Implementation of a Design of Experiments to Improve Periplasmic Yield of Functional ScFv Antibodies in a Phage Display Platform.

Autor: Aghdam MA; Department of Biological Science, Faculty of Basic Science, Higher Education Institute of Rab-Rashid, Tabriz, Iran., Tohidkia MR; Research Center for Pharmaceutical Nanotechnology, Tabriz University of Medical Sciences, Tabriz, Iran., Ghamghami E; Department of Biological Science, Faculty of Basic Science, Higher Education Institute of Rab-Rashid, Tabriz, Iran., Ahmadikhah A; Faculty of Life Sciences and Biotechnology, Shahid Beheshti University, G.C Velenjak, Tehran, Iran., Khanmahamadi M; Chemical Engineering Faculty, Sahand University of Technology, Sahand New Town, Tabriz, Iran., Baradaran B; Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran., Mokhtarzadeh A; Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
Jazyk: angličtina
Zdroj: Advanced pharmaceutical bulletin [Adv Pharm Bull] 2022 May; Vol. 12 (3), pp. 583-592. Date of Electronic Publication: 2021 Jul 03.
DOI: 10.34172/apb.2022.061
Abstrakt: Purpose: Production of functional recombinant antibody fragments in the periplasm of E. coli is a prerequisite step to achieve sufficient reagent for preclinical studies. Thus, the cost-effective and lab-scale production of antibody fragments demands the optimization of culture conditions. Methods: The culture conditions such as temperature, optical density (OD 600 ) at induction, induction time, and IPTG concentration were investigated to optimize the functional expression of a phage-derived scFv molecule using a design of experiment (DoE). Additionally, the effects of different culture media and osmolyte supplements on the expression yield of scFv were examined. Results: The developed 2FI regression model indicated the significant linear effect of the incubation temperature, the induction time, and the induction OD 600 on the expression yield of functional scFv. Besides, the statistical analysis indicated that two significant interactions of the temperature/induction time and the temperature/induction OD 600 significantly interplay to increase the yield. Further optimization showed that the expression level of functional scFv was the most optimal when the cultivation was undertaken either in the TB medium or in the presence of media supplements of 0.5 M sorbitol or 100 mM glycine betaine. Conclusion: In the present study, for the first time, we successfully implemented DoE to comprehensively optimize the culture conditions for the expression of scFv molecules in a phage antibody display setting, where scFv molecules can be isolated from a tailor-made phage antibody library known as "Human Single Fold scFv Library I."
(©2022 The Authors.)
Databáze: MEDLINE
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