title: Novel polyol-responsive monoclonal antibodies against extracellular β-D-glucans from Pleurotus ostreatus.
abstract: β-D-glucans from mushroom strains play a major role as biological response modifiers in several clinical disorders. Therefore, a specific assay method is of critical importance to find useful and novel sources of β-d-glucans with anti-tumor activity. Hybridoma technology was used to raise monoclonal antibodies (Mabs) against extracellular β-d-glucans (EBG) from Pleurotus ostreatus. Two of these hybridoma clones (3F8_3H7 and 1E6_1E8_B3) secreting Mabs against EBG from P. ostreatus were selected and 3F8_3H7 was used to investigate if they are polyol-responsive Mabs (PR-Mabs) by using ELlSA-elution assay. This hybridoma cell line secreted Mab of IgM class, which was purified in a single step by gel filtration chromatography on Sephacryl S-300HR, which revealed a protein band on native PAGE with Mr of 917 kDa. Specificity studies of Mab 3F8_3H7 revealed that it recognized a common epitope on several β-d-glucans from different basidiomycete strains as determined by indirect ELlSA and Western blotting under native conditions. This Mab exhibited high apparent affinity constant (KApp) for β-d-glucans from several mushroom strains. However, it revealed differential reactivity to some heat-treated β-d-glucans compared with the native forms suggesting that it binds to a conformation-sensitive epitope on β-d-glucan molecule. Epitope analysis of Mab 3F8_3H7 and 1E6_1E8_B3 was investigated by additivity index parameter, which revealed that they bound to the same epitope on some β-d-glucans and to different epitopes in other antigens. Therefore, these Mab can be used to assay for β-d-glucans as well as to act as powerful probes to detect conformational changes in these biopolymers.
authors.affiliation: Chemical Engineering and Biotechnology Research Center and Departmental Area of Chemical Engineering of Instituto Superior De Engenharia De Lisboa, R. Conselheiro Emídio Navarro 1, 1959-007, Lisboa, Portugal.|Centre for the Research and Technology Agro-Environment and Biological Sciences, Universidade De Trás-os-Montes E Alto Douro, Quinta De Prados, Apartado 1013, 5001-801 Vila Real, Portugal.
authors.forename: Magda  C
authors.initials: M
authors.lastname: Semedo
authors.affiliation: Chemical Engineering and Biotechnology Research Center and Departmental Area of Chemical Engineering of Instituto Superior De Engenharia De Lisboa, R. Conselheiro Emídio Navarro 1, 1959-007, Lisboa, Portugal.|Centre for the Research and Technology Agro-Environment and Biological Sciences, Universidade De Trás-os-Montes E Alto Douro, Quinta De Prados, Apartado 1013, 5001-801 Vila Real, Portugal.
authors.forename: Amin
authors.initials: A
authors.lastname: Karmali
authors.affiliation: Chemical Engineering and Biotechnology Research Center and Departmental Area of Chemical Engineering of Instituto Superior De Engenharia De Lisboa, R. Conselheiro Emídio Navarro 1, 1959-007, Lisboa, Portugal.|Centre for the Research and Technology Agro-Environment and Biological Sciences, Universidade De Trás-os-Montes E Alto Douro, Quinta De Prados, Apartado 1013, 5001-801 Vila Real, Portugal.
authors.forename: Sónia
authors.initials: S
authors.lastname: Martins
authors.affiliation: Dept. of Bioengineering, Centre for Biological and Chemical Engineering of Instituto Superior Técnico, Av. Rovisco Pais, 1, 1049-001, Lisboa, Portugal.
authors.forename: Luís
authors.initials: L
authors.lastname: Fonseca
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000627:therapeutic use; D000939:Epitopes / Q000276:immunology; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007075:Immunoglobulin M / Q000276:immunology; D009369:Neoplasms / Q000188:drug therapy*; D020076:Pleurotus / Q000737:chemistry*; D011108:Polymers / Q000737:chemistry; D047071:beta-Glucans / Q000276:immunology* / Q000627:therapeutic use
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D007075:Immunoglobulin M; D011108:Polymers; D047071:beta-Glucans; C024617:polyol
keywords: IgM Mab class; Pleurotus ostreatus; epitope analysis of Mab; extracellular β-d-glucans; hybridoma technology