Campylobacter jejuni bacterial pathogen is among the primary causes of food-borne acute gastroenteritis in North America and the world. It has also been linked to severe post-infection sequelae such as Guillain-Barré syndrome. Previous studies identified C. jejuni surface capsular polysaccharide (CPS) as a target for creation of a carbohydrate based vaccine in which the CPS is conjugated to a carrier protein. In this thesis, following sample purification, aspects of C. jejuni HS:5 CPS structure were characterized using numerous analytical techniques such as NMR and GC-MS. CPS is comprised of α-DD-Heptoses linked at C2 to the anomeric carbons of glucose. The α-Glucose molecules are linked though C4 to the α-DD-Heptose anomeric carbon. The α-DD-Heptose structure also has an occasional ring structured amino acid modification. Following characterization the CPS was oxidized and developed into a prototype glycoconjugate vaccine using TEMPO oxidation and EDC-CRM197 coupling methods. / The Natural Sciences and Engineering Research Council of Canada (NSERC)

CARACTERISATION DE VACCINS A BASE DE GLUCIDESVariables influençant l'immunogénicité et propriétés physico-chimiques des vaccins glycoconjuguésDe nombreux aspects peuvent influer sur l'immunogénicité des vaccins conjugués et les principales variables étudiées jusqu'ici sont la taille du fragment saccharide et la nature des glycosides: taux de protéine dans le conjugué purifié, la stratégie de conjugaison, nature de l’espaceur et la protéine porteuse.La taille de la partie saccharidique et le ratio de cette partie / protéine a été étudiée dans différents travaux de Seppälä et Mäkelä,.Dans l'une des premières études sur l'effet de la taille sur l'immunogénicité de la protéine conjuguées à des dextrans, il a été montré que des dextrans de faible poids moléculaire conjugué à l’albumine sérique de poluet induit des réponses anti-dextran fortes chez la souris. L’augmentation de la taille du dextran, a abouti à une réduction de l’immunogénicité. Peeters et al. a montré qu’un tétramère synthétique de Hib unité capsulaire polysaccharide, conjuguée à un support protéique, induit chez des souris adultes, des niveaux d'anticorps primates non humains comparables à un conjugué Hib commercial. Ces niveaux sont plus élevées que ceux induits par un trimère répété, ce qui indique que pour le Hib un minimum de huit sucres est nécessaire pour une bonne réponse immunologique. Laferrière et al. a trouvé peu d'influence de la longueur de la chaîne glucidique sur l'immunogénicité des vaccins antipneumococciques conjugués chez la souris. Pozsgay et al. a étudié chez la souris, l'immunogénicité de l’oligosaccharides du LPS de Shigella dysenteriae de type 1 conjugué à l'albumine sérique humaine (HSA). Les auteurs ont constaté que les octa-, dodéca-, et des fragments de hexadécasaccharides induit des niveaux élevés d'anticorps IgG après trois injections. Ces niveaux sont supérieurs à ceux obtenus avec un conjugué tétrasaccharidique. L'influence du ratio glucides / protéine est différente pour les trois conjugués. Le conjugué octasaccharide-HSA avec la plus forte densité provoque une bonne réponse immunitaire, tandis que dans le cas des conjugués dodéca- et hexadécasaccharides, la densité médiane est optimal. Ces études suggèrent que la longueur de la chaîne d'oligosaccharides et le chargement de l’haptène peuvent être liés entre eux pour déterminer l'immunogénicité des vaccins glycoconjugués.L'espaceur est une molécule linéaire courte qui est généralement liée à la chaîne polysaccharidique et la protéine ou de fragments. Il ya des évidences dans la littérature qui suggèrent que les espaceurs rigides, contraints comme le maléimide cyclohexyle, provoquent une importante quantité d'anticorps indésirables, avec le risque de conduire à une réponse immunitaire éloignée de l'épitope ciblé sur la haptène. L'utilisation d'un alkyle souple type maleimido a été rapporté comme un moyen de surmonter l'immunogénicité observée précédemment. Un certain nombre de transporteurs protéiques ont été utilisés jusqu'ici dans l'évaluation préclinique et clinique de vaccins conjugués. Des protéines telles que les anatoxines diphtériques et tétaniques, qui dérivent des toxines respectives, après la décontamination chimique avec le formaldéhyde, ont été initialement choisies comme transporteur en raison de inocuité (tétanos et la vaccination contre la diphtérie). CRM 197, un mutant non toxique de la toxine 61 de la diphtérie a été largement utilisé comme support pour Hib. Un complexe protéique de la membrane externe de méningocoque du sérogroupe B a été utilisé par Merck comme support pour leur vaccin conjugué Hib. GSK dans leur vaccin antipneumococcique, conjugué multivalent, introduit la protéine D Hib liée à la plupart des polysaccharides inclus dans le vaccin. L'équipe de John Robbins fait un large usage de la forme recombinante non toxique de l’exo-toxine de Pseudomonas aeruginosa comme support contre Staphylococcus aureus de type 5 et 8 ainsi que pour Salmonella. / CHARACTERIZATION OF CARBOHYDRATE BASED VACCINES Variables influencing the immunogenicity and physicochemical properties of glycoconjugate vaccinesMany aspects can influence the immunogenicity of conjugate vaccines and the main variables investigated so far are the size of the saccharide moiety, the saccharide:protein ratio in the purified conjugate, the conjugation strategy, the nature of the spacer and the protein carrier. The size of the saccharide moiety and saccharide/protein ratio were investigated in different works such as Seppälä and Mäkelä in one of the first studies on the effect of size and chemistry on the immunogenicity of dextrans-protein conjugates found that dextrans of low molecular weight conjugated to chicken serum albumin, induced strong anti-dextran responses in mice, while increasing the dextrans' size resulted in reduced immunogenicity.47 Peeters et al. showed that a synthetic tetramer of Hib capsular polysaccharide repeating unit, conjugated to a protein carrier, induced in adult mice and non-human primates antibody levels comparable to a commercial Hib conjugate and higher than those induced by a trimer, indicating that for Hib a minimum of eight sugars is needed for a proper immunological response.48 Laferriere et al. found little influence of the carbohydrate chain length on the immunogenicity of pneumococcal conjugate vaccines in mice.49 Pozsgay et al. studied the immunogenicity in mice of synthetic Shigella dysenteriae type 1 LPS oligosaccharides conjugated to human serum albumin (HSA). The authors found that octa-, dodeca-, and hexadecasaccharide fragments induced high levels of lipopolysaccharide binding IgG antibodies in mice after three injections and were superior to a tetrasaccharide conjugate. The influence of the carbohydrate/protein ratio was different for the three conjugates. The octasaccharide-HSA conjugate with the highest density evoked a good immune response, while in the case of dodeca- and hexadecasaccharide conjugates, the median density was optimal.50 These studies suggest that oligosaccharide chain length and hapten loading might be interconnected in determining the immunogenicity of glycoconjugate vaccines. The spacer is a short linear molecule that is generally linked to the polysaccharide chain or to the protein or to both moieties, depending on the chemistry, used to facilitate the coupling between the protein and sugar. There are evidences in the literature which suggest that rigid, constrained spacers like cyclohexyl maleimide, elicit a significant amount of undesirable antibodies, with the risk of driving the immune response away from the targeted epitope on the hapten.51 52 The use of a flexible alkyl type maleimido spacer has been reported as a way to overcome the previous observed immunogenicity of cyclic maleimide linkers.53 A number of protein carriers have been used so far in preclinical and clinical evaluation of conjugate vaccines. 54 55 56 57 58 59 60Proteins such as diphtheria and tetanus toxoids, which derive from the respective toxins after chemical detoxification with formaldehyde, were initially selected as carrier because of the safety track record accumulated with tetanus and diphtheria vaccination. CRM197, a non-toxic mutant of diphtheria toxin61 which instead does not need chemical detoxification, has been extensively used as carrier for licensed Hib, pneumococcal, meningococcal conjugate vaccines and for other vaccines being developed. An outer membrane protein complex of serogroup B meningococcus has been used by Merck as carrier for their Hib conjugate vaccine.62 GSK in their multivalent pneumococcal conjugate vaccine introduced the use of the Hib-related protein D as carrier for most of the polysaccharides included into the vaccine.63 64 The team of John Robbins made extensive use of the recombinant non toxic form of Pseudomonas aeruginosa exo-toxin as carrier for Staphylococcus aureus type 5 and 8 as well as for Salmonella

Vaccination is one of the most effective public health interventions in the world, saving millions of lives and preventing the onset of debilitating diseases. With widespread emergence of multi-drug resistant pathogens, the importance of preventative medicine has become even more apparent. However, one of the limiting factors in developing novel vaccines that are both safe and highly immunogenic is the availability of adjuvant delivery systems licensed for human use. The purpose of this study was to investigate the role gold nanoparticles could play as an effective vaccine delivery system. A variety of coupling chemistries were explored for their ability to conjugate protein and polysaccharide antigens onto the surface of gold nanoparticles for the development of vaccines against a number of biologically important human pathogens including Y. pestis, B. mallei and S. pneumoniae. Retention of antigenicity and coupling efficiency of conjugated molecules was measured using characterisation techniques such as localised surface plasmon resonance and immunoblotting. Gold nanoparticle coupled antigens were then used to immunise mice and to measure the protective efficacy and the immunological response induced. The findings indicate antigen-specific immune responses are elevated when an antigen is coupled onto gold nanoparticles. Moreover, immunological data from nanoparticle coupled glycoconjugate vaccines against B. mallei and S. pneumoniae indicate the likely presence of a strong T cell immune response which is essential for providing immunological memory. Finally, an intracellular trafficking assay was carried out to identify some of the mechanisms that might be involved in uptake of gold nanoparticles into professional phagocytes. Confocal imaging of receptors associated with endosomal compartments revealed that gold nanoparticles may enter cells through multiple pathways. The findings reported in this study suggest that gold nanoparticles may be an excellent candidate for further investigation as a novel vaccine delivery system.

Le cancer est l’une des principales causes de mort dans le pays développés ; bien que les opérations chirurgicales, la radiothérapie et la chimiothérapie représentent aujourd’hui les principales options de traitement des patients souffrants de tumeurs malignes, leurs effets secondaires sévères ont ouvert la voie au développement de l’immunothérapie antitumorale. A part l’immunothérapie passive, qui est basée sur les anticorps ou tout autre composant du système immunitaire synthétisés en dehors du corps dont la potentielle menace de réactions immunes a été prouvée, nous avons concentré nos efforts sur l’immunothérapie active, qui réside dans la stimulation du système immunitaire du patient pour éradiquer sélectivement les cellules malignes. L’identification d’antigènes carbohydrates associés aux tumeurs (TACAs), surexprimés à la surface des cellules cancéreuses, a permis le développement de vaccins spécifiques à cet antigène. Il est connu depuis plus de 40 ans que la majorité des cancers chez l’homme sont caractérisés par une glycosylation aberrante. Les cellules tumorales peuvent surexprimer des versions tronquées d’oligosaccharides, une séquence terminale inhabituelle ou une augmentation de la sialylation des glycolipides et des glycoprotéines de surface. Un oligosaccharide d’une glycoprotéine tronqué peut rendre une partie de la chaîne principale du peptide, d’habitude caché par le sucre, plus accessible au système immunitaire. Parmi les différents TACAs, nous avons concentré notre attention sur les antigènes Tn et Tf, qui peuvent être trouvés sur des glycoprotéines comme MUC-1, surexprimés sur plus de 90% des carcinomes du sein. Bien que la conception de ces immunomodulateurs repose toujours sur des règles empiriques, il est important de déclencher à la fois la réponse humorale et cellulaire, ainsi qu’un effet de mémoire. Ce défi peut être relevé en combinant, sur une seule molécule, l’antigène carbohydrate exprimés à la surface des tumeurs (épitope des cellules B), les peptides capables de stimuler les cellules CD4+ et CD8+ (épitopes des cellules T) et un adjuvant, pour recueillir tous les éléments du système immunitaire au niveau du site d’injection et renforcer l’absorption des antigènes. De précédentes études faites dans notre groupe de recherche ont publié pour la première fois la synthèse et l’évaluation immunologique d’un prototype de vaccin anticancéreux à quatre composant capable d’induire une réponse immunitaire de longue durée sur des modèles murins. Dans mon travail de thèse, nous avons voulu synthétiser des prototypes de vaccin anticancéreux basés sur les TACAs avec des propriétés immunologiques accrues. Notre stratégie de conception a été guidée par (i) l’importance d’une haute densité de carbohydrates pour promouvoir une capture d’antigène plus efficace et un traitement par les cellules présentatrices d’antigène, et (ii) l’expression hétérogène des TACAs au cours de la maladie et parmi différents patients. En respectant ces deux aspects, il sera possible de déclencher une réponse immunitaire plus forte et à plusieurs facettes. / Cancer is one on the leading causes of death in developed countries; although surgical resection, direct irradiation and cytotoxic chemotherapy represent nowadays the main treatment options for patients suffering with malignancies, their severe side effects paved the way for the rise in popularity of antitumoral immunotherapy. Apart from passive immunotherapy, which is comprised of antibodies or other immune system components that are made outside of the body and has been shown to be associated to potentially life threatening immune reactions, we focused our efforts towards active immunotherapy, which purpose is stimulate the patient immune system to selectively eradicate malignant cells. The identification of tumor-associated carbohydrate antigens (TACAs) on the surface of cancer cells has allowed the development of antigen-specific vaccines. It has been known for over four decades that the majority of human cancers are characterized by aberrant glycosylation. Tumor cells may over-express truncated versions of oligosaccharides, unusual terminal oligosaccharide sequences, or increase sialylation of cell-surface glycolipids and O- and N-linked glycoproteins. A truncated oligosaccharide of a glycoprotein may render a part of the peptide backbone, which is normally shielded by the glycan, more accessible to the immune system. Among the assortment of TACAs we focussed our attention on Tn and TF-antigens, which can be found in membrane-bound glycoproteins like MUC-1, over-expressed in more than 90% of breast carcinomas. Although the design of such immuno-modulators still relies on empiric rules, it is noteworthy important to trigger both humoral and cellular responses, and a memory effect. This challenge can be achieved by combining, within a single molecule, carbohydrate antigen expressed on the surface of tumors (B-cell epitope), peptides capable to stimulate both CD4+ and CD8+ T-cells (T-cell epitopes) and an adjuvant, to gather immune system elements in the injection site and boost the antigen uptake. Previous studies of our research group reported for the first time the synthesis and immunological evaluation of a four-component anticancer vaccine prototype capable of inducing a long-lasting immune response in mice models. In my PhD work we aimed to synthesize TACA-based anticancer vaccine prototypes with improved immunological properties. The principles which guided our design strategies rely on (i) the importance of a high density of carbohydrate epitopes to promote a more effective antigen capture and processing by antigen-presenting cells, and (ii) the evidence of heterogenic expression patterns of TACAs during the course of the disease and among different individuals. Addressing these two aspects would provide a stronger and multifaceted immune response.

CHARACTERIZATION OF CARBOHYDRATE BASED VACCINES Variables influencing the immunogenicity and physicochemical properties of glycoconjugate vaccinesMany aspects can influence the immunogenicity of conjugate vaccines and the main variables investigated so far are the size of the saccharide moiety, the saccharide:protein ratio in the purified conjugate, the conjugation strategy, the nature of the spacer and the protein carrier. The size of the saccharide moiety and saccharide/protein ratio were investigated in different works such as Seppälä and Mäkelä in one of the first studies on the effect of size and chemistry on the immunogenicity of dextrans-protein conjugates found that dextrans of low molecular weight conjugated to chicken serum albumin, induced strong anti-dextran responses in mice, while increasing the dextrans' size resulted in reduced immunogenicity.47 Peeters et al. showed that a synthetic tetramer of Hib capsular polysaccharide repeating unit, conjugated to a protein carrier, induced in adult mice and non-human primates antibody levels comparable to a commercial Hib conjugate and higher than those induced by a trimer, indicating that for Hib a minimum of eight sugars is needed for a proper immunological response.48 Laferriere et al. found little influence of the carbohydrate chain length on the immunogenicity of pneumococcal conjugate vaccines in mice.49 Pozsgay et al. studied the immunogenicity in mice of synthetic Shigella dysenteriae type 1 LPS oligosaccharides conjugated to human serum albumin (HSA). The authors found that octa-, dodeca-, and hexadecasaccharide fragments induced high levels of lipopolysaccharide binding IgG antibodies in mice after three injections and were superior to a tetrasaccharide conjugate. The influence of the carbohydrate/protein ratio was different for the three conjugates. The octasaccharide-HSA conjugate with the highest density evoked a good immune response, while in the case of dodeca- and hexadecasaccharide conjugates, the median density was optimal.50 These studies suggest that oligosaccharide chain length and hapten loading might be interconnected in determining the immunogenicity of glycoconjugate vaccines. The spacer is a short linear molecule that is generally linked to the polysaccharide chain or to the protein or to both moieties, depending on the chemistry, used to facilitate the coupling between the protein and sugar. There are evidences in the literature which suggest that rigid, constrained spacers like cyclohexyl maleimide, elicit a significant amount of undesirable antibodies, with the risk of driving the immune response away from the targeted epitope on the hapten.51 52 The use of a flexible alkyl type maleimido spacer has been reported as a way to overcome the previous observed immunogenicity of cyclic maleimide linkers.53 A number of protein carriers have been used so far in preclinical and clinical evaluation of conjugate vaccines. 54 55 56 57 58 59 60Proteins such as diphtheria and tetanus toxoids, which derive from the respective toxins after chemical detoxification with formaldehyde, were initially selected as carrier because of the safety track record accumulated with tetanus and diphtheria vaccination. CRM197, a non-toxic mutant of diphtheria toxin61 which instead does not need chemical detoxification, has been extensively used as carrier for licensed Hib, pneumococcal, meningococcal conjugate vaccines and for other vaccines being developed. An outer membrane protein complex of serogroup B meningococcus has been used by Merck as carrier for their Hib conjugate vaccine.62 GSK in their multivalent pneumococcal conjugate vaccine introduced the use of the Hib-related protein D as carrier for most of the polysaccharides included into the vaccine.63 64 The team of John Robbins made extensive use of the recombinant non toxic form of Pseudomonas aeruginosa exo-toxin as carrier for Staphylococcus aureus type 5 and 8 as well as for Salmonella

Invasive fungal infections are prevalent and often deadly in immunocompromised patients. There continues to be a pressing need for the development of novel antifungal compounds since there are currently only 13 compounds licensed for the treatment of invasive fungal infections and antibiotic-resistant strains have been emerging. CAY-1 is an antifungal steroidal saponin which was isolated from the fruit of the cayenne pepper plant in 0.1% yield. In Vitro studies of CAY-1 have shown it to be an effective antifungal agent against sixteen pathogenic fungal strains and it showed no cytotoxicity toward mammalian cells up to 100 ìg/mL. The development of a practical synthesis of CAY-1 will potentially allow for further exploration of its medicinal utility and provide the opportunity to synthesize derivatives of CAY-1 which could be investigated in structure-activity relationship studies. To this end, methods for the preparation of they CAY-1 aglycone and pentasaccharide moieties have been investigated. Through this work, several partially protected stereoisomers of the CAY-1 aglycone have been prepared which can be used for the synthesis of saponin derivatives of CAY-1 for structure-activity relationship studies. Definitive characterization of one of these isomers, 3á-hydroxy-(22S, 25R)-5á-spirostan-2â-yl acetate, was achieved by X-ray crystallography. Furthermore, a quantitative inversion of the C-3 stereochemical configuration of this compound was achieved via an acetate group migration of the corresponding mesylate. The possibility of competition between the acetate migration and substitution mechanisms with various nucleophiles was explored. The results, however, indicate that this inversion only occurs via the acetate migration. Additionally, the CAY-1 pentasaccharide synthesis poses two significant challenges. First, these results demonstrate that the central 2, 3-branched portion can be synthesized efficiently from a partially protected glucopyranosyl acceptor since the C-2 and C-3 alcohols differ in their reactivity in glycosylation reactions. The second challenge is the ƒÀ-(1¨4) linkage to the galactosyl acceptor which significantly increases the complexity of the synthesis as compared to literature reported syntheses of other branched oligosaccharides. Nonetheless, this ƒÀ-(1¨4) linkage was achieved using a disarmed trichloroacetimidate glucosyl donor.

<p>Glycoconjugates are biologically important molecules with diverse functions. They consist of carbohydrates of varying size and complexity, attached to a non-sugar moiety as a lipid or a protein. Glycoconjugate structures are often very complex and their intricate biosynthetic pathways makes overexpression difficult. This renders the isolation of pure, structurally defined compounds from natural sources cumbersome. Therefore, to better address questions in glycobiology, synthetic glycoconjugates are an appealing alternative. In addition, synthetic methods allow for the preparation of non-natural glycoconjugates that can enhance the understanding of the influence of structural features on the biological responses.</p><p>In this thesis, synthetic methods for the preparation of glycoconjugates, especially glycolipid analogues, have been developed. These methods make use of solid-phase chemistry and are amenable to library synthesis of series of similar compounds. Solid-phase synthesis is a technique where the starting material of the reaction is attached to small plastic beads through a linker. This allows large excess of reagents to speed up the reactions and the sometimes difficult purifications of intermediate products are reduced to simple washings of the beads.</p><p>One problem with solid-phase synthesis is the difficulties to monitor the reactions and characterize the intermediate products. Gel-phase 19 F-NMR spectroscopy, using fluorinated linkers and protecting groups, is an excellent tool to overcome this problem and to monitor solid-phase synthesis of e.g. glycoconjugates. Two novel fluorinated linkers for the attachment of carboxylic acids have been developed and are presented in the thesis. These linkers can be cleaved with both acids of varying strengths and nucleophiles like hydroxide ions, and they are stable to glycosylation conditions. In addition, a novel filter reactor for solid-phase synthesis was designed. The reactor fits into an ordinary NMR spectrometer to facilitate the reaction monitoring with gel-phase 19 F-NMR spectroscopy.</p><p>The biological applications of the synthesized glycolipids were demonstrated in two different settings. The CD1d restricted binding of glycolipids carrying the monosaccharide α-GalNAc as carbohydrate could be detected on viable cells of mouse origin. CD1d is one of several antigen presenting molecules (the CD1 proteins) that presents lipids and glycolipids to circulating T-cells that in turn can initiate an immune response. The CD1 molecules are relatively sparsely investigated, and the method to measure glycolipid binding on viable cells, as described in the thesis, has the possibility to greatly enhance the knowledge of the structural requirements for CD1-binding.</p><p>Serine-based neoglycolipids with terminal carboxylic acids were used to prepare glycoconjugate arrays with covalent bonds to secondary amines on microtiter plates. Carbohydrate arrays have great possibilities to simplify the study of interactions between carbohydrates and e.g. proteins and microbes. The usefulness of the glycolipid arrays constructed in the thesis was illustrated with two lectins, RCA120 from Ricinus communis and BS-1 from Bandeiraea simplicifolia. Both lectins bound to the array of neoglycolipids in agreement with their respective specificity for galactosides.</p><p>Glycobiology is a large area of great interest and the methods described in this thesis can be used to answer a variety of glycoconjugaterelated biological questions.</p>

Glycoconjugates are biologically important molecules with diverse functions. They consist of carbohydrates of varying size and complexity, attached to a non-sugar moiety as a lipid or a protein. Glycoconjugate structures are often very complex and their intricate biosynthetic pathways makes overexpression difficult. This renders the isolation of pure, structurally defined compounds from natural sources cumbersome. Therefore, to better address questions in glycobiology, synthetic glycoconjugates are an appealing alternative. In addition, synthetic methods allow for the preparation of non-natural glycoconjugates that can enhance the understanding of the influence of structural features on the biological responses. In this thesis, synthetic methods for the preparation of glycoconjugates, especially glycolipid analogues, have been developed. These methods make use of solid-phase chemistry and are amenable to library synthesis of series of similar compounds. Solid-phase synthesis is a technique where the starting material of the reaction is attached to small plastic beads through a linker. This allows large excess of reagents to speed up the reactions and the sometimes difficult purifications of intermediate products are reduced to simple washings of the beads. One problem with solid-phase synthesis is the difficulties to monitor the reactions and characterize the intermediate products. Gel-phase 19 F-NMR spectroscopy, using fluorinated linkers and protecting groups, is an excellent tool to overcome this problem and to monitor solid-phase synthesis of e.g. glycoconjugates. Two novel fluorinated linkers for the attachment of carboxylic acids have been developed and are presented in the thesis. These linkers can be cleaved with both acids of varying strengths and nucleophiles like hydroxide ions, and they are stable to glycosylation conditions. In addition, a novel filter reactor for solid-phase synthesis was designed. The reactor fits into an ordinary NMR spectrometer to facilitate the reaction monitoring with gel-phase 19 F-NMR spectroscopy. The biological applications of the synthesized glycolipids were demonstrated in two different settings. The CD1d restricted binding of glycolipids carrying the monosaccharide α-GalNAc as carbohydrate could be detected on viable cells of mouse origin. CD1d is one of several antigen presenting molecules (the CD1 proteins) that presents lipids and glycolipids to circulating T-cells that in turn can initiate an immune response. The CD1 molecules are relatively sparsely investigated, and the method to measure glycolipid binding on viable cells, as described in the thesis, has the possibility to greatly enhance the knowledge of the structural requirements for CD1-binding. Serine-based neoglycolipids with terminal carboxylic acids were used to prepare glycoconjugate arrays with covalent bonds to secondary amines on microtiter plates. Carbohydrate arrays have great possibilities to simplify the study of interactions between carbohydrates and e.g. proteins and microbes. The usefulness of the glycolipid arrays constructed in the thesis was illustrated with two lectins, RCA120 from Ricinus communis and BS-1 from Bandeiraea simplicifolia. Both lectins bound to the array of neoglycolipids in agreement with their respective specificity for galactosides. Glycobiology is a large area of great interest and the methods described in this thesis can be used to answer a variety of glycoconjugaterelated biological questions.

No description available.

No description available.