scholarly journals Model Transferability and Reduced Experimental Burden in Cell Culture Process Development Facilitated by Hybrid Modeling and Intensified Design of Experiments

2021 ◽  
Vol 9 ◽  
Author(s):  
Benjamin Bayer ◽  
Mark Duerkop ◽  
Gerald Striedner ◽  
Bernhard Sissolak
Keyword(s):  
Cell Culture ◽  
Design Of Experiments ◽  
Hybrid Model ◽  
Process Development ◽  
Cell Concentration ◽  
Viable Cell ◽  
Hybrid Modeling ◽  
Bioprocess Development ◽  
Viable Cell Concentration ◽  
Product Titer

Reliable process development is accompanied by intense experimental effort. The utilization of an intensified design of experiments (iDoE) (intra-experimental critical process parameter (CPP) shifts combined) with hybrid modeling potentially reduces process development burden. The iDoE can provide more process response information in less overall process time, whereas hybrid modeling serves as a commodity to describe this behavior the best way. Therefore, a combination of both approaches appears beneficial for faster design screening and is especially of interest at larger scales where the costs per experiment rise significantly. Ideally, profound process knowledge is gathered at a small scale and only complemented with few validation experiments on a larger scale, saving valuable resources. In this work, the transferability of hybrid modeling for Chinese hamster ovary cell bioprocess development along process scales was investigated. A two-dimensional DoE was fully characterized in shake flask duplicates (300 ml), containing three different levels for the cultivation temperature and the glucose concentration in the feed. Based on these data, a hybrid model was developed, and its performance was assessed by estimating the viable cell concentration and product titer in 15 L bioprocesses with the same DoE settings. To challenge the modeling approach, 15 L bioprocesses also comprised iDoE runs with intra-experimental CPP shifts, impacting specific cell rates such as growth, consumption, and formation. Subsequently, the applicability of the iDoE cultivations to estimate static cultivations was also investigated. The shaker-scale hybrid model proved suitable for application to a 15 L scale (1:50), estimating the viable cell concentration and the product titer with an NRMSE of 10.92% and 17.79%, respectively. Additionally, the iDoE hybrid model performed comparably, displaying NRMSE values of 13.75% and 21.13%. The low errors when transferring the models from shaker to reactor and between the DoE and the iDoE approach highlight the suitability of hybrid modeling for mammalian cell culture bioprocess development and the potential of iDoE to accelerate process characterization and to improve process understanding.

Response to phage infection of immobilized lactococci during continuous acidification and inoculation of skim milk

1994 ◽  
Vol 61 (4) ◽  
pp. 537-544 ◽  
Author(s):  
Flavia M. L. Passos ◽  
Todd R. Klaenhammer ◽  
Harold E. Swaisgood
Keyword(s):  
Steady State ◽  
Cell Concentration ◽  
Skim Milk ◽  
Viable Cell ◽  
Free Cell ◽  
Viable Cell Concentration ◽  
Immobilized Cell ◽  
Stainless Steel Mesh ◽  
Immobilized Cell Bioreactor ◽  
Column Bioreactor

SummaryA laboratory scale bioreactor was used for continuous acidification and inoculation of milk with a proteinase-negative, lactose-fermenting strain,Lactococcus lactissubsp.lactisC2S. Calcium alginate-entrapped cells were immobilized on a spiral stainless steel mesh incorporated into a column bioreactor and used to acidify and inoculate reconstituted skim milk. Characteristics of the immobilized cell bioreactor (ICB) were compared with those of a free cell bioreactor (FCB) during challenge with a virulent phage. Steady state biomass and lactate productivities were respectively 25-fold and 12-fold larger with the ICB than with the FCB. The ICB and the FCB were inoculated with the prolate phage c2 at multiplicities of infection of 0·25 and 0·02 respectively. Within 90 min of the infection, the FCB viable cell concentration dropped by five orders of magnitude and never recovered, while the plaque forming units/ml increased dramatically. In the ICB, released cells decreased immediately after infection, but subsequently increased, while the plaque forming units/ml steadily declined, indicating that phage were being washed out of the bioreactor. Productivity of FCB decreased to zero, whereas productivity of the ICB only decreased ∼ 60% and subsequently recovered to its initial steady state value.

scholarly journals Microbes in deionized water: Implications for maintenance of laboratory water production system

2016 ◽  
Author(s):  
Wenfa Ng ◽  
Yen-Peng Ting
Keyword(s):  
Ion Exchange ◽  
Microbial Diversity ◽  
Production System ◽  
Cell Concentration ◽  
Tap Water ◽  
Viable Cell ◽  
Just In Time ◽  
Water Production ◽  
Viable Cell Concentration ◽  
Salt Solutions

Microbes, with their diverse metabolic capabilities and great adaptability, occupy almost every conceivable ecological niche on Earth – thus, could they survive in the oligotrophic (i.e., nutrient-poor) deionized (DI) water that we use for our experiments? Observations of white cauliflower-like lumps and black specks in salt solutions after months of storage in plastic bottles prompted the inquisition concerning the origin and nature of the “contaminants”. Hypothesizing that the “contaminants” may be microbes from DI water, a series of growth experiments was conducted to detect and profile the microbial diversity in fresh DI water - produced on a just-in-time basis by a filter-cum-ion-exchange system with tap water as feed. While microbes could also be present on the surfaces and headspace of the unsterilized polyethylene bottles, investigating whether microbes are present in freshly produced DI water provides a more stringent performance test of the production system. Inoculation of DI water on R2A agar followed by multi-day aerobic cultivation revealed the presence of a wide variety of microbes (total viable cell concentration of ~103 colony forming units (CFU) per mL) with differing pigmentations, growth rates as well as colony sizes and morphologies. Additionally, greater abundance and diversity of microbes was recovered at 30 oC relative to 25 and 37 oC; most probably due to adaptation of microbes to tropical ambient water temperatures of 25 to 30 oC. Comparative experiments with tap water as inoculum recovered a significantly smaller number and diversity of microbes; thereby, suggesting that monochloramine residual disinfectant in tap water was effective in inhibiting cell viability. In contrast, possible removal of monochloramine by adsorption onto ion-exchange resins – and thus, alleviation of a source of environmental stress - might explain the observed greater diversity and abundance of viable microbes in DI water. Collectively, this study confirmed the presence of microbes in fresh DI water – and suggested a possible source of the “contaminants” in prepared salt solutions. Propensity of microbes for forming biofilm on various surfaces suggested that intermittent flow in just-in-time DI water production provided opportunities for cell attachment and biofilm formation in the system during water stagnation, and subsequent dislodgement and resuspension of cells upon water flow. Thus, regular maintenance and cleaning of the production system should help reduce DI water’s microbial load. Additionally, simple and low-cost culture experiments on agar medium can provide a qualitative and semi-quantitative estimate of microbial diversity and viable cell concentration in DI water, respectively, and along with regular monitoring of water resistivity or conductivity, comprise a trio of tests useful for detecting possible contamination, or deterioration of DI water’s chemical and microbiological quality.

scholarly journals Microbes in deionized water: Implications for maintenance of laboratory water production system

2013 ◽  
Author(s):  
Wenfa Ng
Keyword(s):  
Ion Exchange ◽  
Microbial Diversity ◽  
Production System ◽  
Cell Concentration ◽  
Tap Water ◽  
Viable Cell ◽  
Just In Time ◽  
Water Production ◽  
Viable Cell Concentration ◽  
Salt Solutions

Microbes, with their diverse metabolic capabilities and great adaptability, occupy almost every conceivable ecological niche on Earth – thus, could they survive in the oligotrophic (i.e., nutrient-poor) deionized (DI) water that we use for our experiments? Observations of white cauliflower-like lumps and black specks in salt solutions after months of storage in plastic bottles prompted the inquisition concerning the origin and nature of the “contaminants.” Hypothesising that the “contaminants” may be microbes from DI water, a series of growth experiments was conducted to detect and profile the microbial diversity in fresh DI water - produced on a just-in-time basis by a filter-cum-ion-exchange system with tap water as feed. While microbes could also be present on the surfaces and headspace of the unsterilized polyethylene bottles, investigating whether microbes are present in freshly produced DI water provides a more stringent performance test of the production system. Inoculation of DI water on R2A agar followed by multi-day aerobic cultivation revealed the presence of a wide variety of microbes (total viable cell concentration of ~103 colony forming units (CFU) per mL) with differing pigmentations, growth rates as well as colony sizes and morphologies. Additionally, greater abundance and diversity of microbes was recovered at 30 oC relative to 25 and 37 oC; most probably due to adaptation of microbes to tropical ambient water temperatures of 25 to 30 oC. Comparative experiments with tap water as inoculum recovered a significantly smaller number and diversity of microbes; thereby, suggesting that monochloramine residual disinfectant in tap water was effective in inhibiting cell viability. In contrast, possible removal of monochloramine by adsorption onto ion-exchange resins – and thus, alleviation of a source of environmental stress - might explain the observed greater diversity and abundance of viable microbes in DI water. Collectively, this study confirmed the presence of microbes in fresh DI water – and suggested a possible source for the “contaminants” in prepared salt solutions. Propensity of microbes for forming biofilm on various surfaces suggested that intermittent flow in just-in-time DI water production provided opportunities for cell attachment and biofilm formation in the system during water stagnation, and subsequent dislodgement and resuspension of cells upon water flow. Thus, regular maintenance and cleaning of the production system should help reduce DI water’s microbial load. Additionally, simple and low-cost culture experiments on agar medium can provide a qualitative and semi-quantitative estimate of microbial diversity and viable cell concentration in DI water, respectively; which, along with regular monitoring of water resistivity or conductivity, comprise a trio of tests useful for detecting possible contamination or, deterioration of DI water’s chemical and microbiological quality.

Developments in Using Off-Line Radio Frequency Impedance Methods for Measuring the Viable Cell Concentration in the Brewery

2000 ◽  
Vol 58 (2) ◽  
pp. 57-62
Author(s):  
J. P. Carvell ◽  
G. Austin ◽  
A. Matthee ◽  
K. Van de Spiegle ◽  
S. Cunningham ◽  
...  
Keyword(s):  
Radio Frequency ◽  
Cell Concentration ◽  
Viable Cell ◽  
Viable Cell Concentration

Encapsulation of probiotics in soybean protein-based microparticles preserves viable cell concentration in foods all along the production and storage processes

2020 ◽  
Vol 37 (3) ◽  
pp. 242-253 ◽  
Author(s):  
Carolina González-Ferrero ◽  
Juan Manuel Irache ◽  
Beatriz Marín-Calvo ◽  
Leticia Ortiz-Romero ◽  
Raquel Virto-Resano ◽  
...  
Keyword(s):  
Cell Concentration ◽  
Soybean Protein ◽  
Viable Cell ◽  
Viable Cell Concentration ◽  
And Storage ◽  
Storage Processes

Antimicrobial Effectiveness of Lysozyme Immobilized on Polyvinylalcohol-Based Film against Alicyclobacillus acidoterrestris

2006 ◽  
Vol 69 (4) ◽  
pp. 861-865 ◽  
Author(s):  
AMALIA CONTE ◽  
MILENA SINIGAGLIA ◽  
MATTEO ALESSANDRO DEL NOBILE
Keyword(s):  
Active Compound ◽  
Apple Juice ◽  
Cell Concentration ◽  
Viable Cell ◽  
Viable Cell Concentration ◽  
Alicyclobacillus Acidoterrestris ◽  
Single Strain ◽  
Microbial Cells ◽  
Active Film ◽  
Antimicrobial Effectiveness

In this study, the effectiveness of an active polyvinylalcohol-based film against Alicyclobacillus acidoterrestris was assessed. The active film was fabricated by immobilizing an active compound on the surface of a polymeric matrix and then tested by putting the film in contact with a medium that had been inoculated with microbial cells. Microbiological tests showed that the film was antimicrobial against both a single strain and a culture cocktail of A. acidoterrestris, at 44°C. By monitoring the viable cell concentration under three different packaging conditions, it was possible to demonstrate that the active film was equally effective against both the single strain and the culture cocktail and that it maintained this efficacy at various medium volumes. The same microbial tests were also conducted on viable spores of the investigated microorganism, inoculated both into a laboratory medium and apple juice. The results indicate that these viable spores were better inhibited than cells by the active film in both investigated media.

Disinfection kinetics of Legionella pneumophila by ultraviolet irradiation

2002 ◽  
Vol 46 (11-12) ◽  
pp. 311-317 ◽  
Author(s):  
K. Yamagiwa ◽  
M. Tsujikawa ◽  
M. Yoshida ◽  
A. Ohkawa
Keyword(s):  
Rate Constant ◽  
Legionella Pneumophila ◽  
Ultraviolet Irradiation ◽  
Cell Concentration ◽  
Viable Cell ◽  
Lag Time ◽  
Viable Cell Concentration ◽  
Initial Cell Concentration ◽  
Initial Cell ◽  
Kinetics Of

Disinfection kinetics of Legionella pneumophila by ultraviolet irradiation was investigated. The change in viable cell concentration with exposure time could be divided into three steps: lag step in which little change in viable cell concentration was observed, fast disinfection step and slow disinfection step. The slow disinfection step was not observed at the initial cell concentrations below about 106 cfu/mL. The disinfection kinetics were well described with two parameters; lag time and disinfection rate constant of the fast disinfection step. The effects of UV intensity, temperature and initial cell concentration in the kinetic parameters were investigated. With increasing initial cell concentration, the lag time decreased and the disinfection rate constant increased. The effects of initial cell concentration on the kinetic parameters were considered to be attributed to the decrease in the effective UV irradiation intensity due to the partial shield of UV light by the disinfected cells. The empirical correlations were presented for predicting the lag time and disinfection rate constant. Furthermore, UV disinfection of L. pneuophila in a model hot-tub connected with external irradiation chamber was also discussed.

Sign in / Sign up

Export Citation Format

Share Document