Rat monoclonal antibodies against new guanine nucleotide exchange factor, mouse Zizimin2.
We recently isolated and identified Zizimin2 as a functional factor that is highly expressed in murine splenic germinal center B cell after immunization with T-cell-dependent antigen. Zizimin2 was revealed that it is a new family member of Dock (dedicator of cytokinesis), Dock11, which was the guanine nucleotide exchange factor for Cdc42, the Rho family of low molecular weight GTPases. However, neither the molecular function of Zizimin2 in the acquired immunity nor the physiological role in decrease of immunological activity along with aging has been known. To facilitate the further analysis of physiological function of this molecule, we have established rat monoclonal antibodies (MAbs) against mouse Zizimin2 protein. Recombinant His-tagged partial mouse Zizimin2 protein was expressed in Escherichia coli, purified, and applied as an antigen for immunization. The desired hybridomas were selected by the recombinant Zizimin2 protein coated enzyme-linked immunosorbent assay screening, and we generated stable hybridoma cell lines that produced the antibody against murine Zizimin2 protein. We determined their isotypes and further examined capability or availability in immunoblotting, immunoprecipitation, or immunofluorescence microscopy. Here we demonstrated the several appropriate antibodies for immunoblotting and immunofluorescence microscopy. These MAbs should therefore be very useful tools for study of the characterization of the Zizimin2 protein and the understanding of the biological function of Zizimin2-mediated immunosenescence.
Jia
Yanjun
Y
Department of Mechanism of Aging Research Institute, National Center for Geriatrics and Gerontology, Obu-city, Aichi, Japan.
Asai
Azusa
A
Sakabe
Isamu
I
Maruyama
Mitsuo
M
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification; D000918:Antibody Specificity; D015153:Blotting, Western; D002460:Cell Line; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005455:Fluorescent Antibody Technique; D020662:Guanine Nucleotide Exchange Factors / Q000276:immunology*; D006801:Humans; D006825:Hybridomas; D047468:Immunoprecipitation; D051379:Mice; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000276:immunology / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; C498343:Dock11 protein, mouse; D020662:Guanine Nucleotide Exchange Factors; D011994:Recombinant Proteins