The aim of this study was to investigate the effect of r-insulin in ExpiSf9 cell cultures growing in ExpiSF CD medium. An initial phase of adaptation to different r-insulin concentration was performed prior to experimentation. The first part focused on the evaluation of r-insulin as a supplement for cell growth and viability maintenance. In the second part, r-insulin was investigated as an enhancer for HIV-1 Gag VLP production through baculovirus infection (BV). The Gag gene was fused in frame to eGFP to ease process characterization and product quantification.