No abstract

The monitoring and control of bioprocesses is of the utmost importance in order to provide a consistent, safe, and high-quality product for consumers. Current monitoring and control schemes rely on infrequent and time consuming offline sampling methods, which inherently leads to some variability in the process which may impact the product quality profile. As part of Lonza's dedication to process analytical technology (PAT) initiatives this study evaluated the ability to generate generic calibration models, which are independent of the cell line, using Raman probes to monitor changes in glucose, lactate, glutamate, ammonium, viable cell concentration (VCC), total cell concentration (TCC) and product concentration. Calibration models were developed from cell culture using two different CHOK1SV GS-KO cell lines producing different monoclonal antibodies (mAbs). Developed predictive models, measured changes in glucose, lactate, ammonium, VCC, and TCC with average prediction errors of 0.44, 0.23, 0.03 g L , 1.90 × 10 cells mL , and 1.85 × 10 cells mL , respectively over the course of cell culture with minimal cell line dependence. The development of these generic models allows the application of spectroscopic PAT techniques in clinical and commercial manufacturing environments, where processes are typically run once or twice in GMP manufacturing based on a common platform process. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:730-737, 2018.

Miniaturized and integrated components for electrochemical detection in micro- and nano-fluidic devices are of great interest as they directly yield an electrical signal and promise sensitive, label-free, real-time detection. One of the challenges facing electrochemical sensing is the lack of reliable reference electrode options. This paper describes the fabrication and characterization of a microscale palladium hydride reference electrode in a single microfabrication step. The reference electrode was integrated inside of a nanoscale constriction along with a gold working electrode to create a complete electrochemical sensor. After charging the palladium electrode with hydrogen, the device was used to detect pyocyanin concentrations from 1-100 μM, with a 0.597 micromolar detection limit. This is the first time that a palladium hydride reference electrode has been integrated with a microfabricated electrochemical sensor in a nanofluidic setup. The device was then used over the course of 8 days to measure pyocyanin produced by four different Pseudomonas aeruginosa strains in growth media. By utilizing square wave and differential pulse voltammetry, the redox active molecule, pyocyanin, was selectively detected in a complex solution without the use of any electrode surface modification.

The condition of cells in Pseudomonas aeruginosa biofilms was monitored via the electrochemical detection of the electro-active virulence factor pyocyanin in a fabricated microfluidic growth chamber coupled with a disposable three electrode cell. Cells were exposed to 4, 16, and 100 mg L(-1) colistin sulfate after overnight growth. At the end of testing, the measured maximum peak current (and therefore pyocyanin concentration) was reduced by approximately 68% and 82% in P. aeruginosa exposed to 16 and 100 mg L(-1) colistin sulfate, respectively. Samples were removed from the microfluidic chamber, analyzed for viability using staining, and streaked onto culture plates to confirm that the P. aeruginosa cells were affected by the antibiotics. The correlation between electrical signal drop and the viability of P. aeruginosa cells after antibiotic exposure highlights the usefulness of this approach for future low cost antibiotic screening applications.