A novel LED‐based 2D‐fluorescence spectroscopy system for in‐line bioprocess monitoring of Chinese hamster ovary cell cultivations—Part II

Abstract

This study was performed in order to evaluate a new LED‐based 2D‐fluorescence spectrometer for in‐line bioprocess monitoring of Chinese hamster ovary (CHO) cell culture processes. The new spectrometer used selected excitation wavelengths of 280, 365, and 455 nm to collect spectral data from six 10‐L fed‐batch processes. The technique provides data on various fluorescent compounds from the cultivation medium as well as from cell metabolism. In addition, scattered light offers information about the cultivation status. Multivariate data analysis tools were applied to analyze the large data sets of the collected fluorescence spectra. First, principal component analysis was used to accomplish an overview of all spectral data from all six CHO cultivations. Partial least square regression models were developed to correlate 2D‐fluorescence spectral data with selected critical process variables as offline reference values. A separate independent fed‐batch process was used for model validation and prediction. An almost continuous in‐line bioprocess monitoring was realized because 2D‐fluorescence spectra were collected every 10 min during the whole cultivation. The new 2D‐fluorescence device demonstrates the significant potential for accurate prediction of the total cell count, viable cell count, and the cell viability. The results strongly indicated that the technique is particularly capable to distinguish between different cell statuses inside the bioreactor. In addition, spectral data provided information about the lactate metabolism shift and cellular respiration during the cultivation process. Overall, the 2D‐fluorescence device is a highly sensitive tool for process analytical technology applications in mammalian cell cultures.

Modern Sensor Tools and Techniques for Monitoring, Controlling, and Improving Cell Culture Processes

“…The 2-D fluorescence spectrometry has been used along with multivariate data analysis to differentiate between viable, dead, and lysed cell populations in mammalian cell culture. This is of value given that, with standard methodologies, it can be problematic to differentiate between populations with high resolution and accuracy [64].…”

Realization of user‐friendly bioanalytical tools to quantify and monitor critical components in bio‐industrial processes through conceptual design

“…Limits in the figure depend on analyte and process. As the arrows and positions of the citations [14–52] illuminate, many of the methodologies have undergone gradual improvements in performance. These achievements have moved their placement of the circular diagrams towards the intersected areas.…”

“…The strength of these methods is the simplicity of transduction of the signal, compared to methods with biorecognition or other reaction schemes (through antibodies, enzymes, samples treatments, etc.) [14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29].…”

“…with biosensors based on enzyme electrodes, microgravity sensors, and thermal biosensors) but few are used in industrial practice and if so, only in process R&D. Successful commercially examples could be noted, F I G U R E 5 Analytical performance with critical limits for response time, sensitivity, selectivity, and cost effectivity. Selected methodologies are cited in the diagram: near-infrared- [15][16][17], 2D fluorescence- [18][19][20], and Raman spectroscopies [21][22][23], in-situ microscopy [24][25][26], holography [27,28], calorimetry [29,30], enzyme electrodes [31], enzyme thermistors [32][33][34], localized surface plasmon resonance [35], immuno-capacitive sensors [36][37][38][39], electrodes [40], aptamers [45], molecular imprinted polymers [47], electronic noses and tongues [49], and lateral flow sensors [52]. Optical and spectroscopic methods labelled blue, calorimetric red, methods based on biorecognition green, other methods with grey.…”

Implementation of QbD strategies in the inoculum expansion of a mAb production process

“…This work will focus on the QbD implementation of a monoclonal Antibody (mAb) production process using a Chinese Hamster Ovary (CHO) cell cultivation process [14, 15]. Concrete QbD case studies, like the CMC Biotech Working Group for mAb production processes and the following downstream part are published [16].…”