Staudinger Ligation-based Cell Surface Chemical Conjugation Glycoengineering Service

Staudinger Ligation-based Cell Surface Glycoengineering Service at CD BioGlyco

Cell surface glycans determine cell interactions with other biomolecules and are dynamic indicators of cell physiology. It changes as cells develop, activate, and as the effects of disease transition from healthy to pathological states. Therefore, imaging of cell surface glycans is of great significance for the diagnosis and treatment of diseases. Glycoengineering is a discipline that studies the structure, function, and metabolic regulation of glycoconjugates and their applications. At CD BioGlyco, we use Glycoengineering Techniques to introduce target molecules carrying bioorthogonal groups such as azides, alkynes, ketones, or aldehydes into the cell surface without the need for genetic manipulation.

Glycan imaging is performed by two methods: imaging with lectins and antibodies, but in vivo use of this method is limited by low affinity. Another method is to use aldehydes or ketones to form Schiff bases, but when used in vivo, glycans cause problems by non-specific binding to endogenous metabolites. Staudinger ligation is a chemically selective ligation. The functional groups involved in the reaction do not react inside the biomolecule or on the cell surface. It is very suitable for labeling and imaging intracellular glycans. It provides new possibilities for visualizing the processes in which glycans participate in various biological changes in their native environment and detecting intracellular interactions.

The introduction of bioorthogonal phosphines and azides in Staudinger ligation has promoted the development of cell surface glycan imaging. We apply Staudinger ligation to the imaging and detection of cell surface glycans. Our process is as follows:

Given the smaller size of the azide and the difficulty in synthesizing azidosugar as a metabolic precursor, we load the azide on a glycoconjugate on the cell surface to form N-azidoacetylmannosamine (ManNAz). It is a bioorthogonal chemical reporter group and the addition of the azide group will not affect the properties of the glycoconjugate itself.
The Staudinger ligation reaction occurs with triphenylphosphine labeled with biotin or fluorophores to generate stable cell surface adducts, thereby enabling imaging and detection of cell surface glycans.

Fig.1 Staudinger ligation-based cell surface glycoengineering service. (CD BioGlyco)

Publication

Paper Title: A FRET-based fluorogenic phosphine for live-cell imaging with the Staudinger ligation

Technology: Cell surface glycan imaging with the Staudinger ligation

Journal: Angewandte Chemie International Edition

IF: 16.6

Published: 2008

Results: Here, the authors reported a fluorescent phosphine reagent 1 that was used to label azides displayed on Chinese hamster ovary (CHO) cells. The results showed that CHO cells treated with N-α-azidoacetylmannosamine (Ac₄ManNAz) labeled with fluorescent phosphine reagent 1 showed strong fluorescent labeling. Control cells treated with fluorescent phosphine reagent 1 and lacking azide treatment showed weaker fluorescence.

Fig.2 Flow cytometry analysis of live Ac₄ManNAz-treated CHO cells labeled by fluorescent phosphine reagent 1. (Hangauer & Bertozzi, 2008)

Highlights

Staudinger ligation is suitable for biotin labeling, fluorophore labeling, DNA labeling, polymer and material synthesis, and the connection of biomolecules, etc.
Staudinger-connected reagents have no obvious toxicity and can be used for fluorescence imaging of living cells or organisms.
Staudinger ligation of fluorescent reagents to Ac₄ManNAz results in ester cleavage accompanied by non-quenching, overcoming the shortcomings of non-specific ester hydrolysis that releases the quencher prematurely and produces unwanted background fluorescence.

CD BioGlyco is a global professional glycobiology company dedicated to providing the most comprehensive glycobiology services to researchers around the world. We have developed advanced Glycoengineering Platforms to help clients purify, label, and detect glycans in organisms. If you happen to need glycoengineering services, please feel free to contact us and our experts will reply to you in time to provide you with the optimal experimental plan.

Reference

Hangauer, M.J.; Bertozzi, C.R. A FRET-based fluorogenic phosphine for live-cell imaging with the Staudinger ligation. Angewandte Chemie international edition. 2008, 47(13): 2394-2397.

This service is for Research Use Only, not intended for any clinical use.