Abstract
Antibody titer and viable cell density (VCD) are two important parameters that need to be closely monitored during the process of cell line development and manufacturing of therapeutic antibodies. Typically, determination of each parameter requires 10–100 μL of supernatant sample, which is not suitable for small scale cultivation. In this study, we demonstrated that as low as 2 μL of culture supernatants were sufficient for the analysis using UV-Vis spectrum assisted with partial least squares (PLS) model. The results indicated that the optimal PLS models could be used to predict antibody titer and VCD with the linear relationship between reference values and predicted values at R2 values ranging from 0.8 to > 0.9 in supernatant samples obtained from four different single clones and in polyclones that were cultured in various selection stringencies. Then, the percentage of cell viability and productivity were predicted from a set of samples of polyclones. The results indicated that while all predicted % cell viability were very similar to the actual value at RSEP value of 6.7 and R2 of 0.8908, the predicted productivity from 14 of 18 samples were closed to the reference measurements at RSEP value of 22.4 and R2 of 0.8522. These results indicated that UV-Vis combined with PLS has potential to be used for monitoring antibody titer, VCD, and % cell viability for both online and off-line therapeutic production process.Graphical abstractThe process of multivariate analysis and partial least squares regression of UV-Vis spectrum for the determination of CHO cell densities and antibody titers obtained from small volume of cell culture supernatant samples.
Highlights
Since the production of therapeutic antibodies involves the use of living cells, the manufacturing process is much more complicated than generating small molecule drugs, which requires chemical reactions [1]
The investigations were conducted on four different single clones that were cultured in the same culture conditions, and 14 polyclones that were cultured under 4 different MTX selection pressures; a total of 70 samples were used in the assay
From 48 samples obtained from different passages of four different single clones, 30 samples were used to construct the calibration model for the prediction of antibody titer and only 27 samples were used to construct the calibration model for the prediction of viable cell density (VCD) because 3 samples were outlier sample and was not selected for the analysis
Summary
Since the production of therapeutic antibodies involves the use of living cells, the manufacturing process is much more complicated than generating small molecule drugs, which requires chemical reactions [1] Multiple bioprocess parameters such as viable cell density (VCD), antibody titer, pH, temperature, %CO2, and level of metabolites need to be tightly monitored in order to obtain maximum yield and to ensure consistency of product quality [2]. Ultraviolet-Visible (UV-Vis) spectra provide less information when compared with other spectroscopy techniques as the bands are less specific and have high chances of spectral overlapping [17] This method is more attractive in terms of ease of handling and cost, which is more suitable for lab-scale or manufacturing of therapeutic antibody in low-to-middle-income countries. There has been a previous report on successful application of this technique to monitor glutamine and glucose concentrations in BHK-21 cell cultures [18]
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