title: IgE and IgG epitope mapping by microarray peptide-immunoassay reveals the importance and diversity of the immune response to the IgG3 equine immunoglobulin.
abstract: The presence of whole horse IgG in therapeutic snake antivenom preparations of high purity is a contamination that can cause IgE-mediated allergic reactions in patients. In this study, the immunodominant IgE and IgG-binding epitopes in horse heavy chain IgG3 were mapped using arrays of overlapping peptides synthesized directly onto activated cellulose membranes. Pooled human sera from patients with and without horse antivenom allergies were used to probe the membrane. We have demonstrated that, for both cases, individuals produce antibodies to epitopes of sequential amino acids of horse heavy chain IgG3, although the signal strength and specificity appear to be distinct between the two groups of patients. A single region was found to contain the dominant allergic IgE epitope. The critical residues involved in the binding of human IgE to the epitope were determined to include four hydrophobic amino acids followed by polar and charged residues that formed a coil structure. This is the first study to describe the specific amino acid sequences involved with the immune recognition of human IgG and IgE to horse antivenom.
authors: Center for Technological Development in Health (CDTS) National Institute of Science and Technology for Innovation in Neglected Diseases (INCT-IDN), FIOCRUZ, Rio de Janeiro, RJ, Brazil; Molecular and Cellular Biology Department, Federal Fluminense University, Niterói, RJ, Brazil. Electronic address: salvatore.giovanni@pq.cnpq.br.
authors: Salvatore G
authors: SG
authors: De-Simone
authors: Molecular and Cellular Biology Department, Federal Fluminense University, Niterói, RJ, Brazil.
authors: Paloma
authors: P
authors: Napoleão-Pêgo
authors: Molecular and Cellular Biology Department, Federal Fluminense University, Niterói, RJ, Brazil.
authors: Luiz A L
authors: LA
authors: Teixeira-Pinto
authors: Research and Development Laboratory, Vital Brazil Institute, Niterói, RJ, Brazil.
authors: Anibal R
authors: AR
authors: Melgarejo
authors: Research and Development Laboratory, Vital Brazil Institute, Niterói, RJ, Brazil.
authors: Aniesse S
authors: AS
authors: Aguiar
authors: Center for Technological Development in Health (CDTS) National Institute of Science and Technology for Innovation in Neglected Diseases (INCT-IDN), FIOCRUZ, Rio de Janeiro, RJ, Brazil.
authors: David W
authors: DW
authors: Provance
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology; D019295:Computational Biology; D018604:Epitope Mapping / Q000379:methods / Q000662:veterinary*; D014644:Genetic Variation; D006736:Horses / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007118:Immunoassay / Q000662:veterinary; D007073:Immunoglobulin E / Q000235:genetics*; D007074:Immunoglobulin G / Q000235:genetics* / Q000276:immunology; D046228:Microarray Analysis / Q000662:veterinary; D008969:Molecular Sequence Data
chemical_list: D000906:Antibodies; D007074:Immunoglobulin G; D007073:Immunoglobulin E
keywords: 3,3′,5,5′-tetramethylbenzidine; 50 mM citrate-buffer saline; Anaphylactic reaction; B-linear epitopes; CBS; ELISA; HCS; HRP; Horse immunoglobulin; IgE epitope; IgG epitope; NST; ST; Spot-synthesis; TBS-CT; TBS-T; TMB; enzyme linked immunosorbent assay; hIg; health control sera; hhcIg; horse heavy chain immunoglobulin; horse immunoglobulin; horseradish peroxidase; not sensitive treatment; sensitive treatment; tris-buffer saline containing 3% casein and 0.1% Tween 20, pH 7.0; tris-buffer saline, 0.1% Tween 20, pH 7.0