Search results for "Epitope Mapping"
showing 10 items of 28 documents
Monoclonal anti-fosB antibody specific for predetermined, nonstructural region of the fosB protein.
1997
Comparison of the primary structures and theoretical prediction of the potential antigenic determinant of the deduced Fos proteins reveals the presence of a nonstructural and hydrophilic region juxtaposed to the leucine zipper and nonconserved among the Fos protein family. To develop monoclonal anti-peptide antibodies capable of distinguishing all Fos-proteins, synthetic peptides specific for the mentioned predicted region were synthesized manually by the "tea-bag" method. Immunization of Balb/c mice with fosB-related synthetic peptide BSA gave rise to mouse hybridoma cell line K21 (IgG1, kappa) secreting highly specific antibodies against corresponding human fosB protein. Fine mapping of t…
The Major Conformational IgE-binding Epitopes of Hevein (Hev b6.02) Are Identified by a Novel Chimera-based Allergen Epitope Mapping Strategy
2002
A novel approach to localize and reconstruct conformational IgE-binding epitope regions of hevein (Hev b6.02), a major natural rubber latex allergen, is described. An antimicrobial protein (AMP) from the amaranth Amaranthus caudatus was used as an immunologically non-IgE-binding adaptor molecule to which terminal or central parts of hevein were fused. Hevein and AMP share a structurally identical core region but have different N-terminal and C-terminal regions. Only 1 of 16 hevein-allergic patients showed weak IgE binding to purified native or recombinant AMP. Chimeric AMP with the hevein N terminus was recognized by IgE from 14 (88%) patients, and chimeric AMP with the hevein C terminus wa…
Identification and quantification of a new family of peptide endocannabinoids (Pepcans) showing negative allosteric modulation at CB1 receptors.
2012
The α-hemoglobin-derived dodecapeptide RVD-hemopressin (RVDPVNFKLLSH) has been proposed to be an endogenous agonist for the cannabinoid receptor type 1 (CB(1)). To study this peptide, we have raised mAbs against its C-terminal part. Using an immunoaffinity mass spectrometry approach, a whole family of N-terminally extended peptides in addition to RVD-Hpα were identified in rodent brain extracts and human and mouse plasma. We designated these peptides Pepcan-12 (RVDPVNFKLLSH) to Pepcan-23 (SALSDLHAHKLRVDPVNFKLLSH), referring to peptide length. The most abundant Pepcans found in the brain were tested for CB(1) receptor binding. In the classical radioligand displacement assay, Pepcan-12 was th…
Generation of monoclonal antibodies of desired specificity using chimeric polyomavirus-derived virus-like particles.
2005
Foreign protein sequences presented on hamster polyomavirus (HaPyV) major capsid protein VP1-derived virus-like particles (VLPs) have been demonstrated to be highly immunogenic. The current study was aimed to evaluate VP1-derived chimeric VLPs as tools for hybridoma technology to generate monoclonal antibodies (mAbs) of desired specificity. Chimeric VLPs containing inserts of different size and origin were used as immunogens. Chimeric VLPs carrying a 9 amino acid (aa)-long cytotoxic T-cell epitope (STAPPVHNV) of human mucin 1 (MUC1) elicited a strong epitope-specific humoral immune response in mice and promoted the production of MUC1-specific mAbs. From a total of seven mAbs of IgG isotype …
Anti-ADAMTS13 autoantibody profiling in patients with immune-mediated thrombotic thrombocytopenic purpura.
2021
Anti-A Disintegrin and Metalloproteinase with a ThromboSpondin type 1 motif, member 13 (ADAMTS13) autoantibodies cause a severe ADAMTS13 deficiency in immune-mediated thrombotic thrombocytopenic purpura (iTTP). ADAMTS13 consists of a metalloprotease (M), a disintegrin-like (D) domain, 8 thrombospondin type 1 repeats (T1-T8), a cysteine-rich (C), a spacer (S), and 2 CUB domains (CUB1-2). We recently developed a high-throughput epitope mapping assay based on small, nonoverlapping ADAMTS13 fragments (M, DT, CS, T2-T5, T6-T8, CUB1-2). With this assay, we performed a comprehensive epitope mapping using 131 acute-phase samples and for the first time a large group of remission samples (n = 50). Ne…
Multiple IgE recognition on the major allergen of the Parietaria pollen Par j 2
2015
The interaction between IgE antibodies and allergens is a key event in triggering an allergic reaction. The characterization of this region provides information of paramount importance for diagnosis and therapy. Par j 2 Lipid Transfer Protein is one of the most important allergens in southern Europe and a well-established marker of sensitization in Parietaria pollen allergy. The main aim of this study was to map the IgE binding regions of this allergen and to study the pattern of reactivity of individual Parietaria-allergic patients. By means of gene fragmentation, six overlapping peptides were expressed in Escherichia coli, and their IgE binding activity was evaluated by immunoblotting in …
Participation of Two Ser-Ser-Phe-Tyr Repeats in Interleukin-6 (IL-6)-Binding Sites of the Human IL-6 Receptor
1996
The alpha-subunit of interleukin-6 (IL-6) receptor is a member of the hematopoietin receptor family. The alignment of its amino acid sequence with those of other members of this family (human somatotropin receptor/murine IL-3 receptor beta and human IL-2 receptor beta) has suggested that amino acids included in two SSFY repeats found in each of its hematopoietin receptor domains, contribute to the binding of the ligand. The involvement of these amino acids in IL-6 binding and signal transduction was studied by site-directed mutagenesis and molecular modelling. We present a computer-derived three-dimensional model of the IL-6/IL-6 receptor complex based on the structure of the human somatotr…
Analysis of Epitope Spreading over an Eleven-year Period in a Patient with Systemic Lupus Erythematosus: CASE REPORT
1998
During a period of more than eleven years serum samples of a patient with Systemic Lupus Erythematosus were collected and analyzed for anti-nuclear autoantibodies. High titer of anti-La/SS-B were detectable in all serum samples. The La/SS-B epitopes remained constant. Besides anti-La/SS-B antibodies all serum samples contained traces of anti-Ro/SS-A including anti-Ro52 and anti-Ro60 antibodies. During disease flares anti-Ro/SS A antibodies were upregulated and anti-dsDNA antibodies appeared, thus supporting the concept of an antigen driven intermolecular epitope spreading to Ro/SS-A and dsDNA.
Identification of T cell epitopes by the use of rapidly generated mRNA fragments.
2004
Abstract Although the number of defined T cell epitopes of clinically relevant antigens is constantly increasing, there is still an enormous need to identify further peptides, processed from new antigens or presented by rare HLA molecules, respectively. Here we introduce a novel two-step approach for the rapid identification of T cell epitopes. It was established in the CMV infection model. From the peripheral blood of healthy donors sharing HLA-A1 according to HLA serotyping we isolated CD8 + T lymphocytes and generated dendritic cells (DCs). DCs were electroporated with CMV pp65 mRNA and tested for recognition by autologous CD8 + T lymphocytes in IFN-γ ELISPOT assays. In all 10 CMV-seropo…
Immunogenic hotspots in the spacer domain of ADAMTS13 in immune‐mediated thrombotic thrombocytopenic purpura
2021
International audience; Background Immune-mediated thrombotic thrombocytopenic purpura (iTTP) is caused by anti-ADAMTS13 autoantibodies inducing a severe deficiency of ADAMTS13. Epitope mapping studies on samples obtained during acute iTTP episodes have shown that the iTTP immune response is polyclonal, with almost all patients having autoantibodies targeting the spacer domain of ADAMTS13.Objectives To identify the immunogenic hotspots in the spacer domain of ADAMTS13.Patients/methods A library of 11 full-length ADAMTS13 spacer hybrids was created in which amino acid regions of the spacer domain of ADAMTS13 were exchanged by the corresponding region of the spacer domain of ADAMTS1. Next, th…