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Microorganisms are very tiny creatures that play an important part in the biotechnology industry. In certain environmental conditions, microbes metabolize and convert carbohydrates into alcohol or organic acids, a process known as fermentation. Different factors, such as temperature, air composition, nutrients and pH affects microbial growth. The purpose of this thesis was to study oxygen transfer behaviour and its influence on fermentation in a baffleless flask -fermentor-hybrid stirred-tank reactor (STR) (US patented Pat. 8,162,295) of Moubio Knowledge Co. The innovative design and portability of this tank make it a potentially important piece of laboratory equipment. The off-center impeller, the plastic vent tube and the thermometer well work together to create a possible mechanism for continuous gas exchange between the tank and the surrounding air. The tank (kLa = 29.57 h-1, 925 rpm) had 3 times oxygen transfer of Zoro’s [1] reactor (kLa = 10 h-1, 1000 rpm). In addition, this stirred tank reactor could grow microbes even at room temperature with 64 % of the same results as a shake flask incubating at an optimal temperature of 37 oC. In this project, Escherichia coli BL21 was used to produce green fluorescent protein (GFP) in the aerobic system. Oxygen transfer rate (OTR) was observed by iodometric titration, which was then used to calculate kLa values to form kLa correlations. OTR values changed based on the stirring speed, the depth of the plastic vent tube, and the depth of the thermometer well in the working volume. kLa correlation curves can be utilized in future tank design and gas-exchange research., Mikrobin kokoiset organismit ovat tärkeä osa biotekniikan teollisuudessa. Tietyissä ympäristön olosuhteissa mikrobit metaboloivat ja muuttavat hiilihydraatit alkoholeiksi ja orgaanisiksi hapoiksi, mitä kutsutaan fermentoinniksi. Erilaiset tekijät kuten lämpötila, ilma, ravinteet ja pH vaikuttavat mikrobikasvatukseen. Tutkimuksen tarkoituksena oli tarkastella hapen siirron käyttäytymistä ja sen vaikutusta fermentointiin Moubio Knowledge Co. omistamassa hybridi STR bioreaktorissa (US patentoitu 8,162,295). Reaktorin innovatiivinen muotoilu ja kannettavat ominaisuudet ovat potentiaalisia osia uudenaikaiseen laboratoriovälineistöön. Epäkeskinen sekoitin, muoviputki ja metallinen lämpömittariputki muodostavat yhdessä mahdollisen jatkuvan ilmanvaihtomekanismin tankin ja ympäristön välillä. Tankilla (kLa = 29.57 h-1, 925 rpm) oli kolme kertaa suurempi hapen siirto kuin Zoron [1] reaktorissa (kLa = 10 h-1, 1000 rpm). Lisäksi STR pystyi kasvattamaan mikrobit huoneenlämmössä jopa 64 % kasvatuspullon 37 oC:ssa tapahtuvaan optimaalisen lämpötilakasvatukseen verrattuna. Tässä projektissa käytettiin Escherichia coli BL21 kantaa tuottamaan GFP proteiinia aerobisessa systeemissä. Lisäksi, jodimetristä titrausta käytettiin tunnistamaan hapen siirtonopeutta, jonka avulla laskettiin kLa arvoja. Hapen siirtonopeusarvot vaihtelivat eri sekoitusnopeudesta sekä muovi- ja lämpömittariputken sijainneista riippuen. Sen jälkeen huomioimalla väliaineen tilavuutta, sekoitusvoimaa ja yleistä kLa korrelaatiota muodostettiin omat kLa korrelaatiot, joita voidaan hyödyntää tulevien uusien tankkien suunnitelussa ja ilmanvaihtotutkimuksissa.

BACKGROUND: Many reports have suggested that mild hypothermic culture conditions improve the specific monoclonal antibody (mAb) productivity of mammalian cells. The effect of active hypothermic growth on the mAb productivity of the hybridoma C2E7 was investigated. Hybridoma growth under hypothermic conditions (32 °C) was stimulated by supplementation of the culture medium with high serum concentrations (up to 30%). RESULTS: Specific and volumetric mAb productivity of a stimulated, active growth, mildly hypothermic hybridoma culture (30% FBS supplemented, 32 °C) were 1.38‐ and 1.34‐fold greater than the control culture (10% FBS supplemented, 37 °C). The enhanced specific mAb productivity under hypothermic conditions was associated with an increase in IgM mRNA levels during both the lag and early exponential phases of hypothermic growth. CONCLUSION: Stimulation of hybridoma growth under mildly hypothermic conditions increased both the specific and volumetric mAb productivity of hybridoma cells.

BACKGROUND: Many reports have suggested that mild hypothermic culture conditions improve the specific monoclonal antibody (mAb) productivity of mammalian cells. The effect of active hypothermic growth on the mAb productivity of the hybridoma C2E7 was investigated. Hybridoma growth under hypothermic conditions (32 °C) was stimulated by supplementation of the culture medium with high serum concentrations (up to 30%). RESULTS: Specific and volumetric mAb productivity of a stimulated, active growth, mildly hypothermic hybridoma culture (30% FBS supplemented, 32 °C) were 1.38‐ and 1.34‐fold greater than the control culture (10% FBS supplemented, 37 °C). The enhanced specific mAb productivity under hypothermic conditions was associated with an increase in IgM mRNA levels during both the lag and early exponential phases of hypothermic growth. CONCLUSION: Stimulation of hybridoma growth under mildly hypothermic conditions increased both the specific and volumetric mAb productivity of hybridoma cells.

The effect of mild hypothermic (32 degrees C) conditions on cell growth, cell-cycle progress, and antibody production of hybridoma C2E7 cells was investigated in the present study. The growth of hybridoma cells was slower during the mild hypothermic condition compared to that at 37 degrees C; this led to about 10% decrease in maximum viable cell density and volumetric antibody productivity. However, under mild hypothermic growth conditions, the culture viability was substantially improved and the specific antibody productivity was enhanced compared to that at 37 degrees C. The average specific productivity for the entire batch culture at 32 degrees C is about 5% higher than that at 37 degrees C. Cell-cycle analysis data showed that there was no growth arrestment during the mild hypothermic growth of hybridoma cells. The G1-phase cells were increased, while the S-phase cells were decreased gradually as the culture time progressed. Further analysis showed that the specific antibody productivity of hybridoma cells was correlated to the fraction of S-phase cells.

The effect of mild hypothermic (32 degrees C) conditions on cell growth, cell-cycle progress, and antibody production of hybridoma C2E7 cells was investigated in the present study. The growth of hybridoma cells was slower during the mild hypothermic condition compared to that at 37 degrees C; this led to about 10% decrease in maximum viable cell density and volumetric antibody productivity. However, under mild hypothermic growth conditions, the culture viability was substantially improved and the specific antibody productivity was enhanced compared to that at 37 degrees C. The average specific productivity for the entire batch culture at 32 degrees C is about 5% higher than that at 37 degrees C. Cell-cycle analysis data showed that there was no growth arrestment during the mild hypothermic growth of hybridoma cells. The G1-phase cells were increased, while the S-phase cells were decreased gradually as the culture time progressed. Further analysis showed that the specific antibody productivity of hybridoma cells was correlated to the fraction of S-phase cells.