Optimizing E. Coli cell growth performance with in-line, real-time OD600 (abs/mm) monitoring during the fermentation process

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The increasing demand of cell and gene therapeutics (CGT) have created a need to optimize manufacturing process steps using Process Analytical Technologies (PAT) to lower costs and increase yields. Plasmid DNA (pDNA) is a precursor to the development of CGTs and is met with many challenges throughout the production process due its size, viscosity, and similarities to other impurities which can impact their yields. The generation of these plasmids in high density E-Coli cultures can be difficult to monitor using traditional off-line OD600 methods due to the careful sample preparation, fast growth rate, and dilutions required to be in the linear range of a standard spectrophotometer. 

This study proposes utilizing an inline- variable pathlength spectrophotometer in a recirculation loop during fermentation to monitor and optimize the growth of E. Coli cells in real-time. This provides distinct advantages vs traditional methods due to the constant modification of path lengths to create a slope-based solution that is always within the linear range of the equipment with no buffer correction required. The growth curve is tracked in real-time using the slope (Absorbance/mm) at 600nm to measure cell density and to optimize growth performance of E. Coli cells. This will serve as a proof-of-concept for the optimal cultivation of E. Coli and production of pDNA.

• What is Variable Pathlength Technology (VPT) and how it differs from traditional off-line UV-Vis methods and equipment.
• FlowVPX System data review of in-line OD600 monitoring during the fermentation process.
• Eliminate careful sample preparation and dilutions required for offline OD600 measurements.
• Increase pDNA production via inline analytics and optimal timepoint selection – Proof of concept.

Tanja Buch

Technical Director at Biofactory Competence Center

Tanja Buch is a Biosystem Engineer with a PhD in Bioprocess Engineering. She has over 10 years of experience in the cultivation of mammalian cells and microorganism as well as the purification of recombinant proteins using filtration and chromatographic tools. Tanja has worked in several cooperation’s with large biopharmaceutical companies to support them with process development, process optimization or analytical method development for their processes. In 2015 she took part in the design and set-up of the Biofactory Competence Center (BCC) in Fribourg, Switzerland, a training and research center for the biopharmaceutical industry. Within the BCC, Tanja is responsible to deliver training courses to the biopharma companies and to design and perform research projects for the industrial partners in all areas of bioprocess manufacturing.

Ramsey Shanbaky

Director, In-Line Applications, Bioprocess Analytics at

Ramsey Shanbaky spent the first 8 years of his career working in R&D, Manufacturing and Product Management designing and building fiber optic products for high powered laser delivery in the medical device industry. In the first part of his tenure at Repligen, he worked alongside his customers to implement the CTech™ SoloVPE® System within biopharmaceutical accounts. For the last 5 years, he has been partnering with customers to implement the CTech™ FlowVPE® and FlowVPX™ Systems for in-line concentration measurements both in the process development and GMP manufacturing areas for bioprocessing. He holds an BSEE from Purdue.

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SPEAKERS

Tanja Buch

Technical Director at Biofactory Competence Center

Ramsey Shanbaky

Director, In-Line Applications, Bioprocess Analytics at

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