E. coli Fermentation Process Monitoring
E. coli fermentation for the production of recombinant proteins and other products is a complex, multistep process that requires rigorous control of culture conditions. For optimal fermentation to take place in the bioreactor, oxygen level and pH must be maintained within a narrow range. The bacterial cells consume oxygen as they replicate, so the culture must be sparged with oxygen as necessary.
Also, the bacterial cells release metabolic by-products during the growth process, requiring periodic rebalancing of pH levels. Maintaining optimal conditions to ensure high yields requires frequent monitoring of oxygen and pH in real time during the reaction.
Various technologies exist for oxygen and pH measurements, including galvanic sensors, paramagnetic sensors, fuel-cell sensors, and even paper. Yet all these techniques are invasive and require sampling the culture or placing a probe or sensor into the reactor vessel, which can introduce contamination or alter growth conditions.
Optical sensors provide noncontact alternatives to more invasive monitoring methods. Fluorescence can be used to monitor oxygen in the liquid and headspace phases of the bioreactor. Partial pressure of dissolved or gaseous oxygen is monitored using a phase fluorometer to measure the phase shift between the excitation signal of a blue LED and the emission signal of the fluorescence.
Noninvasive monitoring of pH in the liquid phase can be accomplished by measuring the colorimetric response of a pH dye using a miniature spectrometer. This method is ratiometric and relatively unaffected by drift. Colorimetric pH sensors also can be implemented as reflective sensors immune to the effects of solution color, sediment, or turbidity.
Typically, reflective pH and oxygen sensors are patches consisting of an active sensor layer formed by doping a thin sheet of sol-gel host matrix with target-sensitive fluorophores for oxygen sensing or indicator dyes for pH sensing. In some pH sensors a gold mesh can improve reflectivity and be customized for transmission levels, pore sizes, thicknesses, and other parameters.
The pH sensor layer is sandwiched between this mesh layer and a layer of adhesive that allows the patch to be affixed to the vessel wall. Oxygen sensor patches can be left bare or coated with a silicone overcoat to make them more robust for harsh environments.