The detection of glycoproteins is an important area in biochemistry and molecular biology, as glycosylation is one of the most common and complex post-transcriptional modifications of proteins in cellular processes.
Figure 1Glycoprotein analysis services.
How is glycoprotein detected?
1. Mass spectrometry (MS).
Mass spectrometry is one of the most powerful tools in glycoprotein analysis, especially for structural identification and complex glycan analysis. It can be used to identify glycosylation sites and the structure of glycans. Mass spectrometry relies on measuring the mass-to-charge ratio (mz) of a protein or peptide. With this method, the quality, type, and connection of the sugar chains can be precisely determined.
1.Advantage: Provides very detailed and precise information on the composition and structure of glycoprotein glycans. Especially when using tandem mass spectrometry (MSMS), it provides in-depth information about glycan branch branches and link types. 2.Disadvantages: The technology is complex and requires professional operation and data analysis skills. In addition, the cost is high and there are certain requirements for sample size. 2.Western blotting
Western blotting first separates the protein by gel electrophoresis, then transfers the protein to a membrane and detects the glycoprotein with specific antibodies.
1.Advantage: Provides information about the relative molecular weight and presence of glycoproteins. This method is relatively simple and inexpensive. 2.Insufficient: Unable to provide detailed information on the structure of the sugar chain. Sensitivity and specificity are limited by the available antibodies. 3.Enzyme-linked immunosorbent assay (ELISA).
The glycoprotein is fixed with a specific antibody or lectin, and then detected by a secondary antibody or lectin (often with an enzyme label) that binds to it.
1.Benefits: Suitable for rapid screening and quantification. High sensitivity allows for the processing of a large number of samples. 2.Disadvantages: As with Western blotting, detailed information on the structure of the sugar chain cannot be provided. 4.Flow cytometry
Glycoproteins on the cell surface can be detected at the single-cell level by flow cytometry using fluorescently labeled antibodies or interpretive markers.
1.Advantage: Ability to analyze at the cellular level, suitable for live-cell analysis. 2.Deficiencies: Mainly limited to glycoproteins on the cell surface, it cannot provide comprehensive information on the structure of sugar chains. 5.Lectin affinity chromatography
Glycoproteins are detected using specific lectins, a protein that naturally binds to specific sugar structures. Lectins bind specifically to glycans and are then detected by staining or fluorescent labeling.
1.Advantages: Ability to detect specific types of glycan structures, suitable for rapid screening and relative quantification. 2.Deficiencies: Unable to provide precise structural information about glycan chains. Specificity depends on the interpretive marker used. 
Figure 2Far-Western blot analysis.
Choosing the right glycoprotein assay depends on the specific purpose of the study and the resources available. For example, for studies that require detailed information on the structure of glycan chains, mass spectrometry may be the best option; For rapid screening or quantification, ELISA may be considered. Each method has its own unique advantages and limitations, and understanding these differences is critical to selecting the most appropriate experimental method.