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Expansins are small proteins that play a key role in modifying the structure of the cell wall during various physiological processes, in particular cell growth by elongation. Expansins are encoded by a large multigene family and are divided into four subfamilies, the main ones being alpha- and beta-expansins; the latter have received special development in cereals. Expansins are thought to modify the interactions of cellulose with xyloglucan (alpha-expansins) or with arabinoxylan (beta-expansins). At the same time, expansins do not have catalytic activity, the specific mechanism of their action is unclear, same as the physiological significance of such a wide variety of isoforms. To study the expression patterns of individual expansins, we conducted a transcriptomic analysis of all expansin genes identified in the maize genome using a convenient model system—zones of the maize growing primary root that differ in the stage of cell development and the composition of their cell walls. Of the 91 maize expansin genes, 67 were expressed in the root, and most expansin genes were characterized by a narrow range of zones with maximum transcript levels. Using a glycoarray containing 183 polysaccharides from the cell walls of plants of various species, we demonstrated that recombinant expansins AtEXPA1 and AtEXPB1 are able to bind to arabinogalactans and rhamnogalacturonans I and a number of other cell wall polysaccharides, which expands the list of their potential carbohydrate targets. Differences in the specificity of the interaction of alpha- and beta-expansins with various polysaccharides, both quantitatively and qualitatively, have been demonstrated. It has been hypothesized that the abundance of expansins in one plant organism and the fine regulation of their expression can be explained, at least in part, by the specific binding of individual expansins to specific cell wall polysaccharides. [ABSTRACT FROM AUTHOR]

Banana peel (BP) is the primary by-product generated during banana processing which causes numerous environmental issues. This study examines the physical attributes, proximate analysis, glycoarray profiling, antioxidant abilities, and prebiotic activity of BP. The analysis demonstrated that carbohydrates constituted the primary components of BP and the glycoarray profiling indicated that BP contains multiple pectin and hemicellulose structures. BP also contained phenolic compounds, including (+)-catechin and gallic acid, flavonoid compounds, and antioxidant activities. BP demonstrated prebiotic effects by promoting the proliferation of advantageous gut bacteria while inhibiting the growth of harmful bacteria. The prebiotic index scores demonstrated that BP exhibited a greater capacity to promote the growth of beneficial bacteria in comparison to regular sugar. The study demonstrated the potential of the BP as a valuable source of dietary fibre, bioactive compounds, and prebiotics. These components have beneficial characteristics and can be utilised in the production of food, feed additives, and functional food. [ABSTRACT FROM AUTHOR]

The dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) is an innate immune C-type lectin receptor that recognizes carbohydrate-based pathogen associated with molecular patterns of various bacteria, fungi, viruses and protozoa. Although a range of highly mannosylated glycoproteins have been shown to induce signaling via DC-SIGN, precise structure of the recognized oligosaccharide epitope is still unclear. Using the array of oligosaccharides related to selected fragments of main fungal antigenic polysaccharides we revealed a highly specific pentamannoside ligand of DC-SIGN, consisting of α-(1 → 2)-linked mannose chains with one inner α-(1 → 3)-linked unit. This structural motif is present in Candida albicans cell wall mannan and corresponds to its antigenic factors 4 and 13b. This epitope is not ubiquitous in other yeast species and may account for the species-specific nature of fungal recognition via DC-SIGN. The discovered highly specific oligosaccharide ligands of DC-SIGN are tractable tools for interdisciplinary investigations of mechanisms of fungal innate immunity and anti-Candida defense. Ligand- and receptor-based NMR data demonstrated the pentasaccharide-to-DC-SIGN interaction in solution and enabled the deciphering of the interaction topology.

Banana peel (BP) is the primary by-product generated during banana processing which causes numerous environmental issues. This study examines the physical attributes, proximate analysis, glycoarray profiling, antioxidant abilities, and prebiotic activity of BP. The analysis demonstrated that carbohydrates constituted the primary components of BP and the glycoarray profiling indicated that BP contains multiple pectin and hemicellulose structures. BP also contained phenolic compounds, including (+)-catechin and gallic acid, flavonoid compounds, and antioxidant activities. BP demonstrated prebiotic effects by promoting the proliferation of advantageous gut bacteria while inhibiting the growth of harmful bacteria. The prebiotic index scores demonstrated that BP exhibited a greater capacity to promote the growth of beneficial bacteria in comparison to regular sugar. The study demonstrated the potential of the BP as a valuable source of dietary fibre, bioactive compounds, and prebiotics. These components have beneficial characteristics and can be utilised in the production of food, feed additives, and functional food.

The specificity of the most plant carbohydrate-binding proteins (CBP), many of which are known only through bioinformatic analysis of the genome, has either not been studied at all or characterized to a limited extent. The task of deciphering the carbohydrate specificity of the proteins can be solved using glycoarrays composed of many tens or even hundreds of glycans immobilized on a glass surface. Plant carbohydrates are the most significant natural ligands for plant proteins; this work shows that plant polysaccharides without additional modification can be immobilized on the surface, bearing N-hydroxysuccinimide activated carboxyl groups. As a result, an array of 113 well-characterized polysaccharides isolated from various plant cell walls, 23 mono- and oligosaccharides – components of polysaccharides, and glycans – ligands for widely known plant lectins was designed. Upon chemical immobilization of polysaccharides, their functional activity was preserved, which was confirmed by the results of interaction with antibodies and the plant lectin ricin. Using the constructed array, a previously unknown ability of ricin to bind polysaccharides was found, which significantly expands the knowledge of its specificity, and it was also found that a large variety of antibodies to plant polysaccharides are present in human peripheral blood. [ABSTRACT FROM AUTHOR]

Monoclonal antibody EBCA-1 is used in the sandwich immune assay for the detection of circulating Candida mannan in blood sera samples for the diagnosis of invasive candidiasis. To reinvestigate carbohydrate specificity of EBCA-1, a panel of biotinylated oligosaccharides structurally related to distinct fragments of Candida mannan were loaded onto a streptavidin-coated plate to form a glycoarray. Its use demonstrated that EBCA-1 recognizes the trisaccharide β-Man-(1→2)-α-Man- (1→2)-α-Man and not homo-α-(1→2)-linked pentamannoside, as was reported previously. [ABSTRACT FROM AUTHOR]

Klebsiella pneumoniae Is a Gram‐negative pathogenic bacterium that emerges a challenge for modern medicine due to intensively rising multi‐drug resistance. Specific LPS O‐chains represent promising targets for development of vaccines for immunoprophylaxis or immunotherapy of K. pneumoniae‐caused diseases. Herein, the synthesis of di‐, tetra‐, and hexasaccharides which represent structural fragments of both galactan I and galactan II of K. pneumoniae LPS O‐chains is described. The galactofuranoside building block used for assembling the galactan I chains was prepared by pyranoside‐into‐furanoside (PIF) rearrangement of the corresponding galactopyranoside precursor bearing an acid‐labile temporary p‐methoxybenzyl protecting group at O(3). The synthesized set of K. pneumoniae antigenic oligosaccharides in the form of their biotinylated derivatives was used to create a glycoarray which was applied for the screening of antibodies to galactan I and galactan II in anti‐K. pneumoniae sera. The obtained results demonstrate the applicability of such a glycoarray‐based assay for typing antibodies to Klebsiella strains belonging to most frequent O1 serogroup. [ABSTRACT FROM AUTHOR]

Toxoplasma gondii rhoptry neck protein 4 (TgRON4) is a component of the moving junction, a key structure for host cell invasion. We previously showed that host cellular β-tubulin is a binding partner of TgRON4 in the invasion process. Here, to identify other binding partners of TgRON4 in the host cell, we examined the binding of TgRON4 to components of the host cell surface. TgRON4 binds to various mammalian cells, but this binding disappeared in glycosaminoglycan- and heparan sulfate-deficient CHO cells and after heparitinase treatment of mammalian cells. The C-terminal half of TgRON4 showed relatively strong binding to cells and heparin agarose. A glycoarray assay indicated that TgRON4 binds to heparin and modified heparin derivatives. Immunoprecipitation of T. gondii -infected CHO cell lysates showed that TgRON4 interacts with glypican 1 during Toxoplasma invasion. This interaction suggests a role for heparan sulfate in parasite invasion. [ABSTRACT FROM AUTHOR]

Abstract: Due to the ability of Pseudomonas aeruginosa (PA) to develop antibiotic resistances, alternative therapeutic strategies have been proposed. Among others, carbohydrate multivalent molecules targeting lectin‐based virulent factors have been widely reported in particular those targeting LecA. LecA is a tetravalent galactose specific lectin involved in biofilm formation and cell internalization. Herein, we report the synthesis of 36 galactoclusters built from galactosides with aromatic and non‐aromatic aglycons and with an additional chain. The chains were either neutral or positively charged. Only the galactoclusters with naphthyl or tyrosine aglycon showed a moderate increase of binding for the positively charged 3‐dimethylammonium propyl chain. In contrast, the non‐aromatic galactoclusters display typically poorer binding properties towards LecA. The introduction of these side chains led to improved affinities up to becoming comparable to the high‐affinity aromatic galactoclusters. [ABSTRACT FROM AUTHOR]

We performed a serological investigation using glycoarray in Guillain-Barré syndrome (GBS), chronic inflammatory demyelinating polyradiculoneuropathy (CIDP), and multifocal motor neuropathy (MMN). Antibodies to 10 glycolipids and 45 glycolipid complexes were tested. Anti-GM1/sulfatide and anti-GA1/sulfatide IgG antibodies were common in GBS (20.0% and 19.0%, respectively). Anti-GQ1b/sulfatide IgG antibody was detected in 14.0% of GBS patients. IgG antibodies to antigens containing GQ1b were significantly correlated with ophthalmoplegia in GBS (p < 0.01). IgM antibodies to antigens containing GM1 or GalNAc-GD1a were in 50% and 37.5% of MMN patients, respectively. Glycoarray is efficient for detecting antibodies against numerous glycolipid complexes in immune-mediated neuropathies. [ABSTRACT FROM AUTHOR]

Abstract: Nowadays, there is a great interest for understanding the structure/function relationship governing recognition of carbohydrates by their receptors for the design of new treatments. Indeed, carbohydrates and glycoconjugates play a major role in key biological events such as cell–cell recognition, pathogenesis inflammation, and host pathogen interactions. Pseudomonas aeruginosa (PA) is one of the predominant bacterium encountered in nosocomial infections. PA infections often lead to chronic inflammation and eventually to death despite aggressive antibiotic therapy: the emergence of resistant strains and biofilm formation seems to give a selective advantage to the bacterium. A promising approach is to inhibit the virulence factors of PA such as PA-IL which is a galactose specific lectin. Herein, we develop a microarray to probe the binding of six galacto-conjugates to PA-IL differing by their spatial configuration and geometry. This microsystem is made of 40 independent microwells in which 64 spots of glycoconjugates probes are arrayed by using DNA Directed Immobilization (DDI). This microsystem allows, in a multiplex fashion, qualitative information on the binding by direct fluorescence readout as well as quantitative information by the determination of IC50 values in a competition assay and surface dissociation constants (K d). According to our data, direct fluorescent signals (FI635), IC50 and K d values provided similar ranking for glycoconjugates with respect to PA-IL binding thus affording a general tool for the selection of galacto-conjugates displaying the best affinities toward PA-IL. [Copyright &y& Elsevier]

A microarray enables high-throughput interaction screening of numerous biomolecules; however, fabrication of a microarray composed of cellular membrane components has proven difficult. We report fabrication of a liposomal glyco-microarray by using an azide-reactive liposome that carries synthetic and natural glycolipids via chemically selective and biocompatible liposome immobilization chemistry. Briefly, liposomes carrying anchor lipid dipalmitoylphosphatidylethanolamine (DPPE)-PEG-triphenylphosphine and ganglioside (GM1 or GM3) were prepared first and were then printed onto an azide-modified glass slide so as to afford a liposomal glyco-microarray via Staudinger ligation. Fluorescent dye release kinetics and fluorescence imaging confirmed successful liposome immobilization and specific protein binding to the intact arrayed glycoliposomes. The liposomal glyco-microarray with different gangliosides showed their specific lectin and toxin binding with different binding affinity. The azide-reactive liposome provides a facile strategy for fabrication of either a natural or a synthetic glycolipid-based membrane-mimetic glycoarray. This liposomal glyco-microarray is simple and broadly applicable and thus will find important biomedical applications, such as studying glycolipid-protein interactions and toxin screening applications. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

Porphyromonas gingivalis is a major pathogen in adult periodontitis. Fimbriae play an important role in the initial interaction between the bacteria and the host. Our earlier studies suggested that the oligosaccharide moiety of lactoferrin is involved in the interaction with fimbriae. Porphyromonas gingivalis fimbriae bound strongly to albumin-fucosylamide (albumin-1-amido-1-deoxy- l-fucose) and to a lesser extent to albumin- N-acetyl- d-galactosamine (albumin-p-aminophenyl- N-acetyl-β- d-galactosaminide, but failed to bind bovine serum albumin. In this study we explored the glycan array to determine the carbohydrate-binding specificity of P. gingivalis fimbriae. Purified fimbriae bind to glycans which have a Lewisx, Galβ1-4(Fucα1-3) GlcNAcβ structure in common. Interestingly, all glycans have a terminal fucose moiety and if fucose is removed, the fimbriae fail to bind. This is the first study that suggests that fucose is important for P. gingivalis fimbriae binding. [ABSTRACT FROM AUTHOR]

Abstract: A biotinylated mannotriose (Man3-bio) was dispersively immobilized in the matrix of biotinylated lactose (Gal-Glc-bio) on a streptavidin-covered, 27-MHz quartz crystal microbalance (QCM), and binding kinetics of concanavalin A (Con A) to Man3-bio in the Gal-Glc-bio matrix could be obtained from frequency decreases (mass increases) of the QCM. Association constants (K a) and binding and dissociation rate constants (k on and k off) could be determined separately as the 1:1 and 1:2 bindings of Con A to Man3-bio on the surface. When Man3-bio was immobilized with content of 1 to 5 mol% in the matrix, the 1:1 binding of Con A to Man3-bio was obtained as K a =(4±1)×106 M−1, k on =(4±1)×104 M−1 s−1, and k off =(12±2)×10–3 s−1. On the contrary, when Man3-bio was immobilized with content of 20 to 100 mol% in the matrix, the 1:2 binding of Con A to Man3-bio was obtained as K a =(14±2)×106 M−1, k on =(14±2)×104 M−1 s−1, and k off =(7±2)×10–3 s−1. Thus, K a for the 1:2 binding was 10 times larger than that for the 1:1 binding, with a three times larger binding rate constant (k on) and a three times smaller dissociation rate constant (k off). This is the first example to obtain separate kinetic parameters for the 1:1 and 1:2 bindings of lectins to carbohydrates on the surface. [Copyright &y& Elsevier]

Abstract: Glycoarrays are powerful tools for the understanding of protein/carbohydrate interactions and should find applications in the diagnosis of diseases involving these interactions. Immobilisation of the carbohydrate probe is a key issue in the elaboration of high performance devices. In the present study, we have compared the fluorescent signal intensity and determined the lower detection limit of glycoconjugates immobilised at two concentrations (0.5 and 25μM) by DNA-directed immobilisation (DDI), to glycoconjugates covalently immobilised on the solid support (borosilicate glass slide). At 0.5μM, DDI led to a stronger fluorescence signal (by a factor of 4.5) and to a lower detection limit (20nM) than covalent immobilisation (higher than 200nM). We also report the development of an IC50 measurement assay of DDI immobilised glycoconjugates. We found that the relative affinity per galactose residue of RCA 120 for glycoconjugates bearing one or three galactose residues was different by a factor of 23 when measured under IC50 conditions or by direct fluorescence reading. [Copyright &y& Elsevier]

Influenza A virus specificity for the host is mediated by the viral surface glycoprotein hemagglutinin (HA), which binds to receptors containing glycans with terminal sialic acids. Avian viruses preferentially bind to α2-3-linked sialic acids on receptors of intestinal epithelial cells, whereas human viruses are specific for the α2-6 linkage on epithelial cells of the lungs and upper respiratory tract. To define the receptor preferences of a number of human and avian H1 and H3 viruses, including the 1918 H1N1 pandemic strains, their hemagglutinins were analyzed using a recently described glycan array. The array, which contains 200 carbohydrates and glycoproteins, not only revealed clear differentiation of receptor preferences for α2-3 and/or α2-6 sialic acid linkage, but could also detect fine differences in HA specificity, such as preferences for fucosylation, sulfation and sialylation at positions 2 (Gal) and 3 (GlcNAc, GalNAc) of the terminal trisaccharide. For the two 1918 HA variants, the South Carolina (SC) HA (with Asp190, Asp225) bound exclusively α2-6 receptors, while the New York (NY) variant, which differed only by one residue (Gly225), had mixed α2-6/α2-3 specificity, especially for sulfated oligosaccharides. Only one mutation of the NY variant (Asp190Glu) was sufficient to revert the HA receptor preference to that of classical avian strains. Thus, the species barrier, as defined by the receptor specificity preferences of 1918 human viruses compared to likely avian virus progenitors, can be circumvented by changes at only two positions in the HA receptor binding site. The glycan array thus provides highly detailed profiles of influenza receptor specificity that can be used to map the evolution of new human pathogenic strains, such as the H5N1 avian influenza. [Copyright &y& Elsevier]

Understanding the specificity of cell-surface carbohydrates interaction with antibodies and receptors is important for the development of new therapeutics and high-sensitivity diagnostics. This approach is, however, limited to the availability of natural and truncated sequences of the oligosaccharides and the sensitivity of the assay system. Reported here is the synthesis of the cancer antigen Globo H hexasaccharide, an epitope found on the cell surface of breast, prostate, and ovarian cancers, and its truncated sequences by using the programmable one-pot synthesis strategy. The saccharides were then arrayed covalently on glass slides with different density and used for the fluorencense-based binding analysis of two monoclonal antibodies against Globo H and the serum from breast cancer patients, to define the specificity of these antibodies. It was shown that the terminal tetrasaccharide binds the monoclonal antibodies equally well as does the hexasaccharide and the fucose residue is required for effective binding. The serum binds both the defucosylated pentasaccharide and the fucosylated hexasaccharide without a significant difference, perhaps because of the polyclonal nature of the serum or the presence of diverse immune responses to different sugar epitopes at various stages. This method requires very small amounts of materials and is more effective and sensitive than the traditional ELISA method, and thus provides another platform to monitor the immune response to carbohydrate epitopes at different stages during differentiation, metastasis, or treatment. [ABSTRACT FROM AUTHOR]

Abstract: A sensitive, specific, and rapid method for the detection of carbohydrate–protein interactions was demonstrated using quantum dots (QDs) as a fluorescence label coupled with protein. 1,3-Dipolar cycloaddition between azide and alkyne was exploited to attach α-d-glucopyranoside to a C14 hydrocarbon chain that noncovalently binds to the microtiter well surface, and the product formation was detected by both electrospray ionization–mass spectrometry (ESI–MS) and QD- (or fluorescein isothiocyanate (FITC))-conjugated lectin binding. It indicated that the peak intensity of the fluorescence emission was proportional to the initial concanavalin A (Con A) concentration in the range of 2×10−3μmol/L to 2×10−2mmol/L with a detection limit at least 100 times lower than that of the FITC-based method. [Copyright &y& Elsevier]

A noncovalent carbohydrate microarray was used to screen for possible inhibitors to fucosyltransferases, which are critical to the synthesis of inflammation mediators like sialyl Lewis x (SLex). Inhibition was followed by observation of the transferred fucose on the carbohydrate array with the lectin Tetragonolobus purpureas. Of these compounds, four inhibitors with nanomolar Kis were discovered, with three of the top five inhibitors exhibiting a common architecture. [Copyright &y& Elsevier]

Previous studies have shown that in the blood of healthy donors (1) there are no natural antibodies against sialylated glycoproteins, which contain Neu5Acα (N-acetylneuraminic acid) as the most widespread form of human sialic acid, and (2) there is a moderate level of antibodies capable of binding unnatural oligosaccharides, where Neu5Ac is beta-linked to a typical mammalian glycan core. In the present study, we investigated antibodies against βNeu5Ac in more detail and verified the presence of Kdn (2-keto-3-deoxy- D-glycero-D-galacto-nonulosonic acid) as a possible cause behind their appearance in humans, taking into account the expected cross-reactivity to Kdn glycans, which are found in bacterial glycoconjugates in both the α- and β-forms. We observed the binding of peripheral blood immunoglobulins to sialyllactosamines (where "sialyl" is Kdn or neuraminic acid) in only a very limited number of donors, while the binding to monosaccharide Kdn occurred in all samples, regardless of the configuration of the glycosidic bond of the Kdn moiety. In some individuals, the binding level of some of the immunoglobulins was high. This means that bacterial Kdn glycoconjugates are very unlikely to induce antibodies to βNeu5Ac glycans in humans. To determine the reason for the presence of these antibodies, we focused on noninfectious pathologies, as well as on a normal state in which a significant change in the immune system occurs: namely, pregnancy. As a result, we found that 2/3 of pregnant women have IgM in the blood against Neu5Acβ2-3Galβ1-4GlcNAcβ. Moreover, IgG class antibodies against Neu5Acβ2-3Galβ1-4GlcNAcβ and Neu5Acβ2-6Galβ1-4GlcNAcβ were also detected in eluates from the placenta. Presumably, these antibodies block fetal antigens., (Copyright ® 2022 National Research University Higher School of Economics.)

Glycolipids cluster in plasma membranes to form heterogenous patches. Whereas lectins and antibodies have been conventionally viewed as binding a single oligosaccharide head group, and assayed accordingly, it is increasingly evident that cis-interactions between glycan headgroups may form unique molecular shapes that either enhance or attenuate binding of antibodies and other proteins. Herein we describe a method for assaying antibody binding to heteromeric glycolipid complexes that allows rapid, simple, inexpensive and high-throughput assessment of binding events, focusing on autoantibodies present in human serum., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

Abstract: The work presented herein is a new noncovalent glycoarray assembly method for microplates created by simply mixing together a carbohydrate and a tetradecylamine. α-Mannose was utilized in the model study and product formation was detected by lectin binding. The method can be further extended to array complex carbohydrates. [Copyright &y& Elsevier]

Methodologies to identify epitopes or ligands of the fungal cell wall polysaccharides influencing the immune response of human pathogens have to date been imperfect. Using the galactomannan (GM) of Aspergillus fumigatus as a model, we have shown that synthetic oligosaccharides of distinct structures representing key fragments of cell wall polysaccharides are the most precise tools to study the serological and immunomodulatory properties of a fungal polysaccharide., (Copyright © 2020 Wong et al.)

A repertoire of monoclonal antibodies was generated by immunization of mice with cancer-associated glycoprotein CA19.9, and two of them were selected as optimal capture and detecting counterparts for sandwich test system for detection of CA19.9. Fine epitope specificity of the antibodies was determined using printed glycan array, enzyme-linked immunosorbent assay, and inhibitory enzyme-linked immunosorbent assay. Unexpectedly, both immunoglobulins did not bind key epitope of CA19.9 glycoprotein, tetrasaccharide SiaLeA, as well as its defucosylated form sialyl LeC (known as CA-50 epitope). The antibodies were found to have different glycan-binding profiles; however, they recognized similar glycotopes with common motif Galβ1-3GlcNAcβ (LeC), thus resembling specificity of human natural cancer-associated anti-LeC antibodies. We propose that cancer-specific glycopeptide epitope includes Galβ1-3GlcNAcβ fragment of a glycoprotein O-chain in combination with proximal hydrophobic amino acid(s) of the polypeptide chain. [ABSTRACT FROM AUTHOR]

A repertoire of monoclonal antibodies was generated by immunization of mice with cancer-associated glycoprotein CA19.9, and two of them were selected as optimal capture and detecting counterparts for sandwich test system for detection of CA19.9. Fine epitope specificity of the antibodies was determined using printed glycan array, enzyme-linked immunosorbent assay, and inhibitory enzyme-linked immunosorbent assay. Unexpectedly, both immunoglobulins did not bind key epitope of CA19.9 glycoprotein, tetrasaccharide SiaLe A , as well as its defucosylated form sialyl Le C (known as CA-50 epitope). The antibodies were found to have different glycan-binding profiles; however, they recognized similar glycotopes with common motif Galβ1-3GlcNAcβ (Le C ), thus resembling specificity of human natural cancer-associated anti-Le C antibodies. We propose that cancer-specific glycopeptide epitope includes Galβ1-3GlcNAcβ fragment of a glycoprotein O-chain in combination with proximal hydrophobic amino acid(s) of the polypeptide chain.

Glycan microarrays have become a powerful platform to investigate the interactions of carbohydrates with a variety of biomolecules. However, the number and diversity of glycans available for use in such arrays represent a key bottleneck in glycan array fabrication. To address this challenge, we describe a novel glycan array platform based on surface patterning of engineered glycophages that display unique carbohydrate epitopes. Specifically, we show that glycophages are compatible with surface immobilization procedures and that phage-displayed oligosaccharides retain the ability to be recognized by different glycan-binding proteins (e.g. antibodies and lectins) after immobilization. A key advantage of glycophage arrays is that large quantities of glycophages can be produced biosynthetically from recombinant bacteria and isolated directly from bacterial supernatants without laborious purification steps. Taken together, the glycophage array technology described here should help to expand the diversity of glycan libraries and provide a complement to the existing toolkit for high-throughput analysis of glycan-protein interactions., (Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)