Fabrication technology of integrated fiber microfluidic electrophoresis chip

The technology of PCB was used to fabricate the chip mould, and the microfluidic electrophoresis chip was fabricated with PDMS material. The fiber integrated on the chip was used as the transmission medium, so the light spot size was near the depth of microchannel. The detection sensitivity was improved, and the optical focusing system was spared. The fabrication process, sealing methods and structure characteristic of PDMS microfluidic electrophoresis chips were discussed. The experiment was achieved by using the fabricated chip to separate FITC fluorescein and FITC-labeled amino acid mixture reagent, and the feasibility of the chip was validated.     

微流控毛细管电泳-流动注射联用技术在分离和测定中药制剂中麻黄碱与伪麻黄碱的应用

采用较短的毛细管作为分离通道,建立了一种微流控毛细管电泳和流动注射联用的分离测定麻黄碱和伪麻黄碱的新体系.该体系由一个H-型微芯片接口、一个水平放置的毛细管(作为分离通道)、两个竖直放置的Tygon管(作为阳极和阴极的电解质流通储液槽)组成.在最佳实验条件下,麻黄碱和伪麻黄碱标准品的进样频率可达到60 h-1,且完全基线分离,重现性良好,检测限(S/N=3)分别为:麻黄1.77 μg/mL,伪麻黄2.03 μg/mL.该微流控体系已用于3种含有麻黄碱和伪麻黄碱的市售药品的测定,结果令人满意.     

应用聚合酶链式反应检测犬细小病毒

将具有犬细小病毒病典型症状的病犬肠溶物经ＳＤＳ－酚裂解法提取总ＤＮＡ,并以此ＤＮＡ为模板,通过人工合成的两条２０ｍｅｒ引物进行ＰＣＲ扩增,该对引物扩增的片段为犬细小病毒结构蛋白ＶＰ２基因中间１／３区段,ＰＣＲ产物经琼脂糖凝胶电泳,试验结果表明：用ＰＣＲ扩增检测犬细小病毒具有良好的特异性,应用该方法检测的２７份病犬肠溶物样品块获得了预计长度的ＤＮＡ片段（５３１ｂｐ）,而健康犬的肠溶物样品ＰＣＲ结果为     

Dynamic changes of apoptosis in duck embryo fibroblasts induced by new type Gosling viral enteritis virus

The monolayer duck embryo fibroblast （DEF） cells were experimentally infected with new type Gosling viral enteritis virus （NGVEV） and the dynamic changes of apoptosis were detected at different time points after NGVEV infection by transmission electron microscopy （TEM）, DNA agarose gel electrophoresis and Annexin V-FITC/PI stained fluorescence-activated cell sorter （FACS）. The result shows that NGVEV can induce infected cells undergoing apoptosis and changing regularly. A series of characteristic apoptotic morphological changes including shrinkage of the cells, chromatin condensation and margination, as well as formation of apoptotic bodies, were observed by TEM. The typical ladder pattern of DNA fragmentation was demonstrated by agarose gel electrophoresis. And using flow cytometry analysis of Annexin V-FITC/PI staining, the dead, viable, apoptotic and necrotic cells could be analyzed quantitatively.     

Programmed cell death in Laminaria japonica (Phaeophyta) tissues infected with alginic acid decomposing bacterium

TdT-mediated dUTP-biotin nick end labeling (TUNEL) is a sensitive and valid method for detecting DNA cleavage in programmed cell death (PCD). Using this method, DNA cleavage was observed in Laminaria japonica sporophytic tissues, which were infected with alginic acid decomposing bacterium. It was found that DNA cleavage occurred 5 min after the infection, the fragments with 3′-OH groups of cleaved nuclear DNA increased with time of infection and spread from the infection site. Although no typical DNA ladder (200 bp/180 bp) was detected by routine agarose gel electrophoresis, the cleavage of nuclear DNA fragments of 97～48.5 kb could be detected by pulsed field gel electrophoresis (PFGE). By using CaspGLOWTM fluorescein active caspase-3 staining method, caspase-3 activity has been detected in response to the infection of alginic acid decomposing bacterium. Our results are similar to the observations in hypersensitive response (HR) of higher plant, suggesting that the rapid cell death of L. Japonica infected by alginic acid decomposing bacterium might be involved in PCD, and indicating that the occurrence of PCD is an active defense process against the pathogen's infection.     

毛细管电泳芯片制备工艺研究

叙述了采用SU-8和PDMS制作毛细管电泳芯片的一种新工艺、新方法,对SU-8的四个工艺参数：前烘温度与时间、后烘温度与时间、曝光时间以及显影时间进行了工艺优化,结果表明使用该方法加工工艺简单,芯片重复性好,成本低,适用于采用激光诱导荧光检测法进行检测,并在制备的毛细管电泳芯片上实现了不同长度DNA片段的快速有效分离．     

改性PDMS/PVDF复合膜对丙烷/氮气分离性能研究

以相转化法制备的聚偏氟乙烯(PVDF)多孔膜为底膜,聚二甲基硅氧烷(PDMS)为分离层膜材料,涂覆法制备了PDMS/PVDF平板复合膜.将PVDF底膜进行等离子体接枝处理,提高PVDF底膜与PDMS选择层的结合力.研究了体系温度,压力,原料气的组成等因素对复合膜分离性能的影响.随着体系温度升高,丙烷(C3H8)的渗透通量减小,而氮气(N2)的渗透通量增大.随着体系的压力增大,C3H8的渗透通量增加,N2的渗透通量几乎不变,分离因子增大.原料气中C3H8的含量增加,渗透通量增大,分离因子也增加.在温度为25℃,压力为0.3MPa,混合气中C3H8含量为10mol%时,制备复合膜的渗透通量为1.625×10-2(c.cm-2.s),分离因子为21.56.     

Semi-crosslinked polyacrylamides as high-resolution and dynamic self-coating sieving matrices for protein capillary electrophoresis

This paper describes non-gel capillary sieving electrophoresis employing semi-crosslinked polyacrylamide as a high performance and low viscous replaceable separation matrix for separation of non-denatured protein separation. Arising from the fine sieving and dynamic coating ability of this polymer, a mixture of basic proteins lysozyme, cytochrome C, ribonuclease A, and trypsin was resolved with excellent reproducibility. Mixing different semi-crosslinked polyacrylamides together further improves the separation. The separtion mechanism was analyzed. With network structure developed to an intermediate state between crosslinked gel and linear polymer solutions, these semi-crosslinked polyacrylamide polymers demonstrate a promise as a new class of size sieving separation medium, not only in capillary electrophoresis, but also in microfluidic chip separation schemes.     

基于ITO玻璃-PDMS复合式低电压电泳芯片的快速制备

提出了一种基于ITO玻璃-PDMS复合式低电压电泳芯片的快速制备新方法.该方法采用Protel软件绘制低电压电泳芯片沟道的形状,利用电路板加工技术加工沟道模具,并采用PDMS整体浇注方法制备芯片微沟道;采用光刻、湿法刻蚀等微加工技术方法对ITO导电玻璃进行加工,并制备出ITO阵列电极.最后经修饰、封装等工序得到基于ITO玻璃-PDMS复合式低电压电泳芯片.实验表明:采用该方法制备的ITO阵列电极线条清晰,沟道侧壁陡直、表面光滑.同时,采用该方法制备芯片周期短,不需要昂贵的制作设备.     

硫酸铈诱导果蝇氧化应激及细胞凋亡的研究

本实验主要研究了稀土硫酸铈（Ce（SO4）2）对果蝇氧化应激生物标记物和细胞凋亡的影响.果蝇培养在不同质量浓度（1,4,16,64,256,1 024 mg/L）的硫酸铈培养基中,分别测定其SOD,CAT和脂质过氧化产物（即MDA含量）,同时用彗星电泳和体外切割DNA实验来检测果蝇细胞中DNA损伤程度,用DNA Laddering法和TUNEL法测定稀土元素Ce对果蝇细胞凋亡的影响.与对照组相比,当Ce（SO4）2质量浓度低于16 mg/L时,果蝇体内SOD和CAT活性显著增加,MDA含量变化不明显;而当Ce（SO4）2质量浓度高于16 mg/L时,SOD和CAT活性明显下降,MDA含量上升.彗星电泳的结果表现为随着硫酸铈剂量的递增,果蝇中肠细胞的彗星率、彗星尾长和Olive尾矩增加,并表现为明显的剂量效应关系.果蝇体外实验结果表明,硫酸铈能打断DNA,使其片段化;同时,TUNEL结果显示果蝇中肠细胞呈现凋亡细胞的特征性蓝绿色颗粒,但DNA琼脂糖电泳没有表现出细胞凋亡特征性的梯形条带图谱.硫酸铈诱导果蝇的氧化应激可以使果蝇中肠细胞SOD和CAT活性降低,MDA含量上升,使中肠细胞出现凋亡特征.由此推断,硫酸铈可诱导果蝇细胞中遗传物质的损伤,对果蝇有一定的氧化毒性和遗传毒性作用．     

基于PDMS的人工视网膜神经微电极阵列

为了利用柔顺性材料实现电极位点与靶细胞的良好接触,同时保证微电极的可靠性,提出了一种新的聚二甲基硅氧烷(PDMS)微电极制作方法.该方法通过在硅基表面沉积金属层、光刻图形化以及电镀形成电极基本结构,然后通过PDMS浇注、湿法刻蚀、释放以及键合完成基于PDMS微电极制作.其中,微电极绝缘层制作和电极位点暴露采用浇注PDMS并结合外力夹压固化和PDMS湿法刻蚀来实现.使用该方法制作的PDMS电极,结构稳定、可靠性好,具有良好的贴附性.同时,通过SEM和阻抗测试对所制作的微电极进行了表面形貌和电学性能的测试和评价.结果显示,相对于传统方法制作的PDMS微电极,电化学阻抗降低了近60%（频率1 kHz处）,基于该方法制作的PDMS微电极在力学和电学性能方面均具明显优势.     

周期可调谐金属光栅的制备工艺研究

介绍了一种制备金属可调谐光栅的新技术。首先采用紫外光刻技术将光栅结构制作至PDMS（聚二甲基硅氧烷）基底，并在其上蒸镀金属Cr层和Ag层，然后采取金属举离工艺得到金属光栅。利用PDMS高弹性的特点，通过控制PDMS纵、横向拉伸程度，对金属光栅周期进行连续、任意调谐，获得预定的目标周期。实验结果表明，金属光栅调谐范围超过其初始周期的100%。     

碳纤维纳米电极用于微流控芯片安培测定单个PC12细胞中神经递质多巴胺

为研究单个PC12细胞中多巴胺的含量,以碳纤维纳米电极为工作电极,自行组装了微流控芯片安培柱末检测系统,设计了一种微流控芯片电泳单细胞进样和溶膜的控制方式,仅需一路高压控制即可实现单细胞进样和溶膜操作.结果表明：采用该系统实现了4种儿茶酚胺类物质的基线分离,通过控制适当的细胞密度及液池的液面高度差,实现了单个细胞连续快速进样、溶膜和分析,得出单个PC12细胞中多巴胺含量为0.80±0.33 fmol（n=5）.     

基于超弹性PDMS的摩擦纳米发电机的性能研究

常用于摩擦纳米发电机的聚二甲基硅氧烷是一种超弹性聚合物,但迄今对其理论分析均采用线弹性本构假设,没有通过实验证明其是否有效.首先基于超弹性本构对PDMS摩擦纳米发电机的接触力学和静电感应性能进行仿真;其次通过光刻和复型工艺制备了PDMS金字塔织构,构建了摩擦纳米发电机实验装置,并将实验与理论结果对比.研究表明:随着施加压力增大,摩擦纳米发电机的接触面积和开路电压均随之增加,之后趋于稳定;相比于超弹性本构,采用线弹性本构高估了金属与PDMS的接触面积,低估了使摩擦纳米发电机开路电压稳定所需施加的压力;采用超弹性PDMS本构的仿真结果与实验更为吻合.      

二肽衍生物的非水毛细管电泳分离

以1,2-苯并-3,4二氢咔唑-9-乙基氯甲酸酯(BCEOC)作为柱前衍生试剂,利用非水毛细管区带电泳法对衍生二肽进行分离,考察了该试剂用于非水毛细管区带电泳模式分离的几个关键条件,在有机溶剂甲醇与乙腈比例75∶25(V/V),醋酸铵浓度45 mmol/L,醋酸浓度0.8 mol/L,柱温17℃,分离电压28 kV条件下,采用二极管阵列检测,实现了11种二肽的基线分离.     

中空纤维复合膜气体分离性能的研究

本论文制备了用于有机蒸汽正庚烷/氮气分离的PDMS/PS,PDMS/PAN,PDMS/PVDF复合膜.研究了PS,PVDF基膜热处理温度和时间对复合膜的分离性能的影响.结果表明：PDMS/PS复合膜的渗透速率和分离因子均随热处理温度的升高而降低,PDMS/PVDF复合膜在393.2K下有最佳分离性能.     

Separation of dopamine and epinephrine by a novel electrophoresis technique with nafion membrane as separation column

A novel electrophoresis technique, in which a strip of perflurosulfonic-acid ( Nafion 117 ) membrane was used to replace the conventional separation column and liquid buffer solution within, was developed and employed to separate the mixture of dopamine and epinephrine under a low separation voltage of 100 V with quadruple pulses amperometry detection. It was showed that the so-called Nation membrane electrophoresis could be one of very simple and easy method and has the potentiality to be used to separate and analyze some small organic biologic molecules.     

电泳羟基磷灰石色谱吸附BSA机理

将电泳与羟基磷灰石色谱技术耦合 ,开发了一种新的电泳色谱技术。建立了羟基磷灰石 (HAP)吸附牛血清白蛋白 (BSA)的物理模型和数学模型 ,建立了不同的实验装置以研究电场下 HAP吸附 BSA的反应动力学特性以及电动色谱吸附 BSA过程的传质动力学特性。实验结果表明 ,施加电场不会改变吸附反应的平衡常数与速率 ,但在电泳羟基磷灰石色谱装置中 ,施加电场在羟基磷灰石颗粒表面产生的电渗流能够加强扰动 ,减少液膜扩散阻力 ,提高羟基磷灰石色谱的动态吸附容量     

樱桃李属坏死环斑病毒病PCR检测技术研究

通过用2种RNA提取方法提取甜樱桃植株样品总RNA,并对其进行电泳分析.结果表明,用方法A提取的RNA完整性比方法B较好,出现了较为典型的RNA带型.以方法A提取的甜樱桃总RNA为模板,用设计一对特异性引物进行RT-PCR反应,可见其DNA谱带与预期的一对引物应扩增的片断吻合,说明PCR检测是特异的.通过30份样品经RT-PCR扩增有2份表现出清晰、明显的特异带,占总样品数的6.7%.     

Conformational dynamics of individual DNA molecules during gel electrophoresis

Gel electrophoresis is widely used in molecular biology to separate DNA molecules according to their sizes. The physical basis of this size separation is, however, poorly understood. Here we report observations of individual, fluorescently stained DNA molecules as they migrate during various kinds of gel electrophoresis. Their movement, under the influence of either a steady electric field or a pulsed-field, is characterized by cycles of elongation and contraction. Initially relaxed coils of DNA lengthen into 'hook-shaped' configurations which temporarily 'hang-up' on obstacles in the gel matrix before sliding off, contracting and entering another cycle. The effects of a new electrophoresis technique, termed 'pulse-oriented electrophoresis', which allows the effective angle of the electric field, and hence the molecular orientation of DNA, to be varied without electrode rearrangement, are also studied. In this case the DNA adopts a 'staircase' configuration showing that the net orientation in a direction is given by the vector sum of the pulses used.