# Optimization of culture condition for Spodoptera frugiperda by design of experiment approach and evaluation of its effect on the expression of hemagglutinin protein of influenza virus

## TL;DR

This study optimizes culture conditions for insect cells to improve the production of a flu protein using experimental design methods.

## Contribution

A novel application of Placket-Burman and Box-Behnken designs to optimize Sf9 cell culture and HA protein expression in BEVS.

## Key findings

- Feed percentage, cell count, and multiplicity of infection were identified as key factors affecting r-HA expression.
- Optimized conditions improved r-HA expression level and potency compared to standard protocols.
- Experimental design effectively selected and optimized parameters for Sf9 cell culture and protein production.

## Abstract

The baculovirus expression vector system (BEVS) is a powerful tool in pharmaceutical biotechnology to infect insect cells and produce the recombinant proteins of interest. It has been well documented that optimizing the culture condition and its supplementation through designed experiments is critical for maximum protein production. In this study, besides physicochemical parameters including incubation temperature, cell count of infection, multiplicity of infection, and feeding percentage, potential supplementary factors such as cholesterol, polyamine, galactose, pluronic-F68, glucose, L-glutamine, and ZnSO4 were screened for Spodoptera frugiperda (Sf9) cell culture and expression of hemagglutinin (HA) protein of Influenza virus via Placket-Burman design and then optimized through Box-Behnken approach. The optimized conditions were then applied for scale-up culture and the expressed r-HA protein was characterized. Optimization of selected parameters via the Box-Behnken approach indicated that feed percentage, cell count, and multiplicity of infection are the main parameters affecting r-HA expression level and potency compared to the previously established culture condition. This study demonstrated the effectiveness of designing experiments to select and optimize important parameters that potentially affect Sf9 cell culture, r-HA expression, and its potency in the BEVS system.