title: Development and evaluation of a single domain antibody against human epidermal growth factor receptor (EGFR).
abstract: Epidermal growth factor receptor (EGFR) plays an important role in cell growth, multiplication and differentiation. Over expression of EGFR is associated with carcinogenesis and seen in variety of cancers. Anti-EGFR monoclonal antibodies can block EGFR downstream signaling pathway resulting in inhibition of uncontrolled cell proliferation. Antibody fragments have a variety of advantages. In comparison to full length antibodies they have smaller size and therefor exhibit better tumor penetration ability. The aim of this study was to prepare a single domain antibody to target extracellular domain of EGFR. mRNA was extracted from C225 hybridoma cells producing anti-EGFR antibody and subjected to reverse transcription reaction (RT-PCR) to obtain cDNA molecules encoding VH domain of mAb C225. The cDNA encoded VH domain was in frame introduced into the pET-22b(+) vector and expressed in BL21 (DE3) bacterial cells. The resultant antibody was purified via Ni- NTA column and its reactivity was assessed by ELISA and western blot techniques using A431 cell lysate. Analysis by ELISA revealed that this single domain antibody was able to bind EGFR on A431cells. This result was further confirmed by western blotting. In conclusion, the results of this study indicated that single domain antibody can identify and bind to EGFR of A431 carcinoma cells. This recombinant fragment antibody would potentially be used for targeting of cancer cells with high EGFR expression.
authors.affiliation: Biotechnology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
authors.forename: Shiva
authors.initials: S
authors.lastname: Ahdi Khosroshahi
authors.affiliation: Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran. Electronic address: farajnias@tbzmed.ac.ir.
authors.forename: Safar
authors.initials: S
authors.lastname: Farajnia
authors.affiliation: Department of Microbiology, Faculty of Basic Sciences, Islamic Azad University, Ahar Branch, Ahar, Iran.
authors.forename: Mehdi
authors.initials: M
authors.lastname: Ghiamirad
authors.affiliation: Maragheh Faculty of Medical Sciences, Maragheh, Iran.
authors.forename: Asghar
authors.initials: A
authors.lastname: Tanomand
authors.affiliation: Immunology Research, Tabriz University of Medical Sciences, Tabriz, Iran.
authors.forename: Kamal
authors.initials: K
authors.lastname: Veisi
authors.affiliation: Tuberculosis and Lung Disease Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
authors.forename: Leila
authors.initials: L
authors.lastname: Rahbarnia
authors.affiliation: Biotechnology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
authors.forename: Bahman
authors.initials: B
authors.lastname: Akbari
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D015153:Blotting, Western; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D066246:ErbB Receptors / Q000276:immunology*; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction; D061905:Single-Domain Antibodies / Q000096:biosynthesis / Q000276:immunology* / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D061905:Single-Domain Antibodies; C512478:EGFR protein, human; D066246:ErbB Receptors
keywords: EGFR; Recombinant expression; Single domain antibody