scholarly journals Antigenic determinants of a plant proteoglycan, the Gladiolus style arabinogalactan-protein

1980 ◽  
Vol 191 (2) ◽  
pp. 437-447 ◽  
Author(s):  
P A Gleeson ◽  
A E Clarke
Keyword(s):  
Potent Inhibitor ◽  
Cross Reactivity ◽  
Arabinogalactan Protein ◽  
Galactose Oxidase ◽  
Antigenic Specificity ◽  
Specific Antiserum ◽  
Antigenic Determinants ◽  
Carbohydrate Component ◽  
Terminal Galactose ◽  
Galactosyl Residues

Antiserum has been raised to the arabinogalactan-protein of Gladiolus style mucilage. This macromolecule has been characterized and has a structure consistent with a 1 leads to 3-linked beta-galactan backbone with side branches of 1 leads to 6-linked beta-galactosyl residues, some of which carry terminal alpha-L-arabinofuranoside residues [Gleeson & Clarke (1979) Biochem. J. 181, 607-621]. The specificity of the antiserum has been investigated by immunoprecipitation with [3H]arabinogalactan-protein. THe 3H label was introduced into the arabinogalactan-protein by oxidation of the terminal galactose residues with galactose oxidase, followed by reduction with NaB3H4. The antigenic specificity of the antiserum was shown to be directed towards the carbohydrate component of the arabinogalactan-protein. D-galactose and L-arabinose were the most effective hapten inhibitors of the antiserum; other monosaccharides, N-acetyl-D-galactono-1,4-lactone, D-glucose, D-mannose, L-rhamnose. L-fucose and D-xylose, were all poor inhibitors. The antiserum showed preference for beta-galactosides over alpha-galactosides. Of the haptens examined, the disaccharide 6-O-beta-D-galactopyranosyl-D-galactopyranose was the most potent inhibitor. The antigenic features of the arabinogalactan-protein were investigated by examining the interaction of the antiserum with chemically and enzymically modified arabinogalactan-protein. Also, the cross-reactivity of structurally related polysaccharides and glycoproteins with the specific antiserum was assessed by a haemagglutination assay using erythrocytes coupled with specific antiserum. The results indicate that the dominant antigenic determinants of the arabinogalactan-protein are probably the side branches of 1 leads to 6 -linked beta-galactose residues bearing the terminal alpha-L-arabinose residues.

The concept and operational definition of protein epitopes

1989 ◽  
Vol 323 (1217) ◽  
pp. 451-466 ◽  
Keyword(s):  
Synthetic Peptides ◽  
Foot And Mouth Disease ◽  
Operational Definition ◽  
Cross Reactivity ◽  
Antigenic Specificity ◽  
Antigenic Determinants ◽  
Immune Recognition ◽  
Segmental Mobility ◽  
X Ray Crystallography ◽  
Relative Contribution

The antigenic determinants or epitopes of a protein correspond to those parts of the molecule that are specifically recognized by the binding sites or paratopes of certain immunoglobulin molecules. Epitopes are thus relational entities that require complementary paratopes for their operational recognition. Some authors consider that the concept of epitope necessarily involves the two properties of antigenic reactivity (ability to bind to a paratope) and immunogenicity (ability to induce an immune response). Such a view creates difficulties because it makes the existence of epitopes in a protein depend on immunogenetic and regulatory mechanisms of the immunized host. The delineation of epitopes can be achieved by antigenic cross-reactivity studies or by X-ray crystallography. Both approaches require specific criteria for deciding which residues of the antigen are in contact with the paratope and are functionally part of the epitope. The relative contribution of static accessibility, segmental mobility and induced fit to immune recognition remains controversial. Each of the methods used for analysing antigenic specificity is subject to various operational constraints originating from the type of experimental probe and from the form at, sensitivity and specificity of the immunoassay used. If a protein is assumed to contain as many epitopes as the number of different monoclonal antibodies that can be raised against it, the delineation of epitopes corresponds to the summation in various hosts of the immune repertoire specific for the antigen. Neutralization epitopes are a special subclass of the epitopes of infectious agents and toxins that are specifically recognized by antibody molecules able to neutralize the biological activity of the antigen. The identification of neutralization epitopes is important for the development of synthetic vaccines because it is this type of epitope that should be mimicked by synthesis and used as a vaccine for eliciting protective immunity. The first demonstration that synthetic peptides could elicit antibodies that neutralized viral infectivity was made by Anderer and his colleagues in the 1960s in their work with tobacco mosaic virus. Nearly 20 years passed before it was shown that antibodies to synthetic peptides were also able to neutralize the infectivity of other viruses such as foot-and-mouth disease, polio and hepatitis B viruses.

scholarly journals Characterization of components of Z-bands in the fibrillar flight muscle of Drosophila melanogaster.

1989 ◽  
Vol 109 (5) ◽  
pp. 2157-2167 ◽  
Author(s):  
J D Saide ◽  
S Chin-Bow ◽  
J Hogan-Sheldon ◽  
L Busquets-Turner ◽  
J O Vigoreaux ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Monoclonal Antibodies ◽  
Flight Muscle ◽  
Cross Reactivity ◽  
Antigenic Determinants ◽  
Immunogold Staining ◽  
Thick Filaments ◽  
The Matrix ◽  
Flight Muscles

Twelve monoclonal antibodies have been raised against proteins in preparations of Z-disks isolated from Drosophila melanogaster flight muscle. The monoclonal antibodies that recognized Z-band components were identified by immunofluorescence microscopy of flight muscle myofibrils. These antibodies have identified three Z-disk antigens on immunoblots of myofibrillar proteins. Monoclonal antibodies alpha:1-4 recognize a 90-100-kD protein which we identify as alpha-actinin on the basis of cross-reactivity with antibodies raised against honeybee and vertebrate alpha-actinins. Monoclonal antibodies P:1-4 bind to the high molecular mass protein, projectin, a component of connecting filaments that link the ends of thick filaments to the Z-band in insect asynchronous flight muscles. The anti-projectin antibodies also stain synchronous muscle, but, surprisingly, the epitopes here are within the A-bands, not between the A- and Z-bands, as in flight muscle. Monoclonal antibodies Z(210):1-4 recognize a 210-kD protein that has not been previously shown to be a Z-band structural component. A fourth antigen, resolved as a doublet (approximately 400/600 kD) on immunoblots of Drosophila fibrillar proteins, is detected by a cross reacting antibody, Z(400):2, raised against a protein in isolated honeybee Z-disks. On Lowicryl sections of asynchronous flight muscle, indirect immunogold staining has localized alpha-actinin and the 210-kD protein throughout the matrix of the Z-band, projectin between the Z- and A-bands, and the 400/600-kD components at the I-band/Z-band junction. Drosophila alpha-actinin, projectin, and the 400/600-kD components share some antigenic determinants with corresponding honeybee proteins, but no honeybee protein interacts with any of the Z(210) antibodies.

scholarly journals IMMUNOLOGIC STUDIES OF WATER-SOLUBLE HUMAN AMYLOID FIBRILS

1969 ◽  
Vol 130 (4) ◽  
pp. 797-808 ◽  
Author(s):  
Edward C. Franklin ◽  
Mordechai Pras
Keyword(s):  
Human Serum ◽  
Comparative Studies ◽  
Amyloid Fibrils ◽  
Complement Fixation ◽  
Cross Reactivity ◽  
Water Soluble ◽  
Antigenic Determinants ◽  
Normal Tissues ◽  
Absorption Studies ◽  
Normal Human

Eight preparations of soluble amyloid and degraded amyloid (DAM) were compared immunologically. Unlike amyloid fibrils, six of eight preparations of DAM proved to be relatively strong immunogens. Antisera to DAM reacted weakly or not at all with normal human serum or extracts of normal tissues, but were specifically reactive with amyloid fibrils or DAM. Comparative studies of DAM'S from eight different subjects showed some degree of cross-reactivity among them, yet demonstrated that they were not identical. Similar conclusions were obtained by quantitative precipitin and complement fixation analyses. Comparison of the amyloid fibrils with the homologous DAM by complement fixation and absorption studies demonstrated the existence in DAM of antigenic determinants that were lacking or inaccessible in the native fibrils. A search for amyloid precursors and antibodies to amyloid in the sera of 12 patients proved unsuccessful.

Cross-reactivity between mammalian myoglobins: Linear vs spatial antigenic determinants

1977 ◽  
Vol 14 (4) ◽  
pp. 283-288 ◽  
Author(s):  
John G.R. Hurrell ◽  
John A. Smith ◽  
Pamela E. Todd ◽  
Sydney J. Leach
Keyword(s):  
Cross Reactivity ◽  
Antigenic Determinants

scholarly journals Chemo-enzymatic modification of poly-N-acetyllactosamine (LacNAc) oligomers and N,N-diacetyllactosamine (LacDiNAc) based on galactose oxidase treatment

2012 ◽  
Vol 8 ◽  
pp. 712-725 ◽  
Author(s):  
Christiane E Kupper ◽  
Ruben R Rosencrantz ◽  
Birgit Henßen ◽  
Helena Pelantová ◽  
Stephan Thönes ◽  
...  
Keyword(s):  
Reductive Amination ◽  
Chemical Conversion ◽  
Product Formation ◽  
Galactose Oxidase ◽  
Binding Motif ◽  
Enzymatic Modification ◽  
Specific Interest ◽  
Subsequent Reduction ◽  
Binding Studies ◽  
Terminal Galactose

The importance of glycans in biological systems is highlighted by their various functions in physiological and pathological processes. Many glycan epitopes on glycoproteins and glycolipids are based on N-acetyllactosamine units (LacNAc; Galβ1,4GlcNAc) and often present on extended poly-LacNAc glycans ([Galβ1,4GlcNAc] n ). Poly-LacNAc itself has been identified as a binding motif of galectins, an important class of lectins with functions in immune response and tumorigenesis. Therefore, the synthesis of natural and modified poly-LacNAc glycans is of specific interest for binding studies with galectins as well as for studies of their possible therapeutic applications. We present the oxidation by galactose oxidase and subsequent chemical or enzymatic modification of terminal galactose and N-acetylgalactosamine residues of poly-N-acetyllactosamine (poly-LacNAc) oligomers and N,N-diacetyllactosamine (LacDiNAc) by galactose oxidase. Product formation starting from different poly-LacNAc oligomers was characterised and optimised regarding formation of the C6-aldo product. Further modification of the aldehyde containing glycans, either by chemical conversion or enzymatic elongation, was established. Base-catalysed β-elimination, coupling of biotin–hydrazide with subsequent reduction to the corresponding hydrazine linkage, and coupling by reductive amination to an amino-functionalised poly-LacNAc oligomer were performed and the products characterised by LC–MS and NMR analysis. Remarkably, elongation of terminally oxidised poly-LacNAc glycans by β3GlcNAc- and β4Gal-transferase was also successful. In this way, a set of novel, modified poly-LacNAc oligomers containing terminally and/or internally modified galactose residues were obtained, which can be used for binding studies and various other applications.

scholarly journals Molecular characterization of a fungal cyclophilin allergen Rhi o 2 and elucidation of antigenic determinants responsible for IgE–cross-reactivity

2019 ◽  
Vol 295 (9) ◽  
pp. 2736-2748 ◽  
Author(s):  
Gaurab Sircar ◽  
Moumita Bhowmik ◽  
Rajat Kanti Sarkar ◽  
Nazanin Najafi ◽  
Angira Dasgupta ◽  
...  
Keyword(s):  
Rhizopus Oryzae ◽  
Allergic Sensitization ◽  
Melting Curve ◽  
Cross Reactivity ◽  
Antigenic Determinants ◽  
Heat Denaturation ◽  
Homologous Sequence ◽  
Ppiase Activity ◽  
Ige Reactivity ◽  
C Terminus

Cyclophilins are structurally conserved pan-allergens showing extensive cross-reactivity. So far, no precise information on cross-reactive IgE-epitopes of cyclophilins is available. Here, an 18-kDa IgE-reactive cyclophilin (Rhi o 2) was purified from Rhizopus oryzae, an indoor mold causing allergic sensitization. Based on LC-MS/MS-derived sequences of natural Rhi o 2, the full-length cDNA was cloned, and expressed as recombinant (r) allergen. Purified rRhi o 2 displayed IgE-reactivity and basophil degranulation with sera from all cyclophilin-positive patients. The melting curve of properly folded rRhi o 2 showed partial refolding after heat denaturation. The allergen displayed monomeric functional peptidyl-prolyl cis-trans isomerase (PPIase) activity. In IgE-inhibition assays, rRhi o 2 exhibited extensive cross-reactivity with various other cyclophilins reported as allergens from diverse sources including its homologous human autoantigen. By generating a series of deletion mutants, a conserved 69-residue (Asn81-Asn149) fragment at C terminus of Rhi o 2 was identified as crucial for IgE-recognition and cross-reactivity. Grafting of the Asn81-Asn149 fragment within the primary structure of yeast cyclophilin CPR1 by replacing its homologous sequence resulted in a hybrid molecule with structural folds similar to Rhi o 2. The IgE-reactivity and allergenic activity of the hybrid cyclophilin were greater than that of CPR1. Therefore, the Asn81-Asn149 fragment can be considered as the site of IgE recognition of Rhi o 2. Hence, Rhi o 2 serves as a candidate antigen for the molecular diagnosis of mold allergy, and determination of a major cross-reactive IgE-epitope has clinical potential for the design of next-generation immunotherapeutics against cyclophilin-induced allergies.

Viral Cross-Reactivity and Antigenic Determinants Recognized by Human Parainfluenza Virus Type 1-Specific Cytotoxic T-Cells

Virology ◽  
1994 ◽  
Vol 199 (2) ◽  
pp. 376-383 ◽  
Author(s):  
Vibhuti P. Dave ◽  
Jane E. Allan ◽  
Karen S. Slobod ◽  
F.Suzette Smith ◽  
Kevin W. Ryan ◽  
...  
Keyword(s):  
T Cells ◽  
Virus Type ◽  
Cytotoxic T Cells ◽  
Cross Reactivity ◽  
Parainfluenza Virus ◽  
Antigenic Determinants ◽  
Human Parainfluenza Virus

scholarly journals Simultaneous Rapid Detection and Serotyping of Cronobacter sakazakii Serotypes O1, O2, and O3 by Using Specific Monoclonal Antibodies

2016 ◽  
Vol 82 (8) ◽  
pp. 2300-2311 ◽  
Author(s):  
Eva J. Scharinger ◽  
Richard Dietrich ◽  
Ina Kleinsteuber ◽  
Erwin Märtlbauer ◽  
Kristina Schauer
Keyword(s):  
Monoclonal Antibodies ◽  
Rapid Detection ◽  
Foodborne Pathogen ◽  
Cross Reactivity ◽  
Antigenic Determinants ◽  
Powdered Infant Formula ◽  
Bacterial Cells ◽  
Cronobacter Sakazakii ◽  
Content Type ◽  
Cell Counts

ABSTRACTCronobacter sakazakiiis a foodborne pathogen associated with rare but often lethal infections in neonates. Powdered infant formula (PIF) represents the most frequent source of infection. Out of the identified serotypes (O1 to O7), O1, O2, and O3 are often isolated from clinical and PIF samples. Serotype-specific monoclonal antibodies (MAbs) suitable for application in enzyme immunoassays (EIAs) for the rapid detection ofC. sakazakiihave not yet been developed. In this study, we created specific MAbs with the ability to bind toC. sakazakiiof serotypes O1, O2, and O3. Characterization by indirect EIAs, immunofluorescence, motility assays, and immunoblotting identified lipopolysaccharide (LPS) and exopolysaccharide (EPS) as the antigenic determinants of the MAbs. The established sandwich EIAs were highly sensitive and were able to detect between 2 × 103and 9 × 106CFU/ml. Inclusivity tests confirmed that 93% of serotype O1 strains, 100% of O2 strains, and 87% of O3 strains were detected at low cell counts. No cross-reactivity with >100 strains ofCronobacterspp. and otherEnterobacteriaceaewas observed, except for that withC. sakazakiiserotype O3 andCronobactermuytjensiiserotype O1. Moreover, the sandwich EIAs detectedC. sakazakiiin PIF samples artificially contaminated with 1 to 10 bacterial cells per 10 g of sample after 15 h of preenrichment. The use of these serotype-specific MAbs not only allows the reliable detection ofC. sakazakiistrains but also enables simultaneous serotyping in a simple sandwich EIA method.

THE ANTIGENIC DETERMINANT OF THE LH-RELEASING HORMONE FOR THREE DIFFERENT ANTISERA

1975 ◽  
Vol 78 (1) ◽  
pp. 222-231 ◽  
Author(s):  
A. Arimura ◽  
H. Sato ◽  
D. H. Coy ◽  
R. B. Worobec ◽  
A. V. Schally ◽  
...  
Keyword(s):  
Serum Albumin ◽  
Rabbit Antiserum ◽  
Cross Reactivity ◽  
Amide Group ◽  
Antigenic Determinants ◽  
Releasing Hormone ◽  
C Terminus ◽  
Lh Rh ◽  
Lh Releasing Hormone ◽  
Antigen Antibody

ABSTRACT Antigenic determinants of LH-releasing hormone (LH-RH) were investigated by testing the cross-reaction of LH-RH analogues and fragments in LH-RH radioimmunoassay (RIA) systems using 3 different antisera against the LH-RH decapeptide. Rabbit antiserum No. 419 was generated against LH-RH adsorbed on polyvinylpyrrolidone (PVP). Antisera Nos. 710 and 742 were produced by immunizing rabbits with LH-RH conjugated either with bovine serum albumin through its C-terminus, or with human serum albumin through the N-terminus, respectively. For antiserum No. 419, the N-terminal (pyro)-glutamic acid and/or histidine in positions 1 and 2 of LH-RH, respectively, were found to enhance the antigen-antibody interaction, but were not indispensable for it. Similarly, the C-terminal amide and glycine-NH2 did not play a major role in these interactions. The LH-RH heptapeptide fragment, corresponding to amino acid sequence from positions 3 to 9, showed a cross-reactivity in this RIA system with LH-RH, although greater amounts than those of cold LH-RH were required for a comparable inhibition of binding of labelled LH-RH. For antiserum No. 710, the LH-RH hexapeptide fragment corresponding to positions 2 to 7 showed considerable cross-reac tivity. Histidine in position 2 played an important role but neither the amide group nor the glycine amide group at the C-terminus were essential. For antiserum No. 742, the C-terminal tetrapeptide-amide fragment of LH-RH showed considerable cross-reactivity in the LH-RH, the amide moiety itself being of crucial importance. These antisera may be useful in investigating peptides related to LH-RH in biological materials.

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