Monoclonal anti-equine IgE antibodies with specificity for different epitopes on the immunoglobulin heavy chain of native IgE.
Abstract: In this study we describe the generation of monoclonal antibodies (mAbs), which recognize different epitopes of the equine IgE constant heavy chain. Equi-murine recombinant IgE (rIgE), composed of the murine V(H)186.2 heavy chain variable region, linked to the equine IgE constant heavy chain and expressed together with the murine lambda(1) chain in J558L cells was used to immunize BALB/C mice. A total of 17 different mAbs were obtained, which recognized the rIgE heavy chain constant region. None of the mAbs reacted with monoclonal equine isotypes IgM, IgG1 (IgGa), IgG3 (IgG(T)), IgG4 (IgGb) or isolated equine light chains, IgGc and IgA from horse serum, or the native mAb B1-8delta, expressing the same heavy chain variable regions and light chains. One of the mAbs (alphaIgE-132) recognized the recombinant equine IgE, but did not recognize any protein in equine serum, i.e. native IgE. A total of 16 mAbs detected a serum protein of approximately 210,000Da on Western blots, corresponding to the expected MW of native IgE. In addition, one of the mAbs (alphaIgE-176) detected a protein of 76,000Da under reducing conditions, most likely the equine IgE heavy chain. According to binding inhibition studies, the equine IgE specific mAbs recognize at least two different epitopes of the equine IgE. In an ELISA using two anti-IgE mAbs which recognized different epitopes, no significant differences in the concentration of total serum IgE could be detected between adult Icelandic horses with IgE-mediated type I allergy (summer eczema) and healthy control animals. In Icelandic horse foals, no serum IgE could be measured 6 months post partum. All anti-IgE mAbs recognized a small population (1.3+/-0.5%) of leukocytes from adult Icelandic horses by surface immunofluorescence, but no cells could be detected in foal blood. The stained leukocytes from adult horses could be enriched by magnetic cell sorting and contained 32% basophils, 53% monocytes and/or large lymphocytes, 13% small lymphocytes and 2% eosinophils.
Publication Date: 2003-03-12 PubMed ID: 12628763DOI: 10.1016/s0165-2427(03)00007-2Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This research involves the production of specific monoclonal antibodies against different sites on the constant heavy chain part of equine (horse) IgE, a type of immunoglobulin involved in allergic reactions. The study found no significant difference in total serum IgE concentrations between adult Icelandic horses suffering from type I allergy and healthy controls, and detected the presence of IgE on certain white blood cells in adult, but not young, horses.
Background and Methods
- The researchers utilized recombinant IgE (genetically engineered antibodies) made up of the heavy chain variable region from a mouse and the constant heavy chain from a horse IgE. This was expressed along with a particular type of mouse light chain in a mammalian cell line.
- They then vaccinated mice with this material, causing the mice’s immune system to manufacture antibodies against the horse IgE components of the recombinant IgE molecules.
- A total of 17 distinct monoclonal antibodies (mAbs) were produced, each of which recognizes different parts (epitopes) of the equine IgE constant heavy chain.
Findings
- None of these mAbs reacted with other types of horse antibodies (IgM, IgG1, IgG3, IgG4, IgGc, IgA) or the isolated light chains of these other antibodies.
- The researchers were able to use these mAbs to detect both native (natural, non-recombinant) horse IgE and the recombinant horse IgE they created.
- Binding inhibition studies showed that the mAbs recognize at least two different epitopes on the equine IgE heavy chain.
- An enzyme-linked immunoassay (ELISA) test found no significant differences in total IgE levels in the blood between adult horses with summer eczema (a type I allergy) and healthy horses.
- However, they found a small population of leukocytes (white blood cells) in adult horses that are recognized by these mAbs. These cells are believed to contain IgE on their surface.
Significance
- The monocyte and lymphocyte populations that are bound by anti-IgE mAbs could provide an important clue to understanding the role of this immunoglobulin in allergic reactions.
- Understanding the mechanism of IgE responses in horses may help veterinarians to better diagnose and treat equine allergies.
- These antibodies can also be used in the future to further study the structure and function of IgE in horses.
Cite This Article
APA
Wagner B, Radbruch A, Rohwer J, Leibold W.
(2003).
Monoclonal anti-equine IgE antibodies with specificity for different epitopes on the immunoglobulin heavy chain of native IgE.
Vet Immunol Immunopathol, 92(1-2), 45-60.
https://doi.org/10.1016/s0165-2427(03)00007-2 Publication
Researcher Affiliations
- Immunology Unit, Hannover School of Veterinary Medicine, Bischofsholer Damm 15, Germany. bw73@cornell.edu
MeSH Terms
- Animals
- Antibodies, Anti-Idiotypic / immunology
- Antibodies, Monoclonal / immunology
- Antibody Specificity / immunology
- Blotting, Western / veterinary
- Cross Reactions / immunology
- Enzyme-Linked Immunosorbent Assay / veterinary
- Epitopes / immunology
- Female
- Flow Cytometry / veterinary
- Horses / immunology
- Immunoglobulin E / immunology
- Immunoglobulin Heavy Chains / immunology
- Immunoglobulin Isotypes
- Immunomagnetic Separation / veterinary
- Leukocytes / immunology
- Mice
- Mice, Inbred BALB C
- Recombinant Proteins / immunology
Citations
This article has been cited 30 times.- Simonin EM, Torsteinsdóttir S, Svansson V, Björnsdóttir S, Freer H, Tarsillo J, Wagner B. Early allergen introduction overrides allergy predisposition in offspring of horses with Culicoides hypersensitivity. Front Immunol 2025;16:1654693.
- Jonsdottir S, Stefansdottir SB, Mirkovitch J, Ziegler A, Torsteinsdottir S, Marti E. Culicoides allergens expressed in insect cells induce sulphidoleukotriene release from peripheral blood leukocytes of horses affected with insect bite hypersensitivity. Front Immunol 2025;16:1597233.
- Brooks MB, Brooks JC, Catalfamo J, Zhu Y, Goggs R, Babasyan S, Wagner B, LeVine DN. Plasma concentration of thrombopoietin in dogs with immune thrombocytopenia. J Vet Intern Med 2024 Sep-Oct;38(5):2507-2517.
- Jentsch MC, Keilhaue A, Wagner B, Rhyner C, Lübke S, Karagulyan M, Arnold C, Lohmann KL, Schnabel CL. Aspergillus fumigatus binding IgA and IgG1 are increased in bronchoalveolar lavage fluid of horses with neutrophilic asthma. Front Immunol 2024;15:1406794.
- Holmes CM, Babasyan S, Wagner B. Neonatal and maternal upregulation of antileukoproteinase in horses. Front Immunol 2024;15:1395030.
- Schnabel CL, Jentsch MC, Lübke S, Kaiser-Thom S, Gerber V, Vrtala S, Huang HJ, Rhyner C, Wagner B, Hoffmann R, Volke D. Immunoproteomics reveal increased serum IgG3/5 binding to Dermatophagoides and yeast protein antigens in severe equine asthma in a preliminary study. Front Immunol 2023;14:1293684.
- Simonin EM, Wagner B. IgE-binding monocytes upregulate the coagulation cascade in allergic horses. Genes Immun 2023 Jun;24(3):130-138.
- Simonin EM, Babasyan S, Tarsillo J, Wagner B. IgE+ plasmablasts predict the onset of clinical allergy. Front Immunol 2023;14:1104609.
- Gressler AE, Lübke S, Wagner B, Arnold C, Lohmann KL, Schnabel CL. Comprehensive Flow Cytometric Characterization of Bronchoalveolar Lavage Cells Indicates Comparable Phenotypes Between Asthmatic and Healthy Horses But Functional Lymphocyte Differences. Front Immunol 2022;13:896255.
- Mair KH, Crossman AJ, Wagner B, Babasyan S, Noronha L, Boyd P, Zarlenga D, Stadler M, van Dongen KA, Gerner W, Saalmüller A, Lunney JK. The Natural Cytotoxicity Receptor NKp44 (NCR2, CD336) Is Expressed on the Majority of Porcine NK Cells Ex Vivo Without Stimulation. Front Immunol 2022;13:767530.
- Kaul S, Sai Keerthana L, Kumar P, Birader K, Tammineni Y, Rawat D, Suman P. Cytotoxin antibody-based colourimetric sensor for field-level differential detection of elapid among big four snake venom. PLoS Negl Trop Dis 2021 Oct;15(10):e0009841.
- Raza F, Babasyan S, Larson EM, Freer HS, Schnabel CL, Wagner B. Peripheral blood basophils are the main source for early interleukin-4 secretion upon in vitro stimulation with Culicoides allergen in allergic horses. PLoS One 2021;16(5):e0252243.
- Larson EM, Babasyan S, Wagner B. IgE-Binding Monocytes Have an Enhanced Ability to Produce IL-8 (CXCL8) in Animals with Naturally Occurring Allergy. J Immunol 2021 May 15;206(10):2312-2321.
- Raza F, Ivanek R, Freer H, Reiche D, Rose H, Torsteinsdóttir S, Svansson V, Björnsdóttir S, Wagner B. Cul o 2 specific IgG3/5 antibodies predicted Culicoides hypersensitivity in a group imported Icelandic horses. BMC Vet Res 2020 Aug 10;16(1):283.
- Larson EM, Babasyan S, Wagner B. Phenotype and function of IgE-binding monocytes in equine Culicoides hypersensitivity. PLoS One 2020;15(5):e0233537.
- Armstrong C, Cassimeris L, Da Silva Santos C, Micoogullari Y, Wagner B, Babasyan S, Brooks S, Galantino-Homer H. The expression of equine keratins K42 and K124 is restricted to the hoof epidermal lamellae of Equus caballus. PLoS One 2019;14(9):e0219234.
- Einhorn L, Hofstetter G, Brandt S, Hainisch EK, Fukuda I, Kusano K, Scheynius A, Mittermann I, Resch-Marat Y, Vrtala S, Valenta R, Marti E, Rhyner C, Crameri R, Satoh R, Teshima R, Tanaka A, Sato H, Matsuda H, Pali-Schöll I, Jensen-Jarolim E. Molecular allergen profiling in horses by microarray reveals Fag e 2 from buckwheat as a frequent sensitizer. Allergy 2018 Jul;73(7):1436-1446.
- Hellman LT, Akula S, Thorpe M, Fu Z. Tracing the Origins of IgE, Mast Cells, and Allergies by Studies of Wild Animals. Front Immunol 2017;8:1749.
- Wagner B, Perkins G, Babasyan S, Freer H, Keggan A, Goodman LB, Glaser A, Torsteinsdóttir S, Svansson V, Björnsdóttir S. Neonatal Immunization with a Single IL-4/Antigen Dose Induces Increased Antibody Responses after Challenge Infection with Equine Herpesvirus Type 1 (EHV-1) at Weanling Age. PLoS One 2017;12(1):e0169072.
- Ortved K, Wagner B, Calcedo R, Wilson J, Schaefer D, Nixon A. Humoral and cell-mediated immune response, and growth factor synthesis after direct intraarticular injection of rAAV2-IGF-I and rAAV5-IGF-I in the equine middle carpal joint. Hum Gene Ther 2015 Mar;26(3):161-71.
- Khatibzadeh SM, Gold CB, Keggan AE, Perkins GA, Glaser AL, Dubovi EJ, Wagner B. West Nile virus-specific immunoglobulin isotype responses in vaccinated and infected horses. Am J Vet Res 2015 Jan;76(1):92-100.
- Sabban S, Ye H, Helm B. Development of an in vitro model system for studying the interaction of Equus caballus IgE with its high-affinity receptor FcεRI. J Vis Exp 2014 Nov 1;(93):e52222.
- Keggan A, Freer H, Rollins A, Wagner B. Production of seven monoclonal equine immunoglobulins isotyped by multiplex analysis. Vet Immunol Immunopathol 2013 Jun 15;153(3-4):187-93.
- Noronha LE, Harman RM, Wagner B, Antczak DF. Generation and characterization of monoclonal antibodies to equine NKp46. Vet Immunol Immunopathol 2012 Jun 15;147(1-2):60-8.
- Noronha LE, Harman RM, Wagner B, Antczak DF. Generation and characterization of monoclonal antibodies to equine CD16. Vet Immunol Immunopathol 2012 Apr 15;146(2):135-42.
- Goodman LB, Wimer C, Dubovi EJ, Gold C, Wagner B. Immunological correlates of vaccination and infection for equine herpesvirus 1. Clin Vaccine Immunol 2012 Feb;19(2):235-41.
- Morán G, Folch H, Araya O, Burgos R, Barria M. Detection of reaginic antibodies against Faenia rectivirgula from the serum of horses affected with Recurrent Airway Obstruction by an in vitro bioassay. Vet Res Commun 2010 Dec;34(8):719-26.
- Wagner B, Burton A, Ainsworth D. Interferon-gamma, interleukin-4 and interleukin-10 production by T helper cells reveals intact Th1 and regulatory TR1 cell activation and a delay of the Th2 cell response in equine neonates and foals. Vet Res 2010 Jul-Aug;41(4):47.
- Morán G, Burgos R, Araya O, Folch H. In vitro bioassay to detect reaginic antibodies from the serum of horses affected with recurrent airway obstruction. Vet Res Commun 2010 Jan;34(1):91-9.
- Langner KF, Jarvis DL, Nimtz M, Heselhaus JE, McHolland LE, Leibold W, Drolet BS. Identification, expression and characterisation of a major salivary allergen (Cul s 1) of the biting midge Culicoides sonorensis relevant for summer eczema in horses. Int J Parasitol 2009 Jan;39(2):243-50.
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