WHY USE MASS SPECTROMETRY FOR FERMENTATION GAS ANALYSIS?

We are living in exciting and challenging times with the increasing use of mammalian cell cultures to produce biopharmaceuticals - ‘exciting’ because they offer the prospect of radical developments in vaccines, monoclonal antibodies and gene therapy, ‘challenging’ because they are more delicate and require more complex processing than simpler cells.

Our gas analysis mass spectrometers have been used with great success for over 30 years to monitor microbial and bacterial processes, at the IBioIC conference we’ll also be showing how they are now being used to improve understanding and increase yields of mammalian cell cultures.

Historically the main application in microbial and bacterial processes has been respiratory gas analysis to generate values for Oxygen Uptake Rate (OUR), Carbon dioxide Evolution Rate (CER) and Respiratory Quotient (RQ). Our mass spectrometers can measure the four air gases (not just oxygen and carbon dioxide, but also nitrogen and argon to provide a complete compositional analysis) in just 10 seconds, including the time to switch from one sample stream to the next. And fast analysis is not at the expense of precision – they typically measure oxygen with a precision of ≤0.005 %mol and carbon dioxide with a precision of ≤0.0003 %mol.  They are also used to detect contamination by looking for small changes in oxygen and carbon dioxide before culture addition, and monitor volatiles at trace levels, such as methanol and ethanol and hydrogen sulphide. Mass spectrometers have been used in both development laboratories and manufacturing plants.

Considering the increasing importance of mammalian cell culture we have recently collaborated with the Department of Biochemical Engineering, University College London (UCL), to look at the usefulness of gas analysis MS for this growing area of biotechnology. We’ll describe the challenges that had to be overcome - for example, in microbial and bacterial fermentations, the feed gas composition is relatively constant, either air or air enriched with oxygen. But in mammalian cell fermentations, the feed gas composition is a frequently changing mixture of several compounds (e.g. nitrogen, oxygen and carbon dioxide). Feed gas concentration ranges can therefore vary dramatically – for example carbon dioxide can vary from tens of part per million to tens of percent – and so the mass spectrometer must be able to see small changes in respiratory gas compositions against this rapidly changing feed gas ‘baseline’.

The work at UCL was carried out with a Thermo Scientific™ Prima BT bench-top magnetic sector MS monitoring two 5 litre bioreactors in which modified Chinese Hamster Ovary (CHO) cells were used to produce monoclonal antibodies. The MS analyzed inlet and outlet gases for oxygen, carbon dioxide, nitrogen and argon on both bioreactors. Besides the on-line MS measurements, cell count (both total and viable), lactate levels and glucose levels were analyzed off-line. The sparge gas to the two bioreactors was a combination of oxygen, carbon dioxide, nitrogen and Air.

We’ll show that the high precision provided by Prima BT’s magnetic sector analyzer was able to overcome the challenges associated with the mammalian cell culture process, for fermentations that ran for weeks rather than days. The online MS gas analysis data provide an invaluable complement to the offline cell count and metabolite data, leading to better understanding of the process dynamics and increase yield of the monoclonal antibodies.

 

Graham Lewis, Thermo Fisher Scientific