共查询到20条相似文献，搜索用时 0 毫秒
1.
Vallejos JR Kostov Y Ram A French JA Marten MR Rao G 《Biotechnology and bioengineering》2006,93(5):906-911
A novel optical sensor was used to study mixing and mean circulation time in a model minibioreactor (12.5 mL stirred vessel, equipped with a paddle impeller). Rotational rates in the range of 10-1,000 rpm corresponding to Reynolds number between 14 and 1,350 were studied. Results suggest that depending on the impeller rotational speed, mixing times up to 214 +/- 87 s can be reproducibly achieved. The minibioreactor was operated in the transitional regime, and it was determined that the non-dimensional form for mixing time, NTheta(M) was linearly dependent on Reynolds number. A linear correlation between mean circulation time and the inverse of rotational speed was also determined. The mean circulation time dependence on rotational speed in the 12.5 mL stirred vessel is similar to those found in large-scale stirred vessels. These results suggest that mixing and circulation times found in large-scale reactors can be replicated in minibioreactors. 相似文献
2.
3.
4.
5.
David Watts Jochen Müller‐Dieckmann Gohar Tsakanova Victor S. Lamzin Matthew R. Groves 《Acta Crystallographica. Section D, Structural Biology》2010,66(8):901-908
Modern X‐ray structure analysis and advances in high‐throughput robotics have allowed a significant increase in the number of conditions screened for a given sample volume. An efficient evaluation of the increased amount of crystallization trials in order to identify successful experiments is now urgently required. A novel approach is presented for the visualization of crystallization experiments using fluorescence from trace amounts of a nonspecific dye. The fluorescence images obtained strongly contrast protein crystals against other phenomena, such as precipitation and phase separation. Novel software has been developed to quantitatively evaluate the crystallization outcome based on a biophysical metric correlated with voxel protein concentration. In >1500 trials, 85.6% of the successful crystallization experiments were correctly identified, yielding a 50% reduction in the number of `missed hits' compared with current automated approaches. The use of the method in the crystallization of three previously uncharacterized proteins from the malarial parasite Plasmodium falciparum is further demonstrated. 相似文献
6.
The present paper describes the development and validation of a simple and sensitive micelle‐enhanced high‐throughput fluorometric method for the determination of niclosamide (NIC) in 96‐microwell plates. The proposed method is based on the reduction of the nitro group of niclosamide to an amino group using Zn/HCl to give a highly fluorescent derivative that was developed simultaneously and measured at λem 444 nm after excitation at λex 275 nm. Tween‐80 and carboxymethylcellulose (CMC) have been used as fluorescence enhancers and greatly enhanced the fluorescence by factors of 100–150%. The different experimental conditions affecting the fluorescence reaction were carefully investigated and optimized. The proposed method showed good linearity (r2≥ 0.9997) over the concentration ranges of 1–5 and 0.5–5 μg/ml with lower detection limits of 0.01 and 0.008 μg/ml and lower quantification limits of 0.04 and 0.03 μg/ml on using Tween‐80 and or CMC, respectively. The developed high‐throughput method was successfully applied for the determination of niclosamide in both tablets and spiked plasma. The capability of the method for measuring microvolume samples made it convenient for handling a very large number of samples simultaneously. In addition, it is considered an environmentally friendly method with lower consumption of chemicals and solvents. 相似文献
7.
In this work an integrated robotic platform has been used for the development of a fully automated microscale process sequence comprising fermentation and bioconversion using E. coli TOP10 [pQR210] expressing cyclohexanone monooxygenase (CHMO). Ninety six-Deep Square Well (96-DSW) microtiter plates were used for microbial culture and enzyme-catalyzed conversion, where plate preparation, reagent addition, and sampling were all carried out without manual intervention. The adoption of automated robotic procedures has enabled the rapid collection of kinetic data for whole process optimization at the microscale. This high-throughput approach enabled a range of amino acid sources for media formulation and well fill volumes to be investigated highlighting when nutritional limitation and oxygen limitations took place. The automated process sequence has been applied to test six CHMO substrates including norcamphor and cycloheptanone all of which to the best of our knowledge have yet to be tested with E. coli TOP10 [pQR210]. Substrate specificity and product selectivity were effectively demonstrated and compared to both the natural substrate cyclohexanone and the model substrate bicyclo[3.2.0]hept-2-en-6-one used to demonstrate asymmetric synthesis. The results obtained using the developed process sequence could be reproduced at 75 L scale when a matched oxygen transfer coefficient k(L) a approach was used. The study demonstrates how automated microscale processing enables the rapid collection of kinetic quantitative data in a robust manner with clear implications for accelerating bioprocess development, optimization, and scale-up. 相似文献
8.
Russell A. Judge Kerry Swift Carlos Gonzlez 《Acta Crystallographica. Section D, Structural Biology》2005,61(1):60-66
Intrinsic ultraviolet fluorescence has been investigated as a rapid non‐invasive method for identifying and distinguishing protein crystals. An epi‐fluorescence microscope, which provides for excitation and viewing of fluorescence from above the sample, and a straight‐through geometry, which provides excitation from above and views fluorescence from underneath the sample, were tested with protein and non‐protein crystal samples. In both systems the protein crystals were observed to fluoresce brightly, providing a high contrast against background solution fluorescence, thus enabling protein crystals to be identified and distinguished from non‐protein crystals. 相似文献
9.
10.
Elizabeth Forsythe Aniruddha Achari Marc L. Pusey 《Acta Crystallographica. Section D, Structural Biology》2006,62(3):339-346
Covalent labeling of macromolecules with trace levels (<1%) of a fluorescent dye is proposed as a means to facilitate finding or detecting crystals in crystallization drops. To test the effects of labeled protein concentration on the resulting X‐ray diffraction data, experiments were carried out with the model proteins insulin, ribonuclease, lysozyme and thaumatin, which were labeled with the fluorescent dye carboxyrhodamine. All proteins were labeled on their N‐terminal amine and lysozyme was also labeled randomly on lysine side chains in a separate series of experiments. Ribonuclease and N‐terminal amine‐labeled lysozyme crystals were poorly formed at 10% label concentration and these were not used in subsequent diffraction experiments. All model proteins were tested to 5% labeled protein, and thaumatin and randomly labeled lysozyme gave well formed crystals to 10% labeled protein. In all cases tested, the presence of the label was found to not significantly affect the X‐ray diffraction data quality obtained. Qualitative visual‐inspection experiments over a range of label concentrations indicated that optimum derivatization levels ranged from 0.025–0.05% for insulin to 0.1–0.25% for thaumatin. Light intensity is a simpler search parameter than straight lines and by virtue of being the most densely packed phase, labeled crystals should be the most intense light sources under fluorescent illumination. For both visual and automated methods of crystal detection, label intensity is a simpler and potentially more powerful search parameter. Screening experiments using the proteins canavalin, β‐lactoglobulins A and B and chymotrypsinogen, all at 0.5% label concentration, demonstrated the utility of this approach to rapidly finding crystals, even when obscured by precipitate. The use of trace‐labeled protein is also proposed to be useful for the automated centering of crystals in X‐ray beamlines. 相似文献
11.
Harms P Kostov Y French JA Soliman M Anjanappa M Ram A Rao G 《Biotechnology and bioengineering》2006,93(1):6-13
Two prototype 24-unit microbioreactors are presented and reviewed for their relative merits. The first used a standard 24-well plate as the template, while the second consisted of 24-discrete units. Both systems used non-invasive optical sensors to monitor pH and dissolved oxygen. The systems were used to cultivate Escherichia coli. Both designs had their merits and the results obtained are presented. In addition, dissolved oxygen control was demonstrated at the milliliter scale and 24 simultaneously monitored fermentations were successfully carried out. These results demonstrated high quality high throughput bioprocessing and provide important insights into operational parameters at small scale. 相似文献
12.
This article describes a simple and potentially scalable microfiltration method for purification of recombinant proteins. This method is based on the fact that when an elastin-like polypeptide (ELP) is fused to a target protein, the inverse phase transition behavior of the ELP tag is imparted to the fusion protein. Triggering the phase transition of a solution of the ELP fusion protein by an increase in temperature, or isothermally by an increase in salt concentration, results in the formation of micron-sized aggregates of the ELP fusion protein. In this article, it is shown that these aggregates are efficiently retained by a microfiltration membrane, while contaminating E. coli proteins passed through the membrane upon washing. Upon reversing the phase transition by flow of Milli-Q water, soluble, pure, and functionally active protein is eluted from the membrane. Proof-of principle of this approach was demonstrated by purifying a fusion of thioredoxin with ELP (Trx-ELP) with greater than 95% recovery of protein and with greater than 95% purity (as estimated from SDS-PAGE gels). The simplicity of this method is demonstrated for laboratory scale purification by purifying Trx-ELP from cell lysate using a syringe and a disposable microfiltration cartridge. The potential scalability of this purification as an automated, continuous industrial-scale process is also demonstrated using a continuous stirred cell equipped with a microfiltration membrane. 相似文献
13.
Yusuke Mizokami Ko Noguchi Mikiko Kojima Hitoshi Sakakibara Ichiro Terashima 《Plant, cell & environment》2019,42(4):1257-1269
C3 photosynthesis is often limited by CO2 diffusivity or stomatal (gs) and mesophyll (gm) conductances. To characterize effects of stomatal closure induced by either high CO2 or abscisic acid (ABA) application on gm, we examined gs and gm in the wild type (Col‐0) and ost1 and slac1‐2 mutants of Arabidopsis thaliana grown at 390 or 780 μmol mol?1 CO2. Stomata of these mutants were reported to be insensitive to both high CO2 and ABA. When the ambient CO2 increased instantaneously, gm decreased in all these plants, whereas gs in ost1 and slac1‐2 was unchanged. Therefore, the decrease in gm in response to high CO2 occurred irrespective of the responses of gs. gm was mainly determined by the instantaneous CO2 concentration during the measurement and not markedly by the CO2 concentration during the growth. Exogenous application of ABA to Col‐0 caused the decrease in the intercellular CO2 concentration (Ci). With the decrease in Ci, gm did not increase but decreased, indicating that the response of gm to CO2 and that to ABA are differently regulated and that ABA content in the leaves plays an important role in the regulation of gm. 相似文献
14.
Chao Chen Yang Zhao Shuaicheng Lu Kanghua Li Yang Li Bo Yang Wenhao Chen Liang Wang Dengbing Li Hui Deng Fei Yi Jiang Tang 《Liver Transplantation》2017,7(20)
Sb2Se3, a V2‐VI3 compound semiconductor, has attracted extensive research attention in photovoltaics due to its non‐toxicity, low cost and earth‐abundant constituents. Herein, a combinatorial approach to optimize the performance of TiO2/Sb2Se3 thin film photovoltaics is employed. By simultaneously conducting a series of experiments in parallel rather than one after another, combinatorial strategy increases experimental throughput and reduces personnel costs. Key parameters such as TiO2 thickness, post‐annealing temperature and Sb2Se3 thickness are identified as 65 nm, 450 °C and 850 nm through the combinatorial approach. Finally, in combination with (NH4)2S back surface cleaning, TiO2/Sb2Se3 solar cells with 5.6% efficiency and decent stability are obtained, showcasing the power of high‐throughput strategy for accelerating the optimization of Sb2Se3 photovoltaics. 相似文献
15.
16.
Joseph Newton Reinhard Oeggl Nils H. Janzen Sandra Abad Daniela Reinisch 《Engineering in Life Science》2020,20(8):331-337
Miniaturization and automation have become increasingly popular in bioprocess development in recent years, enabling rapid high‐throughput screening and optimization of process conditions. In addition, advances in the bioprocessing industry have led to increasingly complex process designs, such as pH and temperature shifts, in microbial fed‐batch fermentations for optimal soluble protein expression in a range of hosts. However, in order to develop an accurate scale‐down model for bioprocess screening and optimization, small‐scale bioreactors must be able to accurately reproduce these complex process designs. Monitoring methods, such as fluorometric‐based pH sensors, provide elegant solutions for the miniaturization of bioreactors, however, previous research suggests that the intrinsic fluorescence of biomass alters the sigmoidal calibration curve of fluorometric pH sensors, leading to inaccurate pH control. In this article, we present results investigating the impact of biomass on the accuracy of a commercially available fluorometric pH sensor. Subsequently, we present our calibration methodology for more precise online measurement and provide recommendations for improved pH control in sophisticated fermentation processes. 相似文献
17.
The flow of cytometry into systems biology. 总被引：1，自引：0，他引：1
Biomedical research is evolving to address biological systems as molecular pathways integrated into complex networks. Tools for molecular and cell analysis are also evolving to address the new challenges and opportunities of this approach. Flow cytometry is a versatile analytical platform, capable of high speed quantitative measurements of cells and other particles. These capabilities are being exploited and extended in a range of new applications stemming from opportunities presented by the advances of genomics, proteomics and systems biology, which are in turn impacting clinical diagnosis, vaccine development and drug discovery. In this review, we highlight some of these advances and consider the future evolution of flow cytometry technology. 相似文献
18.
Robin Kempkes Elizabeth Stofko Kam Lam Edward H. Snell 《Acta Crystallographica. Section D, Structural Biology》2008,64(3):287-301
The Hauptman–Woodward Medical Research Institute runs a high‐throughput crystallization screening service in which macromolecules are screened against 1536 potential crystallization cocktails. Typically, multiple crystallization leads are identified. With a limited amount of sample, the question becomes `How many leads can be optimized and which leads are most likely to produce X‐ray diffraction data?'. In order to prioritize the hits for optimization, the amount of glycerol required to successfully cryocool each cocktail has been determined for the cocktails used in the high‐throughput screen. Those hit conditions that require the minimum amount of cryoprotectant for successful vitrification will be closer in chemical make‐up to the mother liquor. Hence, if the physical properties of the crystals are similar, one could logically prioritize leads that are more likely to produce diffraction based upon the chemical similarity of the native to the cryopreserved mother liquor. 相似文献
19.
20.
Two‐dimensional fluorescence as soft sensor in the monitoring of biotransformation performed by yeast 
下载免费PDF全文
下载免费PDF全文 Marcin Zabadaj Karolina Chreptowicz Jolanta Mierzejewska Patrycja Ciosek 《Biotechnology progress》2017,33(2):299-307
Soft sensors are powerful tools for bioprocess monitoring due to their ability to perform online, noninvasive measurement, and possibility of detection of multiple components in cultivation media, which in turn can provide tools for the quantification of more than one metabolite/substrate/product in real time. In this work, soft sensor based on excitation‐emission fluorescence is for the first time applied for the monitoring of biotransformation production of 2‐phenylethanol (2‐PE) by yeast strains. Main process parameters—such as optical density, glucose, and 2‐PE concentrations—were determined with high accuracy and precision by fluorescence fingerprinting coupled with partial least squares regression. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:299–307, 2017 相似文献