NDLI: Purification and Characterization of Antibodies Directed against the α-Gal Epitope

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Purification and Characterization of Antibodies Directed against the α-Gal Epitope

Content Provider	MDPI
Author	Julia, Rosenlöcher Zappe, Andreas Kohla, Guido Hinderlich, Stephan Parr, Maria
Copyright Year	2021
Description	The α-Gal epitope is an immunogen trisaccharide structure consisting of N-acetylglucosamine (GlcNAc)β1,4-galactose (Gal)α1,3-Gal. It is presented as part of complex-type glycans on glycoproteins or glycolipids on cell surfaces of non-primate mammalians. About 1% of all antibodies in human sera are specific toward α1,3-Gal and are therefore named as anti-α-Gal antibodies. This work comprises the purification and characterization of anti-α-Gal antibodies from human immunoglobulin G (IgG). A synthetically manufactured α Gal epitope affinity resin was used to enrich anti-α-Gal antibodies. Selectivity experiments with purified antibodies were carried out using enzyme-linked immunosorbent assays (ELISA), Western blotting, and erythrocyte agglutination. Furthermore, binding affinities toward α-Gal were determined by surface plasmon resonance (SPR) and the IgG distribution of anti α Gal antibodies (83% IgG2, 14% IgG1, 2% IgG3, 1% IgG4) was calculated applying ELISA and immunodiffusion. A range of isoelectric points from pH 6 to pH 8 was observed in 2D gel electrophoresis. Glycan profiling of anti α Gal antibodies revealed complex biantennary structures with high fucosylation grades (86%). Additionally, low amounts of bisecting GlcNAc (15%) and sialic acids (13%) were detected. The purification of anti-α-Gal antibodies from human IgG was successful, and their use as detection antibodies for α Gal-containing structures was evaluated.
Ending Page	97
Page Count	17
Starting Page	81
e-ISSN	26736411
DOI	10.3390/biochem1020008
Journal	BioChem
Issue Number	2
Volume Number	1
Language	English
Publisher	MDPI
Publisher Date	2021-08-02
Access Restriction	Open
Subject Keyword	BioChem Biochem Biochemical Research Immunology Α-gal Epitope Allergy Anti-α-gal Antibody Igg Subclasses N-glycosylation Surface Plasmon Resonance Gel Electrophoresis
Content Type	Text
Resource Type	Article

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