非折叠蛋白质应答对人胚肾细胞293A迁移特性的影响

为了研究非折叠蛋白质应答对器官发生的影响,应用衣霉素诱导非折叠蛋白质应答并观察其对人胚肾细胞系293A迁移特性的影响。在实验中,应用划痕法对细胞迁移进行观察,并应用细胞黏附实验、荧光染色技术、扫描电镜技术及免疫印迹实验分别对细胞黏附特性、微管及微丝、细胞表面边缘的突起及小分子GTPase的表达水平进行研究。结果表明,非折叠蛋白质应答可以抑制细胞迁移,进一步的研究发现,非折叠蛋白质应答可以降低细胞的黏附能力、引起细胞骨架的重排、抑制伪足的形成并降低RhoA的表达水平。这提示,非折叠蛋白质应答可能通过抑制应激细胞的迁移为应激细胞的功能修复赢得了时间,在器官发生过程中发挥作用。     

CHO细胞工程化改造的研究进展

该文综述了CHO细胞工程化改造相关研究的最新进展,对CHO细胞在调节代谢、抗凋亡和糖基化等方面的工程改造及应用进行了归纳和总结,提出了CHO表达系统应用中可能出现的问题,并对CHO细胞表达系统应用前景进行了展望,以期为后续相关研究提供思路。     

细胞组学及其在药物研发中的应用

细胞组学(cytomics)是一门基于细胞分析技术的科学,它是在细胞水平对生物体系的研究,具有真实、简单和系统性的特点,在生物医学研究中有很好的应用前景。现对细胞组学的概念、特点和内容进行介绍,并结合药物研发的现状和过程,综述了细胞组学在药物研发各阶段的应用,最后对其前景进行了展望。     

诱导多能干细胞及其在神经退行性疾病研究中的应用

用干细胞转录因子OCT4、SOX2、c-MYC和KLF4进行体细胞重编程产生具有胚胎干细胞特性的诱导多能干细胞(iPS细胞)是干细胞研究领域的突破性进展。近年来,iPS细胞的研究从产生方法、重编程机理及实际应用方面不断取得进展。由于iPS细胞的产生可取自体细胞,因而克服了胚胎干细胞应用的伦理学和免疫排斥等缺陷,为iPS细胞的临床应用开辟了广阔的前景。该文将对iPS细胞的产生方法、重编程机理及其在神经性退行性疾病的研究与应用进行文献综述,反映近几年iPS细胞最新研究成果,并阐述了用病人iPS细胞模型探讨帕金森氏病、老年性痴呆症、脊髓侧索硬化症、脊髓肌肉萎缩症及舞蹈症等5种常见神经性退行性疾病发病机理的研究现状。     

细胞谱系示踪技术

细胞谱系示踪(cell lineage tracing)是指利用各种方式标记细胞,并对包括其后代所有细胞的增殖、分化以及迁移等活动进行追踪观察。自20世纪以来,谱系示踪技术为研究器官发育、组织损伤修复以及单细胞的分化命运提供了重要的手段。近些年,随着基因工程技术的飞速发展,细胞谱系示踪技术也有所突破,尤其是诱导性重组酶Cre/loxp系统的应用,极大地拓宽了细胞谱系示踪技术的应用范围。本文将结合细胞谱系示踪技术在多种研究中的应用,对该技术的原理、特点以及最新进展做一综述。     

微生物基因组简化的研究进展

微生物基因组简化是合成生物学研究热点之一。基因组的适度精简可使细胞代谢途径得以优化,改善细胞对底物、能量的利用效率,大大提高细胞生理性能的预测性和可控性。基因组简化细胞将为生物技术的应用提供理想的底盘细胞。同顾了构建基因组简化细胞的研究策略、研究方法及一些模式生物相关研究进展,总结了基因组简化研究所面临的问题及解决办法,对基因组减小化研究发展趋势前景进行了展望。     

嗅鞘细胞治疗脊髓损伤的研究进展

本文通过广泛查阅近几年嗅鞘细胞治疗脊髓损伤的国内外相关文献,发现嗅鞘细胞可以分泌众多的神经营养因子并表达相应受体,且能够调节星型胶质细胞的反应性,降低其神经胶质酸性蛋白和硫酸软骨素糖蛋白的表达水平,且能与其更好的融合在一起.在临床应用方面,嗅鞘细胞供应来源、应用时机及纯度正引起关注及研究.嗅鞘细胞对于治疗脊髓损伤有巨大的应用前景,对嗅鞘细胞进行基因改造、联合应用其他有促进作用的治疗方法将是未来的研究方向.     

单细胞组学技术及其在植物保卫细胞研究中的应用

单细胞组学技术在动物研究中已经得到广泛应用,但在植物学领域尤其是保卫细胞研究中还处于起步阶段。由保卫细胞构成的气孔承担着植物生命过程中水分散发及气体交换大门的作用。将单细胞组学技术应用到保卫细胞功能解析中将有助于了解保卫细胞参与的基本生理过程。该文综述了植物单细胞组学技术的发展、保卫细胞研究现状及单细胞组学技术在植物保卫细胞研究中的初步应用,为借助该技术解决植物生物学中保卫细胞发育、代谢及对环境胁迫响应等基本问题提供研究思路和方法。     

哺乳动物味蕾细胞分型及其细胞间信息传递

味觉是动物基本的生理感觉之一,在人类的感官研究方面,味觉一直滞后于视觉、嗅觉、触觉和听觉.味蕾是味觉的主要感受器,由于传统的细胞生物学研究手段很难在味觉研究中得到应用,人类和动物味觉的信号传递与编码机制目前还处于探索阶段,是目前的研究热点之一.近年来,随着现代分子细胞生物学和微电子传感器技术的发展和应用,味觉研究取得了较大进展.本文主要对近年来对味蕾结构和味细胞间的信号传递研究成果进行了综述,重点介绍了味细胞分型及其特征、味蕾细胞间信号传递途径及其编码机制.     

胚胎干细胞的研究进展

胚胎干细胞是来源于早期胚胎内细胞团的、具有多向分化潜能的细胞,已经广泛用于生命科学的许多领域,它在医学方面的应用成为医学领域的研究热点.本文对胚胎干细胞在诱导分化、基础研究和临床应用上的研究进展进行了综述,对今后的研究方向进行了展望.     

Applications of flow cytometry sorting in the pharmaceutical industry: A review

The article reviews applications of flow cytometry sorting in manufacturing of pharmaceuticals. Flow cytometry sorting is an extremely powerful tool for monitoring, screening and separating single cells based on any property that can be measured by flow cytometry. Different applications of flow cytometry sorting are classified into groups and discussed in separate sections as follows: (a) isolation of cell types, (b) high throughput screening, (c) cell surface display, (d) droplet fluorescent-activated cell sorting (FACS). Future opportunities are identified including: (a) sorting of particular fractions of the cell population based on a property of interest for generating inoculum that will result in improved outcomes of cell cultures and (b) the use of population balance models in combination with FACS to design and optimize cell cultures.     

PC-1基因在前列腺癌细胞系LNCaP和C4-2不同细胞周期的表达

目的:检测PC-1基因在前列腺癌细胞周期中各时间点的表达变化。方法:用200 ng/mL诺可唑(nocoda-zole)处理前列腺癌细胞系LNCaP和C4-2,16 h后使细胞处于G2/M期,在不同时间点收获细胞,分别进行流式分析和Western印迹,检测PC-1基因的表达。结果:流式分析和Western印迹结果显示,在G2/M期,LNCaP和C4-2前列腺癌细胞系中PC-1基因高表达。结论:PC-1基因的表达与前列腺癌细胞的细胞周期有关,提示PC-1可能在细胞周期调控中发挥作用。     

Comparison of methods for assessing integrity of equine sperm membranes

Sperm membrane integrity (SMI) is thought to be an important measure of stallion sperm quality. The objective was to compare three methods for evaluating SMI: flow cytometry using SYBR-14/propidium iodide (PI) stain; an automated cell counting device using PI stain; and eosin-nigrosin stain. Raw equine semen was subjected to various treatments containing 20 to 80% seminal plasma in extender, with differing sperm concentrations, to simulate spontaneous loss of SMI. The SMI was assessed immediately, and after 1 and 2 d of cooled storage. Agreement between methods was determined according to Bland-Altman methodology. Eosin-nigrosin staining yielded higher (2%) overall mean values for SMI than did flow cytometry. Flow cytometry yielded higher (6%) overall mean values for SMI than did the automated cell counter. As percentage of membrane-damaged sperm increased, agreement of SMI measurement between methods decreased. When semen contained 50-79% membrane-intact sperm, the 95% limits of agreement between SMI determined by flow cytometry and eosin-nigrosin staining were greater (range = −26.9 to 24.3%; i.e., a 51.2% span) than for SMI determined by flow cytometry and the automated cell counter (range = −3.1 to 17.0%; 20.1% span). When sperm populations contained <50% membrane-intact sperm, the 95% limits of agreement between SMI determined by flow cytometry and eosin-nigrosin staining were greater (range = −35.9 to 19.0%; 54.9% span) than for SMI determined by flow cytometry and the automated cell counter (range = −11.6 to 28.7%; 40.3% span). We concluded that eosin-nigrosin staining assessments of percent membrane-intact sperm agreed less with flow cytometry when <80% of sperm had intact membranes, whereas automated cell counter assessments of percent membrane-intact sperm agreed less with flow cytometry when <30% of sperm had intact membranes.     

Evaluation of cell numbers and viability of Saccharomyces cerevisiae by different counting methods

Four counting methods (two flow cytometry, one Coulter principle, one microscopic) are compared for measuring cell density and viability of batch-grown yeast. All gave adequate precision in measuring total cell density with no systematic difference between methods. However, the promise of flow cytometry as a rapid means of determining both quantity and quality of a cell population is shown.     

流式细胞术的发展及在植物研究中的应用

流式细胞术是通过对液流中各种荧光标记的颗粒进行多参数快速高效的定性或定量测定的方法,在科学研究的多个领域发挥重要作用。然而,由于植物组织及细胞壁和次生代谢产物等细胞的特殊成分和结构,限制了其在植物研究领域的应用。本文在介绍流式细胞仪发展和组成分类的基础上,着重讨论了流式细胞术在植物领域的应用、研究进展及应用限制,进而展望该研究领域的发展趋势,为拓宽植物流式细胞术的潜在应用范围提供新的思考方向。     

Flow cytometry: basic principles and applications

Flow cytometry is a sophisticated instrument measuring multiple physical characteristics of a single cell such as size and granularity simultaneously as the cell flows in suspension through a measuring device. Its working depends on the light scattering features of the cells under investigation, which may be derived from dyes or monoclonal antibodies targeting either extracellular molecules located on the surface or intracellular molecules inside the cell. This approach makes flow cytometry a powerful tool for detailed analysis of complex populations in a short period of time. This review covers the general principles and selected applications of flow cytometry such as immunophenotyping of peripheral blood cells, analysis of apoptosis and detection of cytokines. Additionally, this report provides a basic understanding of flow cytometry technology essential for all users as well as the methods used to analyze and interpret the data. Moreover, recent progresses in flow cytometry have been discussed in order to give an opinion about the future importance of this technology.     

DNA content analysis of insect cell lines by flow cytometry

The DNA content of insect cell lines (6 lepidoptera, 1 coleoptera and 1 diptera) was determined by flow cytometry. The DNA profiles of the 8 cell lines tested were different. They were characterized by the presence of several peaks (2 to 7) corresponding to different ploidy levels, by differences in the fluorescence intensity of each peak and by the proportion of cells in each peak. Two cell lines (Cf124 and BmN) were constituted of 2 distinct populations of cells. The DNA profiles of the cell lines were stable among the passages and during the length of time culture. This technique was demonstrated to be useful for the detection of mixed cell lines and nucleopolyhedrovirus cell infection, using Autographa californica MNPV. The flow cytometry gives interesting results on the cell cycle and the ploidy level; it appears as a good tool for insect cell lines characterization. This revised version was published online in July 2006 with corrections to the Cover Date.     

Flow cytometry and FACS applied to filamentous fungi

Flow cytometry is an automated, laser- or impedance-based, high throughput method that allows very rapid analysis of multiple chemical and physical characteristics of single cells within a cell population. It is an extremely powerful technology that has been used for over four decades with filamentous fungi. Although single cells within a cell population are normally analysed rapidly on a cell-by-cell basis using the technique, flow cytometry can also be used to analyse cell (e.g. spore) aggregates or entire microcolonies. Living or fixed cells can be stained with a wide range of fluorescent reporters to label different cell components or measure different physiological processes. Flow cytometry is also suited for measurements of cell size, interaction, aggregation or shape using non-labelled cells by means of analysing their light scattering characteristics. Fluorescence-activated cell sorting (FACS) is a specialized form of flow cytometry that provides a method for sorting a heterogeneous mixture of cells into two or more containers based upon the fluorescence and/or light scattering properties of each cell. The major advantage of analysing cells by flow cytometry over microscopy is the speed of analysis: thousands of cells can be analysed per second or sorted in minutes. Drawbacks of flow cytometry are that specific cells cannot be followed in time and normally spatial information relating to individual cells is lacking. A big advantage over microscopy is when using FACS, cells with desired characteristics can be sorted for downstream experimentation (e.g. for growth, infection, enzyme production, gene expression assays or ‘omics’ approaches). In this review, we explain the basic concepts of flow cytometry and FACS, define its advantages and disadvantages in comparison with microscopy, and describe the wide range of applications in which these powerful technologies have been used with filamentous fungi.     

Distinct volume distribution of viable and non-viable hybridoma cells: A flow cytometric study

Light scattering properties of hybridoma cells were examined with flow cytometry. Viable and dead cells form two distinct populations. The distribution of the two populations changes during a batch culture. the concentration of dead cells measured by flow cytometry correlates well to that measured by hemacytometer. The distribution based on small-angle light scattering is similar to the distribution based on volume as measured by Elzone particle counter. It thus appears that viable cells form the population with a larger mean cell volume. The results also indicate that the volume of viable cells decreases during the cultivation while that of dead cells remains relatively constant.     

Cell cycle analysis of synchronized chinese hamster cells using bromodeoxyuridine labeling and flow cytometry

Summary Chinese hamster ovary cells were synchronized into purified populations of viable G1-, S-, G2-, and M-phase cells by a combination of methods, including growth arrest, aphidicolin block, cell cycle progression, mitotic shake-off, and centrifugal elutriation. The DNA content and bromodeoxyuridine (BrdUrd) labeling index were measured in each purified fraction by dual-parameter flow cytometry. The cell cycle distributions determined from the DNA measurements alone (single parameter) were compared with those calculated from both DNA and BrdUrd data (dual parameter). The results show that highly purified cells can be obtained using these methods, but the assessed purity depends on the method of cell cycle analysis. Using the single versus dual parameter measurement to determine cell cycle distributions gave similar results for most phases of the cell cycle, except for cells near the transition from G1- to S-phase and S- to G2-phase. There the BrdUrd labeling index determined by flow cytometry was more sensitive for detecting small amounts of DNA synthesis. As an alternative to flow cytometry, a simple method of measuring BrdUrd labeling index on cell smears was used and gave the same result as flow cytometry. Measuring both DNA content and DNA synthesis improves characterization of synchronized cell populations, especially at the transitions in and out of S-phase, when cells are undergoing dramatic shifts in biochemical activity.