Tabriz University of Medical Sciences Advanced Pharmaceutical Bulletin 2228-5881 12 3 2022 05 21 Implementation of a Design of Experiments to Improve Periplasmic Yield of Functional ScFv Antibodies in a Phage Display Platform 583 592 10.34172/apb.2022.061 EN Marjan Abri Aghdam https://orcid.org/0000-0001-7803-4954 Mohammad Reza Tohidkia https://orcid.org/0000-0002-2949-7482 Elham Ghamghami Asadollah Ahmadikhah Morteza Khanmahamadi Behzad Baradaran Ahad Mokhtarzadeh https://orcid.org/0000-0002-4515-8675 Journal Article 10.34172/apb.2022.061 2021 03 30 2021 07 02 Purpose: Production of functional recombinant antibody fragments in the periplasm of E. coli isa prerequisite step to achieve sufficient reagent for preclinical studies. Thus, the cost-effectiveand lab-scale production of antibody fragments demands the optimization of culture conditions.Methods: The culture conditions such as temperature, optical density (OD600) at induction,induction time, and IPTG concentration were investigated to optimize the functional expressionof a phage-derived scFv molecule using a design of experiment (DoE). Additionally, the effectsof different culture media and osmolyte supplements on the expression yield of scFv wereexamined.Results: The developed 2FI regression model indicated the significant linear effect of theincubation temperature, the induction time, and the induction OD600 on the expression yieldof functional scFv. Besides, the statistical analysis indicated that two significant interactions ofthe temperature/induction time and the temperature/induction OD600 significantly interplay toincrease the yield. Further optimization showed that the expression level of functional scFvwas the most optimal when the cultivation was undertaken either in the TB medium or in thepresence 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 tocomprehensively optimize the culture conditions for the expression of scFv molecules in aphage antibody display setting, where scFv molecules can be isolated from a tailor-made phageantibody library known as "Human Single Fold scFv Library I."