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PRODUCTION AND CHARACTERIZATION OF MOUSE MONOCLONAL ANTIBODY AGAINST RIBOSOMAL PROTEIN S8

Ribosomi su stanični strojevi za sintezu proteina. Eukariotski ribosom (80S) sastavljen je od male i velike podjedinice. Mala podjedinica (40S) građena je od 18S rRNK te 33 proteina, a velika podjedinica (60S) od 28S, 5,8S i 5S rRNK te 47 proteina. Ribosomski protein S8 (RPS8) sastavni je dio male podjedinice ribosoma. Cilj ovog rada bio je proizvesti mišje monoklonsko protutijelo protiv RPS8. Monoklonska protutijela prepoznaju isti epitop istog antigena. Protutijelo je proizvedeno imunizacijom miševa s rekombinantnim RPS8 sintetiziranim u bakterijama i pročišćenim afinitetnom kromatogafijom. Osim toga, slijedeći cilj bio je karakterizirati proizvedeno monoklonsko protutijela protiv RPS8 te ispitati njegovu pimjenjivost u ELISA analizi, Western blot analizi, metodi imunotaloženja te metodi imunofluorescencije. Dobiveni rezultati pokazuju da poizvedeno monoklonsko protutijelo protiv RPS8 specifično prepoznaje protein RPS8 u ELISA analizi, Western blot analizi te metodama imunotaloženja i imunofluorescencije. Protutijela imaju veliku primjenu u biomedicinskim istraživanjima te je od velike važnosti imati kvalitetna protutijela koja specifično prepoznaju protein od interesa.Ribosomes are large molecular machines that function in protein synthesis. Eukaryotic Ribosome (80S) consists of two subunits: the large and small subunits. The large subunit (60S) contains three rRNAs (28S, 5.8S and 5S) complexed with 46 proteins. The small subunit (40S) contains one rRNA (18S) complexed with 33 proteins.Ribosomal protein S8 (RPS8) is a component of a small (40S) ribosomal subunit. The goal of this work was to produce a mouse monoclonal antibody against RPS8. Monoclonal antibodies recognize the same epitope of the same antigen. The antibody was produced by immunization of mice with an recombinant RPS8 synthesized in bacteria and purified by affinity chromatography. Additionaly, the goal of this work was to characterize the mouse monoclonal antibody against the RPS8 and tested its applicability in ELISA analysis, Western blot analysis, immunoprecipitation and immunofluorescence analysis. The results show that the mouse monoclonal antibody against RPS8 specifically recognizes RPS8 protein in ELISA analysis, Western blot analysis, immunoprecipitation and immunofluorescence methods. Antibodies have many applications in biomedical research and it is important to have high quality antibodies that specifically recognize protein of interest