(93g) Characterization of Cell Growth and Recombinant Protein Production in Shaker Batch Cultures for Implementation of Nir Spectroscopy as a Monitoring Technique

Although rarely available, timely process information about fermentation biomass, substrates, intermediates, and products is required to make control decisions. NIR spectroscopy offers the opportunity to improve the fermentation processes by incorporating rapid, nondestructive, and multi-constituent analysis of fermentation broth media directly into the fermentation monitoring and control strategies. The objective of our research project is to use NIR technology to monitor the cell growth and recombinant protein production of a genetically-modified Escherichia coli strain cultures in batch mode. To achieve this objective, preliminary experiments are needed to characterize the optimal growth of the bacteria in their culture media (LB broth + ampicilline) with and without protein induction (L-arabinose addition to media) in order to build models to monitor the biomass and recombinant protein production with the NIR spectral data. The E. coli strain and the recombinant protein selected is commercially available (Bio-Rad pGLO kit), which produces a green fluorescent protein (GFP) when induced with L-arabinose. We will report on the results from a set of experiments that include: monitoring of shaker flask batch cultures of E. coli producing recombinant GFP using UV spectrometry to monitor cell mass and fluorescence spectrophotometry to monitor GFP production and as the primary reference methods to eventually calibrate and build the models with the NIR data. The characterization consists in estimating the Monod parameters of the E. coli using batch culture data for induced and non-induced cultures. Once the characterization is completed, further experiments will be conducted to calibrate the NIR equipment and build the models needed to fully validate the NIR as a tool for monitoring cell growth and protein production. This method would result to be very important to biotechnology industries and the implementation of Process Analytical Technologies (PAT), which has been promoted by the FDA as a preferred mode to produce regulated biotechnology products in the future.