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Glycans or carbohydrates attached to cell membranes and other types of proteins are the primary determinants for binding activity and consecutive cellular function. The study of the function and characteristics of these sugars is a fundamental part of immunology research and is applicable in understanding a variety of intercellular interactions. New technologies such as microarray for glycan-binding applications allow researchers to investigate and reveal new information about this wide and developing field of glycoscience.
2. Basic Concept for Indirect Binding Assay
1. Basic Concept for Direct Binding Assay
Certain broadly neutralizing antibodies need a pre-complexing assay to enhance their relatively weak binding to N-glycans. Below is an illustration for the pre-complexing binding assays:
Z Biotech offers several versions of the market’s most advanced glycan microarray substrates. Our slides feature a multivalent arrangement of functional groups on the surface, which allows the glycan structure to be displayed 3-dimentionally to increase glycan-protein binding affinity.
Z Biotech also provides optimized buffers and blockers for maintaining activity of analysis samples and obtaining high assay signal intensity for microarrays fabricated on the microarray substrates.
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Z Biotech also provides innovative and comprehensive Glycan Array slides for identifying glycan-binding proteins (GBPs):
1. 100 Glycan Array: The 100 Glycan Array is used as a general test to help researchers determine binding characteristics of antibodies, proteins, bacteria, cell cultures, or other potential biological samples to an array of 100 glycans representing the range of glycans found glycosylated to cell membranes.
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2. N-Glycan Array: N-linked oligosaccharides in particular play a major role in intercellular interactions and immune cell functions, making them often preferred glycan candidates for the study of carbohydrate-binding-vaccines, such as HIV Broadly Neutralizing Antibodies (bNAbs). The fundamental N-glycan structures provided in the N-glycan Array can provide understanding of basal binding determinants for antibodies or other proteins of interest.
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3. O-Glycan Array: Tumor-associated carbohydrate antigens (TACAs) represent potential biomarkers for early detection of cancer as well as immunotherapeutic targets. For example, the extracellular glycoprotein MUC1 is overexpressed and aberrantly glycosylated in many types of cancers. The extracellular domain of the MUC-1 contains a variable number of tandem repeats (VNTR) of 20 amino acids residues with serine (Ser) or threonine (Thr) sites for O-glycosylation. Generally, aberrantly expressed O-Glycans aid in the metastasis of diseased cells, yet also distinguish diseased from healthy cells at the cell surface. Therefore, the O-Glycan array can serve as a research tool for determining O-glycan immunodominant epitopes in cancers or sensitively detect the glycan binding epitopes of autoantibodies that target unique O-glycan antigens.
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4. Heparan Sulfate Glycan Array: Heparan sulfate is a sulfated, unbranched polysaccharide found on cell surfaces as well as in the extracellular matrix. Heparan sulfates interact with a variety of proteins and are known to be important to the regulation of many physiological and pathophysiological functions. Because of the prevalence and multifunctionality of heparan sulfates, investigation of the binding roles of this type of glycan has become invaluable for understanding how to combat related diseases. Our Heparan Sulfate Array can be used to determine the specific binding interactions between various types of heparan sulfates and their potential protein partners via microarray assays.
5. Lectin Microarray: Lectins are a class of carbohydrate-binding proteins purified from plants or animals that can bind to different carbohydrate structural epitopes in a highly specific manner. We selected 26 lectins to include in our first-generation lectin microarray. These 26 lectins represent most carbohydrate-binding epitopes and have been characterized on our glycan arrays. There are 8 or 16 identical subarrays on a single array chip so that 8 or 16 samples can be analyzed simultaneously. Our lectin microarray provides scientists with a powerful and sensitive tool for analyzing glycosylation profiles of therapeutic proteins, biomarkers, or other proteins of interest.
6. Neu5Gc/Neu5Ac N-Glycan Array: Neu5Gc is a major mammalian sialic acid synthesized from Neu5Ac by an enzyme that is no longer present in humans. Regardless, nonhuman, diet-derived Neu5Gc is metabolically integrated into a variety of human tissues. Human cells can biochemically recognize Neu5Gc and incorporate it into cell-surface glycoconjugates, however the immune system recognizes Neu5Gc-containing glycans as alien and produces anti-Neu5Gc antibodies resulting in inflammation. Diseases relating to chronic inflammation such as cancer, cardiovascular diseases, and thyroiditis are associated with higher levels of anti-Neu5Gc and Neu5Gc, and it is suspected to be involved in other diseases that are exacerbated by a red meat, Neu5Gc-rich diet. Z Biotech's Neu5Gc microarray represents a broad range of N-glycans found on cell surfaces that have incorporated Neu5Gc. These arrays can be investigated with glycan-binding proteins, antibodies, or cells in order to determine their specific interaction with this Neu5Gc xenoantigen. For comparison, each array includes the precursory, non-xenogenic Neu5Ac sialic acid form of each glycan.
7. HMO Glycan Array: Human milk oligosaccharides (HMOs) are the third most abundant solid ingredient in breast milk. These sugars - unique to human milk - protect newborns from disease and infection, establish healthy gut bacteria, and are essential for infant brain, intestinal, and immune system development. Z Biotech's HMO Glycan Array can be tested with cells, antibodies, or other proteins to determine which HMOs interact and give insight into the functions of these HMOs and their binding partners.
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8. Glycosphingolipid Glycan Array: The cells of the nervous system communicate and are identified by the variety of sialylated glycosphingolipids (GSLs) called gangliosides presented on their surface membranes. The species of gangliosides expressed by each cell are incredibly important for brain function and go through dramatic changes during brain development. The expression of specific types of glycosphingolipids correlate with sensory and motor coordination, nerve regeneration, maintaining neuropsychological balance, and many other crucial neurological functions. Aberrant changes such as truncation, over- or under-expression of these gangliosides is involved in neurodegenerative diseases, Tay-Sachs disease, and tumor growth. Z Biotech presents a new, sensitive research tool specifically for investigating glycosphingolipids. Our Glycosphingolipid Glycan Microarray features 58 glycan epitopes representing a variety of glycosphingolipid structures presented on healthy and abnormal cells.
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