Perfil epidemiológico das uroculturas positivas no Hospital Universitário durante janeiro-dezembro de 2001.

Trabalho de Conclusão de Curso - Universidade Federal de Santa Catarina, Centro de Ciências da Saúde, Departamento de Saúde Pública, Curso de Medicina, Florianópolis, 200

Appraisal of the mixing performance of Single-Use shaken bioreactors

The pharmaceutical industry is at the forefront of the production of antibodies using mammalian cell-based cultures, with single-use technologies gaining prominence in the manufacturing process. At laboratory scale mammalian cells are usually grown in low shear devices, with disposable shaken bioreactors being largely employed in the early stages of bioprocess development. It has been a recent industry trend to use shaken bioreactors at large scale in the upstream process, performing cell culture in single-use bags which eliminate the need for cleaning in place, offer flexibility and lower production down-times. Single-use Orbitally Shaken Bioreactors (OSRs) consist of a shaker, a structural support to which a disposable bag conforms to and all ancillary connections and controllers. Production scale OSRs with single-use bags, employ the agitation principle of shaken flasks and microwell plates, providing a homogeneously single-use upstream scale-up process thus facilitating scaling-up and simplifying regulatory approval. The aim of the work is to characterize the mixing and flow dynamics in a cylindrical orbitally shaken bioreactor with conical bottoms of different heights. The rationale for a conical bottom is to ease the suspension of cells or microcarriers, which are used for the cultivation of cells not yet adapted for suspend culture, such as legacy cell lines and stem cells. The geometry of the conical bottom was designed so to be compatible with single-use bags, as the conical bottom is truncated and the cone obtuse. This study builds upon previous works of the research group (Weheliye et al 2013, Rodriguez et al. 2013, Rodriguez et .al. 2014, Ducci and Weheliye, 2014) for flat bottom reactors, where increases in Froude number were found to determine a mean flow transition and to increase the turbulence levels. The major objective of the current work is to determine the performances of shaken bioreactors with conical bottom, and to assess to what extent the mean flow and flow regime transitions already identified for a flat bottom are affected by the geometry variation. Particle Image Velocimetry, PIV, and Dual Indicator System for Mixing Time, DISMT, were employed to assess the mixing performances in the bioreactor with a conical bottom. DISMT (see Rodriguez et al., 2014) consists in a colorimetric method where two pH indicators are used to visualise the level of mixing reached after insertion of an acid solution. The findings of the current study provides insight into the flow of a single-use shaken bioreactor and offers a novel approach to design the next generation of products and improve scaling methodologies

Automation of a cell culture process with lager vessels

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Orbital shaken bioreactor for influenza A virus production in high cell density cultivations

The majority of large scale cell culture processes is performed in stirred tank bioreactors (STR) and process development employs scale down models of the same bioreactor type. However, for screening of clones or process conditions at even smaller scale, shake flasks (SF) represent the most widely used model. Occasionally, SF allow for robust processes that cannot be transferred to STR because sensitive cells cannot cope with mechanical stress in STR due to stirring and aeration. Orbital shaken bioreactors (OSB) are a valuable alternative to STR as the transfer from SF to OSB is simplified because the systems rely on the same basic principles for mixing and aeration (e.g., bubble-free surface gassing). In particular, high oxygen transfer rates and short mixing times combined with low shear stress can also be achieved in OSB. These benefits may be even more pronounced in high density culture processes. Please click Additional Files below to see the full abstract

Producing viruses in orbit: Current developments for orbital shaken viral vaccine manufacturing

Preculture of suspension cells is successfully performed in shake flasks. Especially newly developed designer cells are passaged up to 100 times in shake flaks at high shaking frequency and are then perfectly adapted to growth in a CO2 incubator with pH control and maximum oxygen supply (typically above 80% pO2). When they are subsequently transferred to stirred tank bioreactors for scaling up, specific cell growth rates are often lower and cells become sensitive to pH control via acid/base addition and shear stress due to submers gassing (bubbles). This was also seen for avian AGE1.CR.pIX and human HEK 293 cells. To avoid these problems, scale up in shaken mode was evaluated. Here we present the latest developments of the SB10-X OSB bioreactor with regard to bag design and improvement of the control unit. A new control strategy was introduced leading to a faster and more precise pH and DO control. Furthermore, the perfusion bag was optimized, so that on TFF or two ATF systems can be easily connected. Both developments have led to a more robust SB10-X system that allows to easily perform batch, fed batch or perfusion runs. Please click Download on the upper right corner to see the full abstract

Scale-down of an orbital shaken bioreactor: High cell density cultivation in perfusion mode and virus production

Application of single-use bioreactors has been commonly shown for several cell culture-based production systems including commercial vaccine production. Compared to stainless steel bioreactors, competitive cell growth characteristics as well as virus yields can be reached [1]. In addition to conventional stirred tank reactors (STR), wave bioreactors or orbital shaken bioreactors (OSBs) are available that rely on alternative mixing regimes. For small-scale screening of clones and media, cell maintenance and process optimization, OSBs are the most widely used system. Besides their simple design and ease of handling, OSBs allow for robust processes due to reduced mechanical stress caused by stirring and aeration [2]. Furthermore, scale-up (£ 2500 L) is simplified as larger OSBs rely on the same basic principles for mixing and aeration (e.g. bubble-free surface gassing). Particularly for high cell density (HCD) processes, high oxygen transfer rates, short mixing times, and low shear stress are beneficial. Until now, the step from spin tubes or shake flasks into larger OSBs was rather large, as only the OSB SB10-X (Kühner AG, Switzerland) with a minimum working volume (wv) of 4-5 L was available. In this study, a novel scale-down 3 L vessel module (wv = 1-3 L) for the OSB SB10-X was evaluated for cultivation of suspension BHK-21 cells (CEVA, Germany) in perfusion mode to HCD. Cultivation was carried out in serum-free medium in a 3 L and 10 L single-use standard bag with 3 L and 5 L initial wv and 100 and 70 rpm shaking frequency with a shaking diameter of 50 mm, respectively. For perfusion, an alternating tangential flow system (ATF2, Repligen) with a cut-off of 0.4 µm (SB10-X) and 0.5 µm (SB3-X), respectively, was used. Following an initial batch phase of 2-3 days, perfusion was initiated. After a complete media exchange, cells in the 3 L vessel module were infected with a fusogenic oncolytic virus (rVSV-NDV, recombinant vesicular stomatitis virus-Newcastle disease virus) at a cell concentration of 44.5x106 cells/mL at a multiplicity of infection (MOI) of 10-4. The obtained data were compared to a cultivation of BHK-21 cells in the standard SB10-X module (infection at a cell concentration of 12.5x106 cells/mL with yellow fever virus WHO 17D-213/77 with an MOI of 10-3) and to a cultivation in a 1 L STR. The novel 3 L vessel module allowed for a successful and direct scale-down utilizing the SB10-X backbone without the need for further optimization. For both the SB10-X and the 3 L vessel module, the ATF system was successfully coupled and cell concentrations of 32.7x106 cells/mL and 45.9x106 cells/mL were reached with high viabilities above 98%, respectively. A faster doubling time (tD=22 h) was observed in the 3 L vessel module compared to the SB10-X system (tD=27 h). For rVSV-NDV production, similar infectious virus titers were reached compared to perfusion cultivations of BHK-21 cells in a 1 L STR. Volumetric media consumption was significantly reduced in the 3 L vessel module, facilitating the implementation of OSB systems in non-industrial research environments. All in all, we demonstrated the adaptability and scalability of the single-use OSB system for the production of various viruses in HCD perfusion mode. References [1] Gallo-Ramirez, L. E., A. Nikolay, Y. Genzel, and U. Reichl. 2015. Bioreactor concepts for cell culture-based viral vaccine production. Expert Rev Vaccines 14 (9):1181-95. doi: 10.1586/14760584.2015.1067144 [2] Klöckner W, Diederichs S, Büchs J. Orbitally shaken single-use bioreactors. Adv Biochem Eng Biotechnol. 2014;138:45-60. doi: 10.1007/10_2013_188. PMID: 23604207

Разработка игрового приложения с искусственным интеллектом в рамках языковой модели

Объект исследования: алгоритмы построения ассоциаций внутри языковой модели и их практическое применение. Цель работы: разработка игрового приложения "Кодовые имена" с возможностью замены реальных игроков искусственным интеллектом. Область применения: распространение с помощью сервиса Google Play Market. Результаты работы: эффективный алгоритм создания базы ассоциаций, алгоритмы искусственного интеллекта для игры "Кодовые имена", мобильное игровое приложение.Object of research: algorithms for constructing associations within a language model and their practical application. The purpose of the work: the development of the game application "Codenames" with the possibility of replacing real players with artificial intelligence. Scope: distribution using Google Play Market. The results of work: an effective algorithm for creation of association database, algorithms for artificial intelligence for the game "Codenames", a mobile game application

On the measurement and scaling of mixing time in orbitally shaken bioreactors

Accurate determination of the mixing time in orbitally shaken bioreactors (OSRs) is essential for the optimization of mixing processes and minimization of concentration gradients that can be deleterious to cell cultures. The Dual Indicator System for Mixing Time (DISMT) was employed to measure mixing times in cylindrical and Erlenmeyer flask bioreactors. Various aspects of importance for the acquisition of accurate data from the measurement methodology are discussed, utilizing also comparisons of DISMT and pH probe results obtained in two stirred reactors. The OSR results are juxtaposed with data previously reported in the literature for both cylindrical reactors and Erlenmeyer flasks. The employment of a critical Froude number shows promise for the establishment of a scaling law for mixing time across the various types and sizes of shaken bioreactors