Development of DNA-Designed Avian IgY Antibodies for Quantitative Determination of Bovine Interferon-Gamma

• Published in: Applied biochemistry and biotechnology 2011-02, Vol.163 (3), p.338-345
• Main Authors: Nikbakht Brujeni, Gholamreza, Jalali, Sayed Amir Hossein, Koohi, Mohammad Kazem
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Antibodies - genetics; Antibodies - immunology; Antigens - immunology; Base Sequence; Bioassays; Biochemistry; Biotechnology; Cattle; Chemistry; Chemistry and Materials Science; Chickens; Chickens - immunology; Cytokines; Deoxyribonucleic acid; DNA; DNA - genetics; Enzyme-Linked Immunosorbent Assay - methods; Enzymes; Human cytomegalovirus; Humans; Immunization; Immunoglobulins; Immunoglobulins - biosynthesis; Immunoglobulins - genetics; Immunoglobulins - immunology; Infectious diseases; Interferon-gamma - analysis; Interferon-gamma - chemistry; Interferon-gamma - genetics; Interferon-gamma - immunology; Lymphocytes; Molecular Sequence Data; Recombinant Proteins; Sequence Alignment; Titrimetry
• ISSN: 0273-2289; 1559-0291
• DOI: 10.1007/s12010-010-9042-9

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 eggyolk 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.