Alizadeh, A., Hamzeh-Mivehroud, M., Haddad, E., Haddad, N., Sharifi, M., Mohammadi, S., Pourtaghi, S., Dastmalchi, S. (2019). Characterization of Novel Fragment Antibodies Against TNF-alpha Isolated Using Phage Display Technique. Iranian Journal of Pharmaceutical Research, 18(2), 759-771. doi: 10.22037/ijpr.2019.1100646
Ali Akbar Alizadeh; Maryam Hamzeh-Mivehroud; Elnaz Haddad; Nazanin Haddad; Mehdi Sharifi; Samin Mohammadi; Samira Pourtaghi; Siavoush Dastmalchi. "Characterization of Novel Fragment Antibodies Against TNF-alpha Isolated Using Phage Display Technique". Iranian Journal of Pharmaceutical Research, 18, 2, 2019, 759-771. doi: 10.22037/ijpr.2019.1100646
Alizadeh, A., Hamzeh-Mivehroud, M., Haddad, E., Haddad, N., Sharifi, M., Mohammadi, S., Pourtaghi, S., Dastmalchi, S. (2019). 'Characterization of Novel Fragment Antibodies Against TNF-alpha Isolated Using Phage Display Technique', Iranian Journal of Pharmaceutical Research, 18(2), pp. 759-771. doi: 10.22037/ijpr.2019.1100646
Alizadeh, A., Hamzeh-Mivehroud, M., Haddad, E., Haddad, N., Sharifi, M., Mohammadi, S., Pourtaghi, S., Dastmalchi, S. Characterization of Novel Fragment Antibodies Against TNF-alpha Isolated Using Phage Display Technique. Iranian Journal of Pharmaceutical Research, 2019; 18(2): 759-771. doi: 10.22037/ijpr.2019.1100646
Characterization of Novel Fragment Antibodies Against TNF-alpha Isolated Using Phage Display Technique
1Biotechnology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
2School of Pharmacy, Tabriz University of Medical Sciences, Tabriz, Iran.
3Student Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran.
4Faculty of Pharmacy, Near East University, POBOX: 99138, Nicosia, North Cyprus, Mersin 10, Turkey.
Abstract
Tumor necrosis factor alpha (TNF-α) is an inflammatory cytokine which plays crucial roles in pathogenesis of inflammatory diseases. The current study aimed to investigate the binding abilities of I44 and I49 domain antibodies to TNF-α. The dAbs were expressed in bacterial expression system and purified by affinity chromatography using Ni-sepharose column. The expression and purity of the proteins were evaluated using western blotting and SDS-PAGE techniques, respectively. ELISA experiment showed that I44 and I49 dAbs bind to TNF-α with the binding constants (Kd) of 5.18 ± 1.41 and 2.42 ± 0.55 µM, respectively. The inhibitory effect of dAbs on TNF-α biological effect was determined in MTT assay in which I44 and I49 prevented TNF-α cell cytotoxicity with IC50 values of 6.61 and 3.64 µM, respectively. The identified anti-TNF-α dAbs could bind to and inhibit TNF-α activity. The dAbs activities can be attributed to their ability to establish hydrogen bonds as well as hydrophobic contacts with TNF-α. The results of the current study can pave the way for further structural studies in order to introduce new more potent anti-TNF-α antibodies.
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