Development of DNA-designed avian IgY antibodies for quantitative determination of bovine interferon-gamma.
Interferon-gamma (IFN-γ), a cytokine produced by sensitized T lymphocytes, is one of the key elements in defining T helper 1 lymphocyte immune responses. Quantitative evaluation of IFN-γ expression could provide an important analytical tool for measurement of cell-mediated immunity and investigating immune responses to infectious diseases. Method of DNA-designed avian IgY antibodies was used for production of monospecific polyclonal antibodies that allows quantification of the recombinant bovine IFN-γ protein. IFN-γ cDNA was subcloned and expressed in mammalian expression plasmid (pcDNA3.1(+)) under the control of the human cytomegalovirus promoter. Chickens were immunized by plasmid DNA, and egg yolk antibodies extracted from eggs were collected after immunization. IgY-specific antibodies were evaluated by an antigen capture enzyme-linked immunosorbent assay (ELISA) using recombinant IFN-γ. Based on the results, developed bovine IFN-γ capture ELISA could detect up to 1 ng/ml of IFN-γ by 64-fold diluted IgY. Monospecific anti-bovine IFN-γ antibodies generated in chickens are useful for quantifying different concentrations of recombinant bovine IFN-γ, which is expressed in cell culture.
Nikbakht Brujeni
Gholamreza
G
Department of Microbiology and Immunology, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran, nikbakht@ut.ac.ir.
Jalali
Sayed Amir Hossein
SA
Koohi
Mohammad Kazem
MK
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000235:genetics / Q000276:immunology*; D000941:Antigens / Q000276:immunology; D001483:Base Sequence; D002417:Cattle; D002645:Chickens / Q000276:immunology*; D004247:DNA / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007136:Immunoglobulins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology*; D007371:Interferon-gamma / Q000032:analysis* / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D008969:Molecular Sequence Data; D011994:Recombinant Proteins; D016415:Sequence Alignment; D017186:Titrimetry
D000906:Antibodies; D000941:Antigens; C016512:IgY; D007136:Immunoglobulins; D011994:Recombinant Proteins; D007371:Interferon-gamma; D004247:DNA