Samples were taken daily during the inoculum expansion as well as during the production process. Viable cell densities and viabilities were measured using a Cedex HiRes (Roche Innovatis, Swiss) with a 1:2 dilution in 10 % phosphate‐buffered saline (PBS). Growth rates of the production process were calculated by linear regression of the natural logarithm of viable cell densities against culture duration. The inoculum growth rate relates to the respective last passage of every inoculum expansion (N‐1‐step), in order to ensure optimal comparability of the examined factor effects. Viabilities were calculated as average over the culture duration for the inoculum expansion as well as the production process. The mAb titer was analyzed during the production process by using the Cedex Bio (Roche, Swiss). Therefore, cell separation was performed by centrifugation for 5 min at 190 x g. Specific antibody titers were calculated by dividing the final antibody titer by the integral viable cell concentration over the entire production process duration.
Cedex bio
Manufactured by Roche
Sourced in Denmark
The Cedex Bio is a compact, automated cell analysis system designed for routine cell culture monitoring and analysis. It provides accurate and reliable measurements of cell count, viability, and other key parameters essential for cell-based bioprocessing and research.
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Lab products found in correlation
4 protocols using cedex bio
1
Characterizing mAb Production Process
2
Raman Spectroscopy for Fermentation Monitoring
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The Raman spectrometer, fiber, and ball lens probe used for the measurements
were the same as those in Materials and Methods for Preparing Band Graph
section. The measurement conditions were also the same as those, with a
laser power of 200 mW, exposure time of 3200 ms, and average number of
spectra of 60. Dark spectral measurement was taken before a batch of
culture. Each spectrum took approximately 3 min to be obtained and
recorded.
Off-line measurements for glucose, lactate, and antibody concentrations were
taken once a day under normal conditions. Additional measurements for
glucose and lactate were taken before and after the addition of the feed
agent on the day it was added. In total, Raman spectra of glucose and
lactate were measured 80 times, and those of antibodies were measured 45
times offline in all three fermentations. A 300 µL sampling was taken for
glucose, lactate, and antibody concentrations on an off-line analyzer (Cedex
Bio, Roche).
were the same as those in Materials and Methods for Preparing Band Graph
section. The measurement conditions were also the same as those, with a
laser power of 200 mW, exposure time of 3200 ms, and average number of
spectra of 60. Dark spectral measurement was taken before a batch of
culture. Each spectrum took approximately 3 min to be obtained and
recorded.
Off-line measurements for glucose, lactate, and antibody concentrations were
taken once a day under normal conditions. Additional measurements for
glucose and lactate were taken before and after the addition of the feed
agent on the day it was added. In total, Raman spectra of glucose and
lactate were measured 80 times, and those of antibodies were measured 45
times offline in all three fermentations. A 300 µL sampling was taken for
glucose, lactate, and antibody concentrations on an off-line analyzer (Cedex
Bio, Roche).
3
2D Growth Characterization of hASCs
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2D growth characterization of previously isolated hASCs was performed in precoated T25-flasks (5 µg/cm2 r-fibronectin; Sigma Aldrich, St. Louis, MO, USA) with the UrSuppe stem cell culture medium (5 mL). For this purpose, the cryopreserved, patient-derived hASCs (P1, 080-PDLcum. 3.9, 085-PDLcum. 3.7) were thawed and precultured in T75-flasks (10,000 hASCs/cm2; 37 °C, 5% CO2, 80% rH) in order to achieve the required cell numbers to inoculate 22 × T25-flasks per donor (P2, 080-PDLcum. 6.3, 085-PDLcum. 6.5, 10,000 cells/cm2). The hASC growth characteristics were assessed over 11 days by harvesting two T25-flasks per donor (2 mL TrypLE Select at 37 °C, 2 min) every day. The cell density, substrate and metabolite measurements were carried out using a NucleoCounter NC-200 (Chemometec, Allerod, Denmark) and a Cedex Bio (Roche Diagnostics, Rotkreuz, Switzerland), respectively. In addition to standard T25-flasks, T25-flasks equipped with pH and DO sensor spots (PreSens, Regensburg, Germany) were also inoculated in parallel for each donor in order to assess the pH and DO profiles during cell growth (37 °C, 5% CO2, 80% rH). In each case, partial medium exchanges of 40% and 60% were performed for each donor on days 4 and 8.
4
Comparison of Cell Lines for Antibody Production
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Two Chinese hamster ovary (CHO) cell lines, one human embryo kidney (HEK) 293 and one insect cell line (Trichoplusia ni BTI-Tn-5B1-4, High Five) were used for the animal cell culture experiments (Table 5 ).
Both CHO cell lines produce the antibody immunoglobulin G (IgG) and behave in the same manner as industrial mammalian production cell lines. HEK cells are also widely used, but tend to form larger aggregates [86 (link)]. High Five insect cells have similar morphological characteristics to CHO cells but are cultured without CO2 based pH control and at lower temperatures. The applied cultivation conditions are summarized inTable 6 . The effect of different clones, media and cultivation conditions should test the robustness of the methodology.
Cell density and viability were measured using a Cedex HiRes (Roche Custom Biotech) for the CHO and High Five cells, and with a NucleoCounter NC200 (Chemometec A/S, Allerod, Denmark) for the HEK293 cells. Metabolites (glucose, glutamine, ammonia), as well as the product IgG, were determined using a Cedex Bio (Roche Custom Biotech) and the corresponding analytic kits.
Both CHO cell lines produce the antibody immunoglobulin G (IgG) and behave in the same manner as industrial mammalian production cell lines. HEK cells are also widely used, but tend to form larger aggregates [86 (link)]. High Five insect cells have similar morphological characteristics to CHO cells but are cultured without CO2 based pH control and at lower temperatures. The applied cultivation conditions are summarized in
Cell density and viability were measured using a Cedex HiRes (Roche Custom Biotech) for the CHO and High Five cells, and with a NucleoCounter NC200 (Chemometec A/S, Allerod, Denmark) for the HEK293 cells. Metabolites (glucose, glutamine, ammonia), as well as the product IgG, were determined using a Cedex Bio (Roche Custom Biotech) and the corresponding analytic kits.
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