Скачать презентацию Respiration Activity Monitoring System Hi Tec Zang Скачать презентацию Respiration Activity Monitoring System Hi Tec Zang

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  • Количество слайдов: 44

Respiration Activity Monitoring System © Hi. Tec Zang Gmb. H - HRE Bioprocessoptimisation Respiration Activity Monitoring System © Hi. Tec Zang Gmb. H - HRE Bioprocessoptimisation

The RAMOS® System Online – respiration activity measurement (OTR, CTR, RQ) Respiration Activity Monitoring The RAMOS® System Online – respiration activity measurement (OTR, CTR, RQ) Respiration Activity Monitoring System © Hi. Tec Zang Gmb. H - HRE in shaking flasks

© Hi. Tec Zang Gmb. H - HRE The Tray © Hi. Tec Zang Gmb. H - HRE The Tray

Fields of Application © Hi. Tec Zang Gmb. H - HRE 4 Online-tracing of Fields of Application © Hi. Tec Zang Gmb. H - HRE 4 Online-tracing of the metabolic activity of pro- and eukaryotic cultures in shaking flasks

Possibilities 4 Easy Determination of parameters: - oxygen transfer rate (OTR) - carbon dioxide Possibilities 4 Easy Determination of parameters: - oxygen transfer rate (OTR) - carbon dioxide transfer rate (CTR) - respiration quotient (RQ) - maximum growth rate (µmax) - volumetric oxygen transfer coefficient (k. La) which afford a safe Scale-Up. © Hi. Tec Zang Gmb. H - HRE …,

Possibilities 4 Detection of characteristic biological phenomena (OTR) Substrate limitation (except C-source) Oxygen limitation Possibilities 4 Detection of characteristic biological phenomena (OTR) Substrate limitation (except C-source) Oxygen limitation Oxygen transfer rate Unlimited growth on minimal media Time of fermentation Product inhibition ( e. g. p. H) Diauxic growth Time of fermentation © Hi. Tec Zang Gmb. H - HRE Time of fermentation = Total oxygen consumption [mol/l] Oxygen transfer rate Time of fermentation maximum oxygen transfer capacity

Possibilities © Hi. Tec Zang Gmb. H - HRE 4 Detection of characteristic biological Possibilities © Hi. Tec Zang Gmb. H - HRE 4 Detection of characteristic biological phenomena CTR development:

Possibilities 4 Recognition of suitable conditions for conventional mass screening (operation duration, culture media, Possibilities 4 Recognition of suitable conditions for conventional mass screening (operation duration, culture media, operation conditions …) 4 Optimisation of substrate concentrations and reduction of media development time 4 Growth control under sterile conditions 4 Targeted sampling depending on oxygen transfer rate 4 Quality control © Hi. Tec Zang Gmb. H - HRE 4 Fermentation balancing (cytotoxycity- and proliferation assays)

State of the Art online-exhaust gas analytik ? stirred bioreactor shaking bioreactor © Hi. State of the Art online-exhaust gas analytik ? stirred bioreactor shaking bioreactor © Hi. Tec Zang Gmb. H - HRE 9 online OTR CTR RQ

Motivation „The disadvantage of the shake flask as an experimental system is that the Motivation „The disadvantage of the shake flask as an experimental system is that the experimenter has only limited capabilities for on-line monitoring and control. “ „Weakness of small-scale liquid fermentations: discontinuous monitoring“ Hilton, 1999 © Hi. Tec Zang Gmb. H - HRE Payne et al. , 1990

What kind of Online Signal? carbon source (glutamine, glucose, . . . ) trace What kind of Online Signal? carbon source (glutamine, glucose, . . . ) trace elements, vitamins nitrogen source (ammonia sulfate, urea, yeast extract, peptone, . . . ) sulfate source (sulfate, cysteine, . . . ) product (proteins, alcohol amino acids, . . . ) Oxygen Carbon dioxide © Hi. Tec Zang Gmb. H - HRE phosphorus source (phosphate, phytin)

Unknown Fermentation Process normal shaking flask: culture process A ? Time end of experiment Unknown Fermentation Process normal shaking flask: culture process A ? Time end of experiment © Hi. Tec Zang Gmb. H - HRE B

Known Fermentation Process culture process A B B Time end of experiment © Hi. Known Fermentation Process culture process A B B Time end of experiment © Hi. Tec Zang Gmb. H - HRE A

Solution measures online the respiration activities (OTR, CTR, RQ) of aerobic biological systems in Solution measures online the respiration activities (OTR, CTR, RQ) of aerobic biological systems in shaking flasks under © Hi. Tec Zang Gmb. H - HRE sterile conditions

Distinct Advantages 4 more information about microbiological processes in shaking flasks 4 rapid characterisation Distinct Advantages 4 more information about microbiological processes in shaking flasks 4 rapid characterisation and targeted optimisation of media 4 replaces expensive experiments in the fermenter 4 parallel technology (time, comparability. . . ) 4 casily handling 4 creates optimal repoducabilty options 4 virtual non-stop operation by very short set-up time 4 reduction of experimental time to the actually required time 4 distinction of process-related and biological effects © Hi. Tec Zang Gmb. H - HRE 4 visualising the perfect inoculation point

© Hi. Tec Zang Gmb. H - HRE Graduated flask © Hi. Tec Zang Gmb. H - HRE Graduated flask

Sample Fermentations Determination of the optimal inoculation- and fed -batch starting time (50 ml Sample Fermentations Determination of the optimal inoculation- and fed -batch starting time (50 ml liquid volume) Cell density [N/m. L] OTR CTR cell density glutamine- and glucose consumption 0 50 glucose consumption 100 150 200 Time of Fermentation [h] © Hi. Tec Zang Gmb. H - HRE OTR/CTR [mol/(L·h)] Mammalian cell culture Hybridoma

Sample Fermentations Media optimisation Example: optimum of osmolarity (50 ml liquid volume) optimum of Sample Fermentations Media optimisation Example: optimum of osmolarity (50 ml liquid volume) optimum of osmolarity at 0, 318 osmol/kg Osmolarity [osmol/kg] © Hi. Tec Zang Gmb. H - HRE Growth rate µ [h-1] Mammalian cell culture Hybridoma

Sample Fermentations Comparison of RAMOS to a stirred reactor with online exhaust gas analytics Sample Fermentations Comparison of RAMOS to a stirred reactor with online exhaust gas analytics OTR [mol/(L·h)] Mammalian cell culture Hybridoma Dipl. -Ing. M. Canzoneri stirred tank reactor (2 litre culture volume) 0 20 40 60 80 Time of Fermentation [h] © Hi. Tec Zang Gmb. H - HRE RAMOS (0, 05 litre culture volume)

Sample Fermentations Effect of different liquid volumes oxygen limitation Flask 1 : 10 m. Sample Fermentations Effect of different liquid volumes oxygen limitation Flask 1 : 10 m. L Flask 2 : 15 m. L Flask 3 : 20 m. L Flask 4 : 30 m. L Flask 5 : 40 m. L Flask 6 : 50 m. L Time of Fermentation [h] © Hi. Tec Zang Gmb. H - HRE OTR [mol/(L·h)] Bacterium Corynebacterium glutamicum

Sample Fermentations Effect of different substrate concentrations OTR [mol/(L·h)] Bacterium Pseudomonas fluorescens fermentation time Sample Fermentations Effect of different substrate concentrations OTR [mol/(L·h)] Bacterium Pseudomonas fluorescens fermentation time [h] © Hi. Tec Zang Gmb. H - HRE 1 x concentrated 2 x concentrated 4 x concentrated

Sample Fermentations Media- and process optimisation OTR [mol/(L·h)] Yeast Hansenula polymorpha Media with 100% Sample Fermentations Media- and process optimisation OTR [mol/(L·h)] Yeast Hansenula polymorpha Media with 100% comp. 1, 30 ml liquid Media with 200% comp. 1, 30 ml liquid Time of Fermentation [h] © Hi. Tec Zang Gmb. H - HRE Media with 200% comp. 1, 20 ml liquid

Sample Fermentations Cell-growth within a RAMOS experiment 23 Dipl. -Ing. M. Canzoneri © Hi. Sample Fermentations Cell-growth within a RAMOS experiment 23 Dipl. -Ing. M. Canzoneri © Hi. Tec Zang Gmb. H - HRE Mammalian cell cultures Hybridoma

Sample Fermentations Cell proliferation within a RAMOS experiment Mammalian cell culture Hybridoma 8 -time Sample Fermentations Cell proliferation within a RAMOS experiment Mammalian cell culture Hybridoma 8 -time parallel measurement 0 40 80 120 160 Time of Fermentation [h] © Hi. Tec Zang Gmb. H - HRE Cell density [N/ml] Dipl. -Ing. M. Canzoneri

Easy Handling 4 little required space – RAMOS fits to normal bench top 4 Easy Handling 4 little required space – RAMOS fits to normal bench top 4 virtual non-stop operation by very short set-up time 4 easy and fast-learnable appliance © Hi. Tec Zang Gmb. H - HRE 4 fully automated user software

© Hi. Tec Zang Gmb. H - HRE Operating Interface © Hi. Tec Zang Gmb. H - HRE Operating Interface

© Hi. Tec Zang Gmb. H - HRE Flask Overview © Hi. Tec Zang Gmb. H - HRE Flask Overview

© Hi. Tec Zang Gmb. H - HRE Oxygen Transfer Rate (OTR) © Hi. Tec Zang Gmb. H - HRE Oxygen Transfer Rate (OTR)

Detail View for each Flask © Hi. Tec Zang Gmb. H - HRE (OTR, Detail View for each Flask © Hi. Tec Zang Gmb. H - HRE (OTR, CTR, RQ)

O 2 -, CO 2 - Transfer Oxygen transfer (OT) Carbon dioxide transfer (CT) O 2 -, CO 2 - Transfer Oxygen transfer (OT) Carbon dioxide transfer (CT) © Hi. Tec Zang Gmb. H - HRE 4 Balancing of the total oxygen transfer during the fermentation process

maximum Growth Rate µ growth rate µ © Hi. Tec Zang Gmb. H - maximum Growth Rate µ growth rate µ © Hi. Tec Zang Gmb. H - HRE maximum growth rate µ

Shedding light on your process 32 CTR © Hi. Tec Zang Gmb. H - Shedding light on your process 32 CTR © Hi. Tec Zang Gmb. H - HRE OTR

OTR [mol/(L·h)] Economic efficiency consideration Media with 100% comp. 1, 30 ml liquid Media OTR [mol/(L·h)] Economic efficiency consideration Media with 100% comp. 1, 30 ml liquid Media with 200% comp. 1, 20 ml liquid 4 The variation of the media concentration led to an reduction of the time of fermentation of ca. 37 % 4 Time of amortisation: ca. 6 months © Hi. Tec Zang Gmb. H - HRE Time of Fermentation [h]

Cell culture (Hybridoma) © Hi. Tec Zang Gmb. H - HRE 34 • Dosing Cell culture (Hybridoma) © Hi. Tec Zang Gmb. H - HRE 34 • Dosing

FTT® Fluid-Train System © Hi. Tec Zang Gmb. H - HRE 35 • Dosing FTT® Fluid-Train System © Hi. Tec Zang Gmb. H - HRE 35 • Dosing and automated samplin

FTT® Fluid-Train System © Hi. Tec Zang Gmb. H - HRE 36 • controlled FTT® Fluid-Train System © Hi. Tec Zang Gmb. H - HRE 36 • controlled loop dosing

RQFeed™ - Feeding algorithm 37 determination of RQ by OUR, CER online measurement exact RQFeed™ - Feeding algorithm 37 determination of RQ by OUR, CER online measurement exact feeding of cultures significant increase in production rates shortening of the fermentation periods © Hi. Tec Zang Gmb. H - HRE ● ●

Cell. Drum™ - Cell force measurement reproducable biomechanical measurement personalised drug and toxin research Cell. Drum™ - Cell force measurement reproducable biomechanical measurement personalised drug and toxin research alternative to animal experiments integrated, fully automated and heat sterilisable pipetting unit ● 24 - 96 Multiwell units with integrated sensorics © Hi. Tec Zang Gmb. H - HRE 38 ● ●

Hi. Sense™ - Precision Gas Analysis 39 1 to 8(5) Measurement Channels for 1 Hi. Sense™ - Precision Gas Analysis 39 1 to 8(5) Measurement Channels for 1 to 4 Fermenters High Resolution Measurement Humidity Compensation (-c Version) "True" OUR, CER and RQ Measurements (-c Version) Low Interference Possible Overpressure Wear-resistant Sensor System Compact Design Additional Functions can be integrated Optionally free Programmability Numerous Coupling Options Data Export is possible © Hi. Tec Zang Gmb. H - HRE ● ● ●

Cell culture (Hybridoma) © Hi. Tec Zang Gmb. H - HRE 40 • Without Cell culture (Hybridoma) © Hi. Tec Zang Gmb. H - HRE 40 • Without dosing

Cell culture (Hybridoma) © Hi. Tec Zang Gmb. H - HRE 41 • Dosing Cell culture (Hybridoma) © Hi. Tec Zang Gmb. H - HRE 41 • Dosing according to OTR controlled loop starting at RQ<1

Cell culture (Hybridoma) © Hi. Tec Zang Gmb. H - HRE 42 • Dosing Cell culture (Hybridoma) © Hi. Tec Zang Gmb. H - HRE 42 • Dosing program

Cell culture (Hybridoma) © Hi. Tec Zang Gmb. H - HRE 43 • Parameterisation Cell culture (Hybridoma) © Hi. Tec Zang Gmb. H - HRE 43 • Parameterisation of taking samples

Cooperations and Publications Cooperations: Prof. Dr. Manfred Biselli Aachen University of Applied Science, Division Cooperations and Publications Cooperations: Prof. Dr. Manfred Biselli Aachen University of Applied Science, Division Jülich Faculty of Biotechnology Prof. Dr. -Ing. Jochen Büchs RWTH Aachen University, Faculty of Bioprocess Engineering Publications: Stöckmann Ch. , Maier U. , Anderlei T. , Knocke Ch. , Gellissen G. , Büchs J. , The Oxygen Transfer Rate as Key Parameter for the Characterisation of Hansenula polymorpha Screening Cultures, J. Ind. Microbiol. Biotechnol. 30, 613 -622, 2003 Anderlei T. , Zang W. , Büchs J. , Online respiration activity measurement (OTR, CTR, RQ) in shake flasks, Biochem. Eng. J. 17(3), 187 -194, 2004 Lotter St. , Büchs J. Utilization of power input measurements for optimisation of culture conditions in shaking flasks, Biochem. Eng. J. 17(3), 195 -204, 2004 Losen M. , Lingen B. , Pohl M. , Büchs. J. , Effect of oxygen-limitation and medium composition on Escherichia coli in small-scale cultures, Biotechnol. Progress. (accepted) © Hi. Tec Zang Gmb. H - HRE Anderlei T. , Büchs J. , Device for sterile online measurement of the oxygen transfer rate in shaking flasks, Biochem. Eng. J. 7(2), 157 -162, 2001