Author/Authors :
Habibi Anbouhi, Mahdi pasteur institute of iran - national cell bank of iran, ايران , Feiz Barazandeh, Aida pasteur institute of iran - national cell bank of iran, ايران , Bouzari, Saeid pasteur institute of iran - molecular biology unit, ايران , Abolhassani, Mohsen pasteur institute of iran - Department of Immunology, Hybridoma Laboratory, ايران , Khanahmad, Hossein pasteur institute of iran - production and research complex, ايران , Golkar, Majid pasteur institute of iran - Department of Parasitology, molecular parasitology laboratory, ايران , Aghasadeghi, Mohammad Reza Pasteur Institute of Iran - Department of Hepatitis and AIDS, ايران , Behdani, Mahdi pasteur institute of iran - national cell bank of iran, ايران , Jahanian-Najafabadi, Ali pasteur institute of iran - molecular biology unit, ايران , Shokrgozar, Mohammad Ali pasteur institute of iran - national cell bank of iran, ايران
Abstract :
Background: Targeting of CD20 antigen with monoclonal antibodies has become the mainstay in the treatment of non-Hodgkin s lymphomas and immunotherapeutic depletion of malignant B cells. Accessibility of antigen is one of the crucial factors in development of monoclonal antibodies against this antigen. One major problem in expression of full length CD20 is aggregation and misfolding. Therefore, production of an alternative polypeptide is easer and favorable comparing to that of a full length transmembrane protein CD20. Methods: In this study, we expressed the extra membrane loop of hCD20 (exCD20) consisting of a non-glycosylated 47-amino acids region. The exCD20 coding sequence was amplified by PCR and cloned in pET32a(+) expression vector. The desired protein was expressed in fusion with thioredoxin and 6× His tag in E. coli Origami strain. ELISA and Westernblotting data were performed to indicate the functionality of this protein. Results: We have obtained the exCD20 recombinant protein which can be detected in ELISA and Western-blot experiments. This recombinant fusion protein was soluble and stable without aggregation and misfolding problems. Conclusion: The recombinant extra membrane loop of human CD20 protein in fusion with thioredoxin (exCD20) can be used in function assays and some applications such as ELISA, immuneblotting, affinity purification, immunization, screening, and development of anti-CD20 antibodies.
