The development of next‐generation molecules such as bispecific antibodies, and biosimilars, will... more The development of next‐generation molecules such as bispecific antibodies, and biosimilars, will lead to products tailored to more specific patient groups. Therefore, these molecules will need to be produced in lower quantities, compared to the blockbusters of recent years. This calls for a change in manufacturing strategy. Recent advances in cell culturing technologies have made it possible to reach higher titers in upstream processes, through different types of manufacturing scenarios. This creates the opportunity to move away from classical 12000l stainless‐steel facilities into flexible facilities, set up around 6×2000l single‐use bioreactors feeding into a single DSP train. This chapter describes the considerations during the design phase of the intensified facility design concept. Several approaches can be taken upstream. One option is to intensify the N‐1 seed bioreactor, to shorten the seed train duration, and make it possible to inoculate the pro- duction bioreactor at a high cell density. Another option is to use a concentrated fed‐batch mode of operation in the production bioreactor. Both strategies will increase the upstream titer. Harvest and purification technologies considered for implementation in the platform process need to accommodate the variability in titer upstream. As analyzed, this facility can yield 500–2000 kg/yr, depending on the upstream manufacturing scenario at a COGs as low as 50 $/g.
The Hyclone Single Use Bioreactors (SUBs) are a new type of disposable bioreactors that mimic cla... more The Hyclone Single Use Bioreactors (SUBs) are a new type of disposable bioreactors that mimic classical stainless steel stirred tank bioreactors, but at a fraction of the costs. We find the systems to be of particular use for our multipurpose cGMP facility. With minimal or no modifications we can hook-up 250 L SUBs to our existing roll-on roll-off controllers for autoclavable 50–100 L bioreactors, thereby increasing DSMs clinical production capacity considerably without significant investments in utilities or the facility. Additional up-sides are the elimination of product carry-over risk, and the reduced changeover time and costs.
The CELL-tainer™ is a novel wave-motion bioreactor characterized by very high mass-transfer capac... more The CELL-tainer™ is a novel wave-motion bioreactor characterized by very high mass-transfer capacities (kLa up to 200 h−1) due to an optimized wave motion. By comparison, this mass-transfer capacity is at least a factor 5 higher than that of stirred tank cell-culture bioreactors. As such, no oxygen supplementation of the overlay is required for oxygenation and the carbon dioxide stripping capacity
The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied syste... more The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied systematically using a statistical experimental design. Aim was to improve the separation of hybridoma cells and their IgG product, by identifying the key variables governing IgG partitioning, and by comparing the IgG partitioning data with the hybridoma cell partitioning data obtained in previous work. The influence of five factors, i.e. the poly(ethylene glycol) molecular weight (PEG Mw), dextran molecular weight (Dx Mw), tie-line length (TLL), pH and potassium phosphate fraction (KPi/(KPi+KCl)), on IgG partitioning was characterized using a full-factorial experimental design. In all of the ATPS's the IgG partitioned predominantly into the lower phase. The partition coefficient varied between 0.78 (Variable settings: PEG Mw = 6000, Dx Mw = 500000, TLL = 0.10 g g-1, KPi/(KPi+KCl) = 1.0 and pH = 7.4) and 0.0002 (Variable settings: PEG Mw = 35000, Dx Mw = 40000, TLL = 0.20 g g-1 KPi/(KPi+KCl) = 1.0 and pH 6.6). The tie-line length, the dextran molecular weight and the PEG molecular weight had the most pronounced effect on IgG partitioning. Matching the partitioning data of the IgG product with previously obtained data of the hybridoma cell partitioning, showed that within the experimental design no ATPS could be found giving a good separation of the hybridoma cells and their IgG product. There are, however, ATPS's available in which the cells partition to, and grow in the lower dextran-rich phase. To achieve a good separation of the hybridoma cells and their IgG product in these ATPSs, the IgG product has to be specifically extracted into the PEG-rich top phase. For this purpose the use affinity ligands coupled to PEG may offer a solution. Therefore, a number of commercially available dye-resins was screened for their ability to bind the BIF6A7 IgG antibody. The mimetic green 1 A6XL dye-resin was found to bind BIF6A7 IgG. The dye-ligand coupled to PEG was used to manipulate the IgG partitioning in an ATPS. In the presence of the PEG-ligand, the IgG partitioned almost completely to the top phase. The IgG-partition coefficient increased three orders of magnitude, resulting in a 25-fold higher IgG concentration in the top phase than in the bottom phase.
Two of the big challenges in modern bioprocesses are process economics and in-depth process under... more Two of the big challenges in modern bioprocesses are process economics and in-depth process understanding. Getting access to online process data helps to understand process dynamics and monitor critical process parameters (CPPs). This is an important part of the quality-by-design concept that was introduced to the pharmaceutical industry in the last decade. Raman spectroscopy has proven to be a versatile tool to allow noninvasive measurements and access to a broad spectrum of analytes. This information can then be used for enhanced process control strategies. This review article will focus on the latest applications of Raman spectroscopy in established protein production bioprocesses as well as show its potential for virus, cell therapy, and mRNA processes.
Two of the big challenges in modern bioprocesses are process economics and in-depth process under... more Two of the big challenges in modern bioprocesses are process economics and in-depth process understanding. Getting access to online process data helps to understand process dynamics and monitor critical process parameters (CPPs). This is an important part of the quality-by-design concept that was introduced to the pharmaceutical industry in the last decade. Raman spectroscopy has proven to be a versatile tool to allow noninvasive measurements and access to a broad spectrum of analytes. This information can then be used for enhanced process control strategies. This review article will focus on the latest applications of Raman spectroscopy in established protein production bioprocesses as well as show its potential for virus, cell therapy, and mRNA processes.
The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied syste... more The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied systematically using a statistical experimental design. Aim was to improve the separation of hybridoma cells and their IgG product, by identifying the key variables governing IgG partitioning, and by comparing the IgG partitioning data with the hybridoma cell partitioning data obtained in previous work. The influence of five factors, i.e. the poly(ethylene glycol) molecular weight (PEG Mw), dextran molecular weight (Dx Mw), tie-line length (TLL), pH and potassium phosphate fraction (KPi/(KPi+KCl)), on IgG partitioning was characterized using a full-factorial experimental design. In all of the ATPS's the IgG partitioned predominantly into the lower phase. The partition coefficient varied between 0.78 (Variable settings: PEG Mw = 6000, Dx Mw = 500000, TLL = 0.10 g g-1, KPi/(KPi+KCl) = 1.0 and pH = 7.4) and 0.0002 (Variable settings: PEG Mw = 35000, Dx Mw = 40000, TLL = 0.20 g g-1 KPi/(KPi+K...
The effect of the important ATPS- and buffer parameters on IgG and hybridoma partitioning in ATPS... more The effect of the important ATPS- and buffer parameters on IgG and hybridoma partitioning in ATPSs containing a PEG-dye-ligand was studied. Objective was to establish selection criteria for effective ligands for extractive fermentations with animal cells in ATPSs. In the presence of 1% PEG-dye-ligand the binding of IgG to the PEG-ligand was affected severely by the Na-chloride concentration. The tie-line length and pH affected IgG partitioning to a lesser extent. The desired partitioning of IgG into the top phase, was only obtained when, in addition to the 10 mmollkg K-phosphate buffer, no Na-chloride was present. In an ATPS culture medium, with ± 35 mmollkg Na-bicarbonate and 60 mmolikg Na-chloride, increasing the PEG-dye-ligand concentration up to 100% did increase the partition coefficient, but was not effective in concentrating the IgG in the top phase of ATPS culture medium at a pH of 7.8. Furthermore, addition of the PEG-dye-ligand to ATPS culture medium changed the hybridoma cell partitioning from the bottom phase to the interface.
The design of aqueous two-phase systems (ATPSs) which support the long-term growth of animal cell... more The design of aqueous two-phase systems (ATPSs) which support the long-term growth of animal cells is described in this paper. It was found that the increase in osmolality caused by the ATPS-forming polymers could be compensated by reducing the NaCl concentration of the culture medium. Cell growth was possible in culture media containing up to 0.025 g g-1 PEG or 0.15 g g-1 dextran. In ATPSs of PEG 35,000; dextran 40,000; and culture medium; the hybridoma cells partitioned to the PEG-lean phase. In two of these ATPSs, hybridoma cells were successfully cultured over a period of snore than two months. The Mab product, however, partitioned along with the cells in the lower phase, but preliminary experiments using PEG ligands showed improved Mab partitioning.
The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied syste... more The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied systematically using a statistical experimental design. im was to improve the separation of hybridoma cells and their IgG product, by identifying the key variables governing IgG partitioning and by comparing the IgG partitioning data with the hybridoma cell partitioning data obtained in previous work.he influence of five factors, i.e. the poly(ethylene glycol) molecular weight (PEG Mw), dextran rnolecular weight (Dx Mw), tie-line length (TLL), pH and potassium phosphate fraction (KPi/(KPi+KCl)), on IgG partitioning was characterized using a full-factorial experimental design. In al of the ATPS’s the IgG artitioned predominantly into the lower phase. The partition coefficient varied between 0.78 (Variable settings: EG Mw = 6000, Dx Mw = 500 000, TLL = 0.10 g g-1, KPi/(KPi+KCl) = 1.0 and pH = 7.4) and 0.0002 (Variable settings PEG Mw = 35 000, Dx Mw = 40 000, TLL = 0.20 g g-1 KPi/(KPi+KCl) = 1.0 and pH 6.6). The tie-line length, the dextran molecular weight and the PEG molecular weight had the most pronounced effect on IgG partitioning. Matching the partitioning data of the IgG product with previously obtained data of the hybridoma cell partitioning, showed that within the experimental design no ATPS could be found giving a good separation of the hybridoma cells and their IgG product. There are, however, ATPS’s available in which the cells partition to, and grow in the lower dextran-rich phase. To achieve good separation of the hybridoma cells and their IgG product in these ATPSs, the IgG product has to be specifically extracted into the PEG-rich top phase. For this purpose the use affinity ligands coupled to PEG may offer a solution. Therefore, a number of commercially available dye-resins was screened for their ability to bind the BIF6A7 IgG antibody. The mimetic green 1 A6XL dye-resin was found to bind BIF6A7 IgG. The dye-liganupled to PEG was used to manipulate the IgG partitioning in an ATPS. In the presence of the PEG-ligand, the IgGpartitioned almost completely to the top phase. The IgG-partition coefficient increased three orders of magnitude, resulting in a 25-fold higher IgG concentration in the top phase than in the bottom phase.
The invention relates to a process for the culturing of cels, preferably E1-immortalized HER cels... more The invention relates to a process for the culturing of cels, preferably E1-immortalized HER cels, more preferably PER.C6 cells in a reactor in suspension in a cell culture medium, wherein the cels produce a biological substance, preferably an antibody, wherein at least one cell culture medium component is fed to the cellculture and wherein the cell culture comprising the cels, the biological substance and cell culture medium is circulated over a separation system and where in the separation system separates the biological substance from substances having a lower molecular weight than the biological substance and where in the biological substance is retained in or fed back into the reactor. Preferably part of the substances of lower molecular weight is continuously removed from the cell culture.
Single‐Use Technology in Biopharmaceutical Manufacture, 2019
The development of next‐generation molecules such as bispecific antibodies, and biosimilars, will... more The development of next‐generation molecules such as bispecific antibodies, and biosimilars, will lead to products tailored to more specific patient groups. Therefore, these molecules will need to be produced in lower quantities, compared to the blockbusters of recent years. This calls for a change in manufacturing strategy.
Recent advances in cell culturing technologies have made it possible to reach higher titers in upstream processes, through different types of manufacturing scenarios. This creates the opportunity to move away from classical 12000l stainless‐steel facilities into flexible facilities, set up around 6×2000l single‐use bioreactors feeding into a single DSP train.
This chapter describes the considerations during the design phase of the intensified facility design concept. Several approaches can be taken upstream. One option is to intensify the N‐1 seed bioreactor, to shorten the seed train duration, and make it possible to inoculate the pro- duction bioreactor at a high cell density. Another option is to use a concentrated fed‐batch mode of operation in the production bioreactor. Both strategies will increase the upstream titer. Harvest and purification technologies considered for implementation in the platform process need to accommodate the variability in titer upstream. As analyzed, this facility can yield 500–2000 kg/yr, depending on the upstream manufacturing scenario at a COGs as low as 50 $/g.
Process for the culturing of eukaryotic cells in a reactor in suspension in a cell culture medium... more Process for the culturing of eukaryotic cells in a reactor in suspension in a cell culture medium, wherein the cells produce a recombinant protein, wherein at least one cell culture medium component is fed to the cell culture and wherein the cell culture comprising the cells, the recombinant protein and cell culture medium is circulated over a filter in a flow substantially parallel to a surface of said filter resulting in a liquid outflow and a flow which contents are kept in or fed back into the reactor, wherein the filter has a pore size suitable to separate the recombinant protein from substances having a lower molecular weight than the recombinant protein and wherein the liquid outflow consists essentially of components having a molecular weight lower than that of the recombinant protein and wherein the recombinant protein is retained in or fed back into the reactor.
The development of next‐generation molecules such as bispecific antibodies, and biosimilars, will... more The development of next‐generation molecules such as bispecific antibodies, and biosimilars, will lead to products tailored to more specific patient groups. Therefore, these molecules will need to be produced in lower quantities, compared to the blockbusters of recent years. This calls for a change in manufacturing strategy. Recent advances in cell culturing technologies have made it possible to reach higher titers in upstream processes, through different types of manufacturing scenarios. This creates the opportunity to move away from classical 12000l stainless‐steel facilities into flexible facilities, set up around 6×2000l single‐use bioreactors feeding into a single DSP train. This chapter describes the considerations during the design phase of the intensified facility design concept. Several approaches can be taken upstream. One option is to intensify the N‐1 seed bioreactor, to shorten the seed train duration, and make it possible to inoculate the pro- duction bioreactor at a high cell density. Another option is to use a concentrated fed‐batch mode of operation in the production bioreactor. Both strategies will increase the upstream titer. Harvest and purification technologies considered for implementation in the platform process need to accommodate the variability in titer upstream. As analyzed, this facility can yield 500–2000 kg/yr, depending on the upstream manufacturing scenario at a COGs as low as 50 $/g.
The Hyclone Single Use Bioreactors (SUBs) are a new type of disposable bioreactors that mimic cla... more The Hyclone Single Use Bioreactors (SUBs) are a new type of disposable bioreactors that mimic classical stainless steel stirred tank bioreactors, but at a fraction of the costs. We find the systems to be of particular use for our multipurpose cGMP facility. With minimal or no modifications we can hook-up 250 L SUBs to our existing roll-on roll-off controllers for autoclavable 50–100 L bioreactors, thereby increasing DSMs clinical production capacity considerably without significant investments in utilities or the facility. Additional up-sides are the elimination of product carry-over risk, and the reduced changeover time and costs.
The CELL-tainer™ is a novel wave-motion bioreactor characterized by very high mass-transfer capac... more The CELL-tainer™ is a novel wave-motion bioreactor characterized by very high mass-transfer capacities (kLa up to 200 h−1) due to an optimized wave motion. By comparison, this mass-transfer capacity is at least a factor 5 higher than that of stirred tank cell-culture bioreactors. As such, no oxygen supplementation of the overlay is required for oxygenation and the carbon dioxide stripping capacity
The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied syste... more The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied systematically using a statistical experimental design. Aim was to improve the separation of hybridoma cells and their IgG product, by identifying the key variables governing IgG partitioning, and by comparing the IgG partitioning data with the hybridoma cell partitioning data obtained in previous work. The influence of five factors, i.e. the poly(ethylene glycol) molecular weight (PEG Mw), dextran molecular weight (Dx Mw), tie-line length (TLL), pH and potassium phosphate fraction (KPi/(KPi+KCl)), on IgG partitioning was characterized using a full-factorial experimental design. In all of the ATPS's the IgG partitioned predominantly into the lower phase. The partition coefficient varied between 0.78 (Variable settings: PEG Mw = 6000, Dx Mw = 500000, TLL = 0.10 g g-1, KPi/(KPi+KCl) = 1.0 and pH = 7.4) and 0.0002 (Variable settings: PEG Mw = 35000, Dx Mw = 40000, TLL = 0.20 g g-1 KPi/(KPi+KCl) = 1.0 and pH 6.6). The tie-line length, the dextran molecular weight and the PEG molecular weight had the most pronounced effect on IgG partitioning. Matching the partitioning data of the IgG product with previously obtained data of the hybridoma cell partitioning, showed that within the experimental design no ATPS could be found giving a good separation of the hybridoma cells and their IgG product. There are, however, ATPS's available in which the cells partition to, and grow in the lower dextran-rich phase. To achieve a good separation of the hybridoma cells and their IgG product in these ATPSs, the IgG product has to be specifically extracted into the PEG-rich top phase. For this purpose the use affinity ligands coupled to PEG may offer a solution. Therefore, a number of commercially available dye-resins was screened for their ability to bind the BIF6A7 IgG antibody. The mimetic green 1 A6XL dye-resin was found to bind BIF6A7 IgG. The dye-ligand coupled to PEG was used to manipulate the IgG partitioning in an ATPS. In the presence of the PEG-ligand, the IgG partitioned almost completely to the top phase. The IgG-partition coefficient increased three orders of magnitude, resulting in a 25-fold higher IgG concentration in the top phase than in the bottom phase.
Two of the big challenges in modern bioprocesses are process economics and in-depth process under... more Two of the big challenges in modern bioprocesses are process economics and in-depth process understanding. Getting access to online process data helps to understand process dynamics and monitor critical process parameters (CPPs). This is an important part of the quality-by-design concept that was introduced to the pharmaceutical industry in the last decade. Raman spectroscopy has proven to be a versatile tool to allow noninvasive measurements and access to a broad spectrum of analytes. This information can then be used for enhanced process control strategies. This review article will focus on the latest applications of Raman spectroscopy in established protein production bioprocesses as well as show its potential for virus, cell therapy, and mRNA processes.
Two of the big challenges in modern bioprocesses are process economics and in-depth process under... more Two of the big challenges in modern bioprocesses are process economics and in-depth process understanding. Getting access to online process data helps to understand process dynamics and monitor critical process parameters (CPPs). This is an important part of the quality-by-design concept that was introduced to the pharmaceutical industry in the last decade. Raman spectroscopy has proven to be a versatile tool to allow noninvasive measurements and access to a broad spectrum of analytes. This information can then be used for enhanced process control strategies. This review article will focus on the latest applications of Raman spectroscopy in established protein production bioprocesses as well as show its potential for virus, cell therapy, and mRNA processes.
The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied syste... more The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied systematically using a statistical experimental design. Aim was to improve the separation of hybridoma cells and their IgG product, by identifying the key variables governing IgG partitioning, and by comparing the IgG partitioning data with the hybridoma cell partitioning data obtained in previous work. The influence of five factors, i.e. the poly(ethylene glycol) molecular weight (PEG Mw), dextran molecular weight (Dx Mw), tie-line length (TLL), pH and potassium phosphate fraction (KPi/(KPi+KCl)), on IgG partitioning was characterized using a full-factorial experimental design. In all of the ATPS's the IgG partitioned predominantly into the lower phase. The partition coefficient varied between 0.78 (Variable settings: PEG Mw = 6000, Dx Mw = 500000, TLL = 0.10 g g-1, KPi/(KPi+KCl) = 1.0 and pH = 7.4) and 0.0002 (Variable settings: PEG Mw = 35000, Dx Mw = 40000, TLL = 0.20 g g-1 KPi/(KPi+K...
The effect of the important ATPS- and buffer parameters on IgG and hybridoma partitioning in ATPS... more The effect of the important ATPS- and buffer parameters on IgG and hybridoma partitioning in ATPSs containing a PEG-dye-ligand was studied. Objective was to establish selection criteria for effective ligands for extractive fermentations with animal cells in ATPSs. In the presence of 1% PEG-dye-ligand the binding of IgG to the PEG-ligand was affected severely by the Na-chloride concentration. The tie-line length and pH affected IgG partitioning to a lesser extent. The desired partitioning of IgG into the top phase, was only obtained when, in addition to the 10 mmollkg K-phosphate buffer, no Na-chloride was present. In an ATPS culture medium, with ± 35 mmollkg Na-bicarbonate and 60 mmolikg Na-chloride, increasing the PEG-dye-ligand concentration up to 100% did increase the partition coefficient, but was not effective in concentrating the IgG in the top phase of ATPS culture medium at a pH of 7.8. Furthermore, addition of the PEG-dye-ligand to ATPS culture medium changed the hybridoma cell partitioning from the bottom phase to the interface.
The design of aqueous two-phase systems (ATPSs) which support the long-term growth of animal cell... more The design of aqueous two-phase systems (ATPSs) which support the long-term growth of animal cells is described in this paper. It was found that the increase in osmolality caused by the ATPS-forming polymers could be compensated by reducing the NaCl concentration of the culture medium. Cell growth was possible in culture media containing up to 0.025 g g-1 PEG or 0.15 g g-1 dextran. In ATPSs of PEG 35,000; dextran 40,000; and culture medium; the hybridoma cells partitioned to the PEG-lean phase. In two of these ATPSs, hybridoma cells were successfully cultured over a period of snore than two months. The Mab product, however, partitioned along with the cells in the lower phase, but preliminary experiments using PEG ligands showed improved Mab partitioning.
The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied syste... more The partitioning of IgG in aqueous two-phase systems (ATPSs) of PEG and Dextran was studied systematically using a statistical experimental design. im was to improve the separation of hybridoma cells and their IgG product, by identifying the key variables governing IgG partitioning and by comparing the IgG partitioning data with the hybridoma cell partitioning data obtained in previous work.he influence of five factors, i.e. the poly(ethylene glycol) molecular weight (PEG Mw), dextran rnolecular weight (Dx Mw), tie-line length (TLL), pH and potassium phosphate fraction (KPi/(KPi+KCl)), on IgG partitioning was characterized using a full-factorial experimental design. In al of the ATPS’s the IgG artitioned predominantly into the lower phase. The partition coefficient varied between 0.78 (Variable settings: EG Mw = 6000, Dx Mw = 500 000, TLL = 0.10 g g-1, KPi/(KPi+KCl) = 1.0 and pH = 7.4) and 0.0002 (Variable settings PEG Mw = 35 000, Dx Mw = 40 000, TLL = 0.20 g g-1 KPi/(KPi+KCl) = 1.0 and pH 6.6). The tie-line length, the dextran molecular weight and the PEG molecular weight had the most pronounced effect on IgG partitioning. Matching the partitioning data of the IgG product with previously obtained data of the hybridoma cell partitioning, showed that within the experimental design no ATPS could be found giving a good separation of the hybridoma cells and their IgG product. There are, however, ATPS’s available in which the cells partition to, and grow in the lower dextran-rich phase. To achieve good separation of the hybridoma cells and their IgG product in these ATPSs, the IgG product has to be specifically extracted into the PEG-rich top phase. For this purpose the use affinity ligands coupled to PEG may offer a solution. Therefore, a number of commercially available dye-resins was screened for their ability to bind the BIF6A7 IgG antibody. The mimetic green 1 A6XL dye-resin was found to bind BIF6A7 IgG. The dye-liganupled to PEG was used to manipulate the IgG partitioning in an ATPS. In the presence of the PEG-ligand, the IgGpartitioned almost completely to the top phase. The IgG-partition coefficient increased three orders of magnitude, resulting in a 25-fold higher IgG concentration in the top phase than in the bottom phase.
The invention relates to a process for the culturing of cels, preferably E1-immortalized HER cels... more The invention relates to a process for the culturing of cels, preferably E1-immortalized HER cels, more preferably PER.C6 cells in a reactor in suspension in a cell culture medium, wherein the cels produce a biological substance, preferably an antibody, wherein at least one cell culture medium component is fed to the cellculture and wherein the cell culture comprising the cels, the biological substance and cell culture medium is circulated over a separation system and where in the separation system separates the biological substance from substances having a lower molecular weight than the biological substance and where in the biological substance is retained in or fed back into the reactor. Preferably part of the substances of lower molecular weight is continuously removed from the cell culture.
Single‐Use Technology in Biopharmaceutical Manufacture, 2019
The development of next‐generation molecules such as bispecific antibodies, and biosimilars, will... more The development of next‐generation molecules such as bispecific antibodies, and biosimilars, will lead to products tailored to more specific patient groups. Therefore, these molecules will need to be produced in lower quantities, compared to the blockbusters of recent years. This calls for a change in manufacturing strategy.
Recent advances in cell culturing technologies have made it possible to reach higher titers in upstream processes, through different types of manufacturing scenarios. This creates the opportunity to move away from classical 12000l stainless‐steel facilities into flexible facilities, set up around 6×2000l single‐use bioreactors feeding into a single DSP train.
This chapter describes the considerations during the design phase of the intensified facility design concept. Several approaches can be taken upstream. One option is to intensify the N‐1 seed bioreactor, to shorten the seed train duration, and make it possible to inoculate the pro- duction bioreactor at a high cell density. Another option is to use a concentrated fed‐batch mode of operation in the production bioreactor. Both strategies will increase the upstream titer. Harvest and purification technologies considered for implementation in the platform process need to accommodate the variability in titer upstream. As analyzed, this facility can yield 500–2000 kg/yr, depending on the upstream manufacturing scenario at a COGs as low as 50 $/g.
Process for the culturing of eukaryotic cells in a reactor in suspension in a cell culture medium... more Process for the culturing of eukaryotic cells in a reactor in suspension in a cell culture medium, wherein the cells produce a recombinant protein, wherein at least one cell culture medium component is fed to the cell culture and wherein the cell culture comprising the cells, the recombinant protein and cell culture medium is circulated over a filter in a flow substantially parallel to a surface of said filter resulting in a liquid outflow and a flow which contents are kept in or fed back into the reactor, wherein the filter has a pore size suitable to separate the recombinant protein from substances having a lower molecular weight than the recombinant protein and wherein the liquid outflow consists essentially of components having a molecular weight lower than that of the recombinant protein and wherein the recombinant protein is retained in or fed back into the reactor.
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Papers by Gerben Zijlstra
There are, however, ATPS’s available in which the cells partition to, and grow in the lower dextran-rich phase. To achieve good separation of the hybridoma cells and their IgG product in these ATPSs, the IgG product has to be specifically extracted into the PEG-rich top phase. For this purpose the use affinity ligands coupled to PEG may offer a solution.
Therefore, a number of commercially available dye-resins was screened for their ability to bind the BIF6A7 IgG antibody. The mimetic green 1 A6XL dye-resin was found to bind BIF6A7 IgG. The dye-liganupled to PEG was used to manipulate the IgG partitioning in an ATPS. In the presence of the PEG-ligand, the IgGpartitioned almost completely to the top phase. The IgG-partition coefficient increased three orders of magnitude, resulting in a 25-fold higher IgG concentration in the top phase than in the bottom phase.
Recent advances in cell culturing technologies have made it possible to reach higher titers in upstream processes, through different types of manufacturing scenarios. This creates the opportunity to move away from classical 12000l stainless‐steel facilities into flexible facilities, set up around 6×2000l single‐use bioreactors feeding into a single DSP train.
This chapter describes the considerations during the design phase of the intensified facility design concept. Several approaches can be taken upstream. One option is to intensify the N‐1 seed bioreactor, to shorten the seed train duration, and make it possible to inoculate the pro- duction bioreactor at a high cell density. Another option is to use a concentrated fed‐batch mode of operation in the production bioreactor. Both strategies will increase the upstream titer. Harvest and purification technologies considered for implementation in the platform process need to accommodate the variability in titer upstream. As analyzed, this facility can yield 500–2000 kg/yr, depending on the upstream manufacturing scenario at a COGs as low as 50 $/g.
There are, however, ATPS’s available in which the cells partition to, and grow in the lower dextran-rich phase. To achieve good separation of the hybridoma cells and their IgG product in these ATPSs, the IgG product has to be specifically extracted into the PEG-rich top phase. For this purpose the use affinity ligands coupled to PEG may offer a solution.
Therefore, a number of commercially available dye-resins was screened for their ability to bind the BIF6A7 IgG antibody. The mimetic green 1 A6XL dye-resin was found to bind BIF6A7 IgG. The dye-liganupled to PEG was used to manipulate the IgG partitioning in an ATPS. In the presence of the PEG-ligand, the IgGpartitioned almost completely to the top phase. The IgG-partition coefficient increased three orders of magnitude, resulting in a 25-fold higher IgG concentration in the top phase than in the bottom phase.
Recent advances in cell culturing technologies have made it possible to reach higher titers in upstream processes, through different types of manufacturing scenarios. This creates the opportunity to move away from classical 12000l stainless‐steel facilities into flexible facilities, set up around 6×2000l single‐use bioreactors feeding into a single DSP train.
This chapter describes the considerations during the design phase of the intensified facility design concept. Several approaches can be taken upstream. One option is to intensify the N‐1 seed bioreactor, to shorten the seed train duration, and make it possible to inoculate the pro- duction bioreactor at a high cell density. Another option is to use a concentrated fed‐batch mode of operation in the production bioreactor. Both strategies will increase the upstream titer. Harvest and purification technologies considered for implementation in the platform process need to accommodate the variability in titer upstream. As analyzed, this facility can yield 500–2000 kg/yr, depending on the upstream manufacturing scenario at a COGs as low as 50 $/g.