Media replenishment: A tool for the analysis of high-cell density perfusion systems

A simple method to analyze media consumption and by-product build-up in high-cell-density perfusion cultures has been developed. This technique makes use of media replenishment strategy which can be used to decouple the two phenomena. Also, the replenished media method can be used to identify limitation by complex and undefined media components such as bovine serum.     

Diethylpyrocarbonate—An effective agent for the sterilization of different types of nutrient media

A simple and convenient method has been tested for the steriltzation of nutrient media for long-term cultivation of plant cells. Diethylpyrocarbonate is suitable for this task in concentrations about 1000 mg l^-1 The cells cultivated for 15 subsequent passages on media treated by DPC had the same growth parameters, production pattern and ability to transform exogeneous organic compounds as did the controls. The method is suitable for the preparatian of both liquid and agar media, for stabilization of stock solutions and for sterilization of cultivation vessels and tubing.Abbreviations DPC diethylpyrocarbonate - medium MS nutrient medium according to Murashige and Skoog - NAA naphtaleneacetic acid     

Modular development enables rapid design of media for alternative hosts

Developing media to sustain cell growth and production is an essential and ongoing activity in bioprocess development. Modifications to media can often address host or product-specific challenges, such as low productivity or poor product quality. For other applications, systematic design of new media can facilitate the adoption of new industrially relevant alternative hosts. Despite manifold existing methods, common approaches for optimization often remain time and labor-intensive. We present here a novel approach to conventional media blending that leverages stable, simple, concentrated stock solutions to enable rapid improvement of measurable phenotypes of interest. We applied this modular methodology to generate high-performing media for two phenotypes of interest: biomass accumulation and heterologous protein production, using high-throughput, milliliter-scale batch fermentations of Pichia pastoris as a model system. In addition to these examples, we also created a flexible open-source package for modular blending automation on a low-cost liquid handling system to facilitate wide use of this method. Our modular blending method enables rapid, flexible media development, requiring minimal labor investment and prior knowledge of the host organism, and should enable developing improved media for other hosts and phenotypes of interest.     

The application of chromogenic media in clinical microbiology

Since 1990, a wide range of chromogenic culture media has been made commercially available providing useful tools for diagnostic clinical microbiology. By the inclusion of chromogenic enzyme substrates targeting microbial enzymes, such media are able to target pathogens with high specificity. Examples of target pathogens include Staphylococcus aureus, Streptococcus agalactiae, Salmonella spp. and Candida spp. The inclusion of multiple chromogenic substrates into culture media facilitates the differentiation of polymicrobial cultures, thus allowing for the development of improved media for diagnosis of urinary tract infections and media for the enhanced discrimination of yeasts. The purpose of this review is to provide some insight into how such media work and appraise their utility in routine clinical diagnostics, in comparison with conventional media.     

Direct measurement of water availability in gelled plant tissue culture media

Summary Water constitutes nearly 100% of the volume and 95% of the mass of gelled plant tissue culture media. Even so, plant growth and development responses observed to occur with relatively small changes in gelling agent concentration (0.1% of media total mass) have been attributed to changes in media water availability. Measurements with three alternative direct techniques, specific for measuring physiochemical water availability indicated the effects of a change of this magnitude in gelling agent concentration negligibly affected the media water potential and water conductivity. Sensitive pressure membrane measurements indicated that incremental gelling agent concentration increases of 0.1% (of media total mass) within the range normally used for plant tissue culture media, depressed water matric potential only 1–2cm H₂O (1–2×10⁻⁴ MPa (mega pascal.)); these values were confirmed with equally sensitive tensiometer measurements. Moreover, no effect of concentration on water movement could be detected with a precise constant-head permeameter over a broader range of gelling agent concentrations. These results indicate that eitherin vitro plants are extremely sensitive to subtle shifts in water status, or other physiochemical factors that also change with gelling agent concentration are contributing to the reported effects on plant growth and development.     

Determination of fumonisins and hydrolyzed fumonisin B1 in microbial culture media by LC/ESI-MS

In order to analyze liquid growth media for fumonisin B₁, B₂ and hydrolyzed fumonisin B1 (HFB1) after incubation with microorganisms for degradation studies, a liquid chromatography/electrospray ionisation mass spectrometry (LC/ESI-MS) method was developed. The use of phytosphingosine hydrochloride (PSH) and T-2-toxin-d3 (T2d3) as internal standard has been tested. The detection limit established with PSH as an internal standard was about 20 ng/mL for FB1 and FB2 and about 50 ng/mL for HFB1. The developed method allows the rapid, simultaneous and quantitative determination of these analytes in microbial culture media without any cleanup. Presented at the 25^th Mykotoxin Workshop in Giessen, Germany May 19–21, 2003     

微生物培养基质量控制技术和标准

微生物培养基的酸碱度、凝胶强度和选择性等直接影响到培养基的质量,在理化试验方法中采用连接可渗透陶器型液体接头的电极和平头电极或者连接微型探头的电极可分别测定液体和固体培养基的pH值,而采用Gelometer和the LFRA Texture Analyser可测定固体培养基的凝胶强度。在微生物学方法中固体培养基采用倾注平板法、涂布法、划线法（半定量法）、改良的Miles-Misra法等测定生长情况,液体培养基采用稀释法测定生长率,用目标菌和杂菌的混合菌株评价选择性增菌培养基的选择性,利用OD值评价液体培养基生长率等。ICFMH（国际食品微生物学和卫生学委员会培养基工作组）、ISO、FDA以及我国卫生部等相继制定了培养基质量控制的标准,但目前还没有一个系统的适合我国国情的培养基质量控制国家标准,以致各相关单位采用的标准不一致,所以制定培养基质量控制国家标准非常关键。     

Towards robust cell culture processes — Unraveling the impact of media preparation by spectroscopic online monitoring

Biopharmaceutical manufacturing processes can be affected by variability in cell culture media, e.g. caused by raw material impurities. Although efforts have been made in industry and academia to characterize cell culture media and raw materials with advanced analytics, the process of industrial cell culture media preparation itself has not been reported so far. Within this publication, we first compare mid‐infrared and two‐dimensional fluorescence spectroscopy with respect to their suitability as online monitoring tools during cell culture media preparation, followed by a thorough assessment of the impact of preparation parameters on media quality. Through the application of spectroscopic methods, we can show that media variability and its corresponding root cause can be detected online during the preparation process. This methodology is a powerful tool to avoid batch failure and is a valuable technology for media troubleshooting activities. Moreover, in a design of experiments approach, including additional liquid chromatography–mass spectrometry analytics, it is shown that variable preparation parameters such as temperature, power input and preparation time can have a strong impact on the physico‐chemical composition of the media. The effect on cell culture process performance and product quality in subsequent fed‐batch processes was also investigated. The presented results reveal the need for online spectroscopic methods during the preparation process and show that media variability can already be introduced by variation in media preparation parameters, with a potential impact on scale‐up to a commercial manufacturing process.     

抗消毒剂型微生物培养基研究

二氧化氯、过氧乙酸、次氯酸钠及过氧化氢等消毒剂高浓度可以杀灭微生物,低浓度可以抑制微生物生长。在微生物检测中,要消除消毒剂的干扰,培养基的抗消毒剂指数必须控制在１２．０—２４.５之间。消毒剂解抑剂Ⅰ的抗消毒剂指数为 １２．０,对细菌和真菌生长无明显抑制作用,加入经改良和优化的普通培养基后,制得５种抗消毒剂型培养基,在灭菌前后和一年的保存期内,其抗消毒剂指数基本保持不变。当采用大样倾注平板法和液体大样法检测残留消毒剂的样品时,使用抗消毒剂型培养基检出细菌和真菌能力大大高于普通培养基。     

Media for cultivation of animal cells: an overview

The increasing interest in products from animal cells has caused an extensive research effort towards development of media for cell cultivation.The basic components in the media used for cultivation of animal cells vary depending upon the characters of the cells and the cultivation method. Basic components consist of an energy source, nitrogen source, vitamins, fats and fatty soluble components, inorganic salts, nucleic acid precursors, antibiotics, oxygen, pH buffering systems, hormones, growth factors and serum. Extensive efforts are directed towards developing serum-free or chemically defined media. Among the serum substitutes is a long list of hormones and growth factors.     

混浊介质后向散射特性的Mueller矩阵实验测量

Mueller矩阵是一种公认的能很好地表述介质偏振特性的方法.由于散射光偏振在生物组织无创伤诊断技术等诸多领域中的重要应用价值,对组织散射特性的Mueller矩阵的研究成为国际上组织光学的热点之一.与现有测量Mueller矩阵的实验方法相比,斜入射正接收装置用来测量Mueuer矩阵是一个更加行之有效的方法,再结合一种新的算法来处理后续数据,由此所获得的Mueller矩阵空间分布图的清晰度不亚于其它文献的报道.这种测量方法结构更简单,具有测量更方便、准确等优点.结果表明:入射角影响Mueller矩阵的空间分布图.随着介质浓度的增大,随机介质后向散射Mueller矩阵各元素的空间分布图样减小.     

非水介质中辣根过氧化物酶（HRP）荧光活性测定法

建立了一种分析ＨＲＰ催化活力的新方法。该方法基于单体（底物）、聚合物（产物）的荧光发射光谱不重叠,使用荧光光谱仪,通过测量底物荧光淬灭来检测ＨＲＰ在非水介质中（二氧六环－水、乙醇－水、丙酮－水体系）催化酚类、芳香胺类物质聚合的活力。此方法迅速、简便,结果是定量并可重复的,并能定量地计算底物转化率。     

Serum-free media for cultures of primitive and mature hematopoietic cells

The in vitro culture of human hematopoietic cells has many research and therapeutic applications. Traditionally, human hematopoietic cultures have been conducted using serum-containing media. The disadvantages inherent in the use of serum could be eliminated by the use of serum-free media. In this review, we summarize and discuss the current status of serum-free media for both mature and immature human hematopoietic cells. The mature hematopoietic cells discussed are of lymphoid (e.g., lymphokine activated killer cells and tumor infiltrating lymphocytes) and myeloid origin (e.g., monocytes/macrophages). The cultures of immature hematopoietic cells discussed are clonogenic and long-term cultures. In addition, we briefly review the types of human hematopoietic cells, their clinical applications, and the basic strategies and components used to formulate serum-free media, Finally, we outline future requirements and directions in the development of serum-free media for primitive hematopoietic cells.     

Evaluation of a new chromogenic agar for the detection of Listeria in food

AIM: Rapid identification of Listeria in food is important in protecting consumers from infection. The development of chromogenic media such as agar Listeria according to Ottaviani and Agosti (ALOA) has allowed more rapid detection of Listeria monocytogenes, with presumptive identification of this pathogenic species after only 24 h of incubation. The aim of this study was to evaluate Oxoid chromogenic Listeria agar (OCLA) in comparison with ALOA and a traditional, nonchromogenic medium, Oxford agar. METHODS AND RESULTS: Media were compared using pure cultures, spiked food samples and naturally contaminated samples. Whilst development of typical colony morphology took 48 h on Oxford agar, Listeria spp. were frequently detected after 24 h of incubation on OCLA and ALOA. There was no significant difference in recovery between the two chromogenic media. CONCLUSIONS: Results indicate that OCLA gives equivalent recovery of Listeria spp. compared with ALOA. Whilst L. monocytogenes was frequently detected after 24 h of incubation, a 48-h incubation time was necessary to ensure detection of both L. monocytogenes and other Listeria spp. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has shown that a commercially available chromogenic medium other than ALOA is appropriate for use in the international standard method. The commercial availability of more than one medium will facilitate the more widespread use of the method, thus increasing confidence in the ability to detect L. monocytogenes in food in the presence of other Listeria spp.     

全媒体时代医院舆情管理危机的应对措施

随着科学技术的快速发展,各类新媒体的出现让部分医疗事件有了更高的关注度。近年来医疗纠纷日益严重,在全媒体时代的背景下,医院舆情危机频繁发生,对医院声誉和医患关系产生较大的不良影响。如何正确的引导网络舆情、降低医院舆情危机带来的不利影响成为急需解决的问题。本研究就全媒体时代医院舆情管理危机展开探讨,从医院舆情管理现状、医院舆情管理所存在的问题及相关的应对措施这三个方面入手,旨在为医院管理者在全媒体背景下控制医院舆情、化解医院突发事件、改善医患关系提供参考。     

The source of a movement: making the case for social media as an informational source using Black Lives Matter

Social media is growing as a technological and social phenomenon, with billions of people using it every day around the world. Given its increasing ubiquity, researchers continue to seek to understand social media and its implications for the social world. Part of the growing body of scholarship on social media includes uncovering the most effective methodologies for exploration that appropriately consider differences between the technology of social media and the ways in which people use them. New directions for social media research include the use of big data and analysing the socio-technical nature of social media. Using data from an ongoing study on college Millennials that includes questions about their thoughts on the Black Lives Matter movement, the author suggests another direction for future research: studying the implications of social media as an informational source.     

A novel method for the quantitative assessment of the percolation of <Emphasis Type="Italic">Metarhizium anisopliae</Emphasis> conidia through horticultural growing media

Incorporation of fungal biological control agents (BCAs) into plant growing media has considerable ergonomic and economic benefits for growers. These agents usually give prophylactic control of target pests and diseases. However, their efficacy is dose dependent and loss of inoculum through leaching could influence the degree of protection they provide. At present there are no protocols to determine the loss of inoculum in the disparate growing media used in horticulture. We describe a method based on a nutrient leaching column to quantify leaching of conidia of the insect-pathogenic fungus Metarhizium anisopliae in a range of growing media. Conidia of this biocontrol agent were applied as a drench or premixed into the medium. Both the application method and growth medium influenced conidial leaching. Inoculum losses were greater following drench application than premixing (95% vs. 15%) irrespective of media type. Comparatively more inoculum was lost from bark and coir following drench application whereas losses were relatively high in peat following premixed application. The leaching column assay provided a simple and accurate method to quantify inoculum loss in real time. This assay could help determine leaching of other fungal BCAs in growing media. It could help in improving pest and disease control by optimizing the rate and frequency of conidial application as well in the design of more efficacious formulations.     

Activities of Urease and Nickel Uptake of Helicobacter pylori Proteins are Media- and Host-Dependent

Background:  Nickel-dependent urease activity and nickel supply are essential for successful colonization of Helicobacter pylori in the acidic environment of the human stomach. A comparison of media effects on these two activities have never been carried out. Additionally to H. pylori we cultivated an Escherichia coli strain expressing the urease and the nickel transporter NixA of H. pylori on the same four media and measured in all cases urease and nickel uptake activity.
Aim:  To compare nickel uptake and urease activity on an inter- and intraspecies level.
Results:  In H. pylori nickel uptake (four to 200 times) and urease activities (400 to 30,000 times) were found to be much higher in comparison to the tested E. coli strain after growth on all media. These differences could not be explained by reduced protein amounts in the heterologous host E. coli . On which media the two bacteria extracted most of the nickel were organism-dependent: E. coli on Brucella Broth, H. pylori on Trypticase Soy Broth, and Minimal Media.
Conclusion:  H. pylori took nickel much more efficiently up than E. coli . The observed differences in urease activity are most likely due to additional protein components absent in the recombinant E. coli strain. The observed variety in nickel uptake and urease activities on the different media in the same organism depended on the intrinsic nickel content and chelating capacities of media components. Different culture conditions may lead to varying results; generalizations should be concluded only after excluding their media dependence.     

曼地亚红豆杉‘Hicksii’花粉活力检测条件优化和适宜诸藏温度分析

采用单因素实验设计对适宜于曼地亚红豆杉‘Hicksii’(Taxus media‘Hicksii')花粉活力检测的TTC染色法和离体培养法的实验条件进行了选择,并采用优化条件研究了在25℃～-196℃条件下储藏13周花粉活力的变化.结果表明:采用TTC染色法测定的花粉活力均高于离体培养法.在TTC染色法的3个影响因素(染液pH、TTC浓度和染色温度)中,染液pH对检测结果有极显著影响,而染色温度和TTC浓度则无显著影响,但温度对染色速率有影响.在离体培养法的3个影响因素(培养基中蔗糖添加量、H3BO3和Ca(NO3)2浓度及培养温度)中,蔗糖添加量对检测结果有极显著影响,在含质量体积分数15％蔗糖的培养基上花粉活力最高,而在含质量体积分数20％和25％蔗糖的培养基上花粉均不能萌发;在含100和200 mg ·L-1H3BO3的培养基中添加200 mg·L-1Ca(NO3)2均能显著提高花粉活力;培养温度对花粉萌发速率有影响但对花粉活力没有明显影响.TTC染色法的最优检测条件为:用5.0g·L-1TTC染液(pH 7.0)在35℃下染色2.0h;离体培养法的最优检测条件为:用含质量体积分数15％蔗糖、100 mg·L-1H3BO3和200 mg·L-1Ca(NO3)2的培养基暗培养4d.在25℃、4℃、0℃、-20℃、-80℃和-196℃条件下储藏13周,‘Hicksii'花粉活力和保持时间有明显差异,其中,于-80℃和-196℃储藏3d 花粉就丧失活力;于25℃和-20℃储藏7周、4 ℃储藏10周,花粉仍有一定的活力;而在0℃条件下花粉活力最高,且储藏11周花粉仍有活力.推测曼地亚红豆杉‘Hicksii’花粉对低温的抗性较差,0℃为其适宜的储藏温度.