detection assist endeavors toward optimal In this proof-of-concept study, IgG-producing Chinese hamster ovary served ideal host optimize glycoengineering. Short interfering RNA targeting Fut8 gene was delivered cells, resulting changes in FUT8 expression were quantified. The results indicate knockdown method efficient, leading ~60% reduction FUT8. Complementary analysis performed rapid yet highly sensitive technique: capillary gel electrophoresis laser-induced fluorescence detection. All experiments showed increase afucosylated glycans; however, greatest shift achieved study ~20%. protocol simplifies efforts harnessing silico design tools, commercially synthesized reagents, quantification assays do not require extensive prior experience. As such, time efficiencies offered investigations into new targets.