Formation of a Monoclonal Antibody-Gold Nanoparticle Conjugate for Oligosaccharide Screening by a Lectin-Based Colorimetric Assay

 

Germarie Sánchez-Pomales, Rebecca A. Zangmeister and Michael J. Tarlov

 

Process Measurements Division, Chemical Science and Technology Laboratory,

National Institute of Standards and Technology

 

 

Glycosylation of monoclonal antibodies (mAbs) offers both a challenge and an opportunity to the biopharmaceutical industry in manufacturing improved and consistent therapeutic antibodies. Glycosylation can significantly affect the safety and efficacy profiles of therapeutic mAbs; it is therefore essential to characterize and control the glycosylation.  One method that is used for glycosylation profiling is lectin binding. Lectins are multivalent carbohydrate-binding proteins with particular specificity. Addition of a lectin with affinity for the glycans of the antibody will cross-link two or more antibodies and induce the formation of aggregates. A major disadvantage of this approach is poor lectin access to the oligosaccharides, which are often buried within the native conformation of the mAb.  Therefore, development of more efficient lectin-based glycosylation analysis should include a strategy to make the oligosaccharides more sterically accessible.

To address this challenge, we adsorbed mAbs on gold nanoparticles (NPs). The rationale behind this method is that once the mAb is adsorbed to the Au NPs, the oligosaccharides will become more accessible to lectin-based analysis. This feature was exploited by developing a colorimetric assay where aggregation of the NPs occurred due to the interaction between the target species immobilized on the NPs (i.e. mAb), and the recognition molecule (i.e. the lectin).

Confirmation of the adsorption of the mAb to the NPs was performed by spectroscopic techniques. Furthermore, we were able to demonstrate that the glycans of the mAb Au NP conjugate are well presented and accessible to interact with lectins, and that multivalent binding between lectins and the presented glycans induced aggregation of the mAb-modified gold nanoparticles.  This work seeks to improve the understanding of the interaction between mAbs and gold and will lead to the development of simple, efficient and reliable glycosylation assays.