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Production of a methylenediphenyl diisocyanate hapten-specific monoclonal antibody.
Green-B; Nayak-A; Law-B; Siegel-P; Beezhold-DH
J Allergy Clin Immunol 2013 Feb; 131(2)(Suppl):AB15
RATIONALE: 4,4'-Methylenediphenyl diisocyanate (MDI) is used in the manufacturing of rigid polyurethane foams, coatings, adhesives, sealants and elastomers. MDI is a low molecular weight allergen and occupational exposure is associated with allergic sensitization, dermatitis, and asthma. Here we describe the production and initial characterization of an MDIhapten specific monoclonal antibody (mAb). METHODS: Female BALB/c mice were immunized intraperitoneally with 25 mg of MDI- keyhole limpet hemocyanin in TiterMax Gold (vol 1:1). Following 6 biweekly immunizations, splenocytes were recovered 72 h after the final immunization and fused to Sp2/0-Ag14 myeloma cell line using 50% polyethylene glycol (1500 MW) for hybridoma production. Hybridomas were then screened in a solid-phase indirect enzyme-linked immunosorbent assay (ELISA) against 40:1 4,4'-MDI-human serum albumin (HSA) and 4,4'-MDI-keratin (KER) protein conjugates. Isotype and mAb reactivity to various protein-isocyanate conjugate epitopes were characterized using an indirect ELISA. Quantification of 2 fold serial dilutions of 4,4'-MDI-HSAwas determined using a colorimetric sandwich ELISA. RESULTS: One hybridoma producing a multimeric IgM mAb(15D4) was produced that reacted with 4,4'-MDI-HSA and 4,4'-MDI-KER. ELISA analysis demonstrated comparable reactivity with hexamethylene diisocyanate- HSA. mAb 15D4 reacted more strongly with 4,4'-MDI-HSA than 2,4' MDI-HSA. Preliminary results demonstrated that concentrations ranging from 3.1-100 mg/mL of 4,4'-MDI-HSA could be quantified using a colorimetric sandwich ELISA. CONCLUSIONS: The preliminary studies of IgM mAb 15D4 suggest potential utility for the development of standardized immunoassays for assessing worker exposure to MDI. Furthermore, this mAb may also be useful for the isolation of endogenous MDI-haptenated proteins following occupational or experimental exposures.
Polyurethane-foams; Coatings; Adhesives; Sealing-compounds; Elastic-properties; Molecular-structure; Allergens; Exposure-levels; Dermatitis; Allergic-reactions; Allergies; Respiration; Pulmonary-system; Pulmonary-system-disorders; Pulmonary-function; Animals; Laboratory-animals
Issue of Publication
Healthcare and Social Assistance; Services
Journal of Allergy and Clinical Immunology
Page last reviewed: September 2, 2020
Content source: National Institute for Occupational Safety and Health Education and Information Division