en ايمونولوژي Immunology پژوهشي Research <p dir="ltr"><strong>Background</strong><strong>:</strong> The ability of polyclonal antibodies to react with many epitopes of an antigen makes them valuable reagents in research and diagnosis. The aim of this study was purification of mouse IgG2a and production of polyclonal antibody against purified mouse IgG2a subclass.</p> <p dir="ltr"><strong>Materials and Methods: </strong>Mouse IgG2a was purified by ProA affinity. Verification method of the purified antibody was SDS-PAGE and ELISA by a mouse isotyping Kit. Rabbit was immunized with purified IgG2a. The production of antibody in rabbit was investigated by direct ELISA method. Rabbit serum was collected and precipitated at the final concentration of 50% ammonium sulfate. Polyclonal antibody was purified by ion-exchange chromatography and labeled with HRP. The titre and cross reactivity of product was detected by direct ELISA method.</p> <p dir="ltr"><strong>Results:</strong> The results of SDS-PAGE in reduced and non-reduced conditions showed bands with 50-KDa, 25-30 KDa MW and a distinct band with 150 KDa MW. Isotype determination showed the presence of mouse IgG2a in related fraction. The titer of Anti-mouse polyclonal antibody was 200000. The optimum titer of prepared HRP conjugated IgG was 4000. Conjugated rabbit IgG has more cross reactivity with mouse IgG2b.</p> <p dir="ltr"><strong>Conclusion:</strong> Taking together, affinity chromatography and ion-exchange chromatography are appropriate techniques for purification of mouse IgG subclasses and rabbit IgG, respectively.</p> <p dir="ltr">.</p> Affinity chromatography, purification, polyclonal antibody, IgG2a, ion-exchange chromatography, 30 35 http://rmm.mazums.ac.ir/browse.php?a_code=A-10-745-1&slc_lang=en&sid=1 Zahra Moradi nebrin 10031947532846007088 10031947532846007088 No Immunology Research Center, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran Jafar Majidi Majidij@Tbzmed.ac.ir 10031947532846007089 10031947532846007089 Yes Tabriz Autonomous University of Medical Sciences, Tabriz University of Medical Sciences, Tabriz, Iran Leili Aghebati Maleki 10031947532846007090 10031947532846007090 No Immunology Research Center, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran Tohid Kazemi 10031947532846007091 10031947532846007091 No Department of Immunology, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran Jalal Abdolalizadeh 10031947532846007092 10031947532846007092 No Department of Immunology, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran Somaieh Dadashi 10031947532846007093 10031947532846007093 No Immunology Research Center, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran Sadeg Eyvazi 10031947532846007094 10031947532846007094 No Immunology Research Center, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran Majid Ahmadi 10031947532846007095 10031947532846007095 No Immunology Research Center, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran Naiemeh Majidi zolbanin 10031947532846007096 10031947532846007096 No Pharmacology Department, Faculty of pharmacy, Tabriz University of Medical Sciences, Tabriz, Iran