Tehran University of Medical Sciences Iranian Journal of Allergy, Asthma and Immunology 1735-1502 14 5 2015 10 20 502 508 EN Mahdi Fasihi-Ramandi Applied Biotechnology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran. Jafar Amani Applied Microbiology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran. AliHatef Salmanian Department of Agricultural Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran. SeyedMohammad Moazzeni Immunology Department, Tarbiat Modares University, Tehran, Iran. Kazem Ahmadi Molecular Biology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran. 2015 10 19 2015 10 19 The B-cell CD20 antigen is one of the most reliable surface targets in immunotherapy of B lymphoma. In this project, we studied the production and characterization of a new monoclonal antibody against chimeric human CD20 extra loops (hCD20 exl). The results showed that clone C12H, IgG2/k isotype reacted with the antigen in ELISA and immunoblot. The Kd value was  found to be 2×10-9M and flow cytometry results showed that 99.9% and 99.7% of the Daudi and Raji cells respectively were stained with C12H monoclonal antibody (mab) but not with Jurkat cell lines. It also effectively competed with Rituximab, thus, the staining of the Daudi and Raji cell lines was reduced to 55.9% and 40.5% of cells respectively. Based on the high affinity reaction of C12H mab and appropriate reactivity of C12H mab with the native antigen on the surface of Raji and Daudi cells in flow cytometry, it was concluded that development and evaluation of C12H mab could be a beneficial candidate for further application in genetically engineered monoclonal antibody. https://ijaai.tums.ac.ir/index.php/ijaai/article/view/600 https://ijaai.tums.ac.ir/index.php/ijaai/article/download/600/565