ELECTROPHORESIS

电泳
  • 文章类型: Journal Article
    使用MultiphysicsCOMSOL模拟了直径为20nm的二氧化硅(SiO2)纳米颗粒被等离子体纳米孔传感器捕获时产生的双峰光电数据,并与传感器测量值进行了比较,以实现紧密匹配的实验参数。纳米传感器,采用自诱导反向作用(SIBA)在固态纳米孔(ssNP)顶部的双纳米孔(DNH)结构的中心光学捕获纳米颗粒。这种SIBA驱动的纳米孔电泳(SANE)传感器能够同时捕获由作用在被捕获的SiO2纳米颗粒上的几个潜在力产生的光学和电学数据:等离子体光学捕获,电渗,电泳,粘性阻力,和热传导力。Multiphysics模拟能够解剖作用在纳米颗粒上的力的相对贡献,作为其上方和通过传感器的ssNP的位置的函数。模拟和实验之间的比较证明了在纳米颗粒进入和离开SANE传感器时生成的光学和电学时间序列数据的定性相似性。这些实验参数匹配的模拟表明,光力和电力之间的竞争将捕获平衡位置移至靠近ssNP顶部开口的位置,相对于位于几nm以上的光学捕获力最大值。实验估计的捕获SiO2纳米粒子所需的光学力的最小值与相应的光电力平衡的模拟预测一致。Multiphysics模拟与实验的比较提高了我们对光和电力之间的相互作用的理解,作为跨等离子体纳米孔传感器的纳米粒子位置的函数。
    Bimodal optical-electrical data generated when a 20 nm diameter silica (SiO2) nanoparticle was trapped by a plasmonic nanopore sensor were simulated using Multiphysics COMSOL and compared with sensor measurements for closely matching experimental parameters. The nanosensor, employed self-induced back action (SIBA) to optically trap nanoparticles in the center of a double nanohole (DNH) structure on top a solid-state nanopores (ssNP). This SIBA actuated nanopore electrophoresis (SANE) sensor enables simultaneous capture of optical and electrical data generated by several underlying forces acting on the trapped SiO2 nanoparticle: plasmonic optical trapping, electroosmosis, electrophoresis, viscous drag, and heat conduction forces. The Multiphysics simulations enabled dissecting the relative contributions of those forces acting on the nanoparticle as a function of its location above and through the sensor\'s ssNP. Comparisons between simulations and experiments demonstrated qualitative similarities in the optical and electrical time-series data generated as the nanoparticle entered and exited from the SANE sensor. These experimental parameter-matched simulations indicated that the competition between optical and electrical forces shifted the trapping equilibrium position close to the top opening of the ssNP, relative to the optical trapping force maximum that was located several nm above. The experimentally estimated minimum for the optical force needed to trap a SiO2 nanoparticle was consistent with corresponding simulation predictions of optical-electrical force balance. The comparison of Multiphysics simulations with experiments improves our understanding of the interplay between optical and electrical forces as a function of nanoparticle position across this plasmonic nanopore sensor.
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  • 文章类型: Journal Article
    分析检测方法在科学研究中起着举足轻重的作用,能够识别和量化各种学科中的特定分析物。本科学报告旨在比较两种非常不同的确定分子质量的方法(MM,也称为分子量,MW)的蛋白质:电泳凝胶和干涉光学检测方法(IODM)。为此,选择具有不同MM的几种蛋白质。电泳技术用于验证基质金属肽酶9抗体(抗MMP9)的不同部分或片段的结构和MM,通过检查具有预期大小的条带的存在,针对S100钙结合蛋白A6的抗体(抗S100A6)和胱抑素S4抗体(抗CST4)。将IODM用于研究上述蛋白质(抗体的一部分)以及蛋白质G,作为将MM和蛋白质大小与测量信号相关联的参考。我们首次报道了IODM作为测定蛋白质MM的竞争性分析方法的证据。这种创新的方法允许使用最小的样品体积和浓度进行准确的MM测定,采用简单的实验程序,消除了蛋白质变性的要求。
    Analytical detection methods play a pivotal role in scientific research, enabling the identification and quantification of specific analytes in various disciplines. This scientific report aims to compare two very different methodologies for determining the Molecular Mass (MM, also known as Molecular Weight, MW) of proteins: electrophoresis gel and the Interferometric Optical Detection Method (IODM). For this purpose, several proteins with different MM were selected. The electrophoresis technique was employed to validate the structure and MM of different parts or fragments of the Matrix Metallopeptidase 9 antibody (anti-MMP9), antibody against S100 calcium binding protein A6 (anti-S100A6) and Cystatin S4 antibody (anti-CST4) by examining the presence of bands with expected sizes. The IODM was applied to study the above-mentioned proteins (part of the antibodies) together with the protein G, as a reference to correlate the MM and protein sizes with the measured signal. We report the evidence of IODM as a competitive analytical approach for the determination of the MM of proteins for the first time. This innovative method allows for accurate MM determination using minimal sample volumes and concentrations, employing a simple experimental procedure that eliminates the requirement for protein denaturation.
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  • 文章类型: Journal Article
    背景:三分之二的免疫球蛋白轻链(AL)淀粉样变性患者有肾脏受累。肾脏损害的生化特征描述不佳。
    方法:在2010年1月1日至2022年4月30日在布宜诺斯艾利斯意大利医院进行了一项横断面研究,涉及被诊断患有AL淀粉样变性和肾脏受累的患者。参与者从机构淀粉样变性登记处进行回顾性鉴定。包括诊断为AL淀粉样变性的患者和肾脏受累的证据。排除患有其他类型淀粉样变性的个体。选择过程涉及对医疗记录和注册数据的彻底审查,以确保准确识别和纳入合格的参与者。
    结果:纳入77例患者。诊断时,90%的受试者有蛋白尿,中位数为4.3克/24小时,61%有肾衰竭,47%的人出现肾病综合征。半自动尿电泳显示55%为非选择性,21%为中度选择性肾小球蛋白尿。尿液免疫固定表明,λ单克隆游离轻链占64%,κ占12%。血清免疫固定显示λ单克隆类型为48%,λIgG为25%。在诊断AL淀粉样变性时,中位年龄为66岁(IQR53~72),49%为男性.除了肾脏受累,其他器官也受到影响:53%的心脏,19%的胃肠系统,周围神经系统占16%,16%的患者的肝脏。
    结论:我们的研究提供了拉丁美洲人群中由于免疫球蛋白轻链引起的肾淀粉样变性的生化特征。蛋白尿是该队列中最常见的多器官受累的发现。
    BACKGROUND: Two-thirds of patients with immunoglobulin light chain (AL) amyloidosis have renal involvement. The biochemical profile of kidney damage is poorly described.
    METHODS: A cross-sectional study was conducted involving patients diagnosed with AL amyloidosis and renal involvement between January 1, 2010, and April 30, 2022 at the Hospital Italiano de Buenos Aires. Participants were retrospectively identified from the Institutional Amyloidosis Registry. Patients diagnosed with AL amyloidosis and evidence of renal involvement were included. Individuals with other types of amyloidosis were excluded. The selection process involved a thorough review of medical records and registry data to ensure accurate identification and inclusion of eligible participants.
    RESULTS: Seventy-seven patients were included. At diagnosis, 90% of the subjects had proteinuria, with a median of 4.3 g/24 h, 61% had renal failure, and 47% presented nephrotic syndrome. Semi-automated urinary electrophoresis revealed 55% with non-selective and 21% with moderately selective glomerular proteinuria. Urine immunofixation indicated 64% with lambda monoclonal free light chains and 12% with kappa. Serum immunofixation demonstrated 48% with lambda monoclonal type and 25% with lambda IgG. At the time of diagnosis of AL amyloidosis, the median age was 66 years (IQR 53-72) and 49% were men. In addition to kidney involvement, other organs were also affected: heart in 53%, gastrointestinal system in 19%, peripheral nervous system in 16%, and liver in 16% of patients.
    CONCLUSIONS: Our study provides a biochemical profile in renal amyloidosis due to immunoglobulin light chains in a Latin American population. Proteinuria emerged as the most common finding in this cohort with frequent multiorgan involvement.
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  • 文章类型: English Abstract
    血管性血友病因子(VWF)多聚体分布的评估,特别是在植入循环支持装置之后,是止血的关键参数。我们的研究旨在使用SebiaHydrasys分析仪评估VWF多聚体的半自动定量。我们的分析集中在量化高分子量,中等重量,和低分子量VWF多聚体。使用Hydrasys2扫描进行电泳迁移,并使用光密度分析与Pholeisation软件进行解释。Hydrasys扫描2基于vonWillebrand病的类型成功地分离了所有预期的VWF多聚体谱。分析显示,在有循环支持装置的患者中,血浆VWF水平升高使得多聚体迁移无法使用制造商推荐的方法进行分析。因此,调整到100%VWF抗原水平提高凝胶精度。我们还建议使用Cryocheck™血浆作为标准化对照,并建立了参考值。总的来说,这种半自动的,标准化,和优化的VWF多聚体分析系统允许VWF多聚体配置文件的有效评估。
    The assessment of von Willebrand factor (VWF) multimer distribution, particularly following the implantation of circulatory support devices, is a crucial parameter in hemostasis. Our study aimed to evaluate the semi-automated quantification of VWF multimers using the Sebia Hydrasys analyzer. Our analysis focused on quantifying high molecular weight, intermediate weight, and low molecular weight VWF multimers. Electrophoretic migration was performed using the Hydrasys 2 scan, and interpretation was carried out using densitometric analysis with the Phoresis software. The Hydrasys scan 2 successfully separated all the expected VWF multimer profiles based on the type of von Willebrand disease. The analysis revealed that in patients with circulatory support devices, elevated levels of plasma VWF rendered multimer migration unanalyzable using the methodology recommended by the manufacturer. Therefore, adjustment to a 100 % VWF antigenic level improved gel precision. We also suggest using as a standardized control the Cryocheck™ plasma, and have established reference values. Overall, this semi-automated, standardized, and optimized VWF multimer analysis system allows for an effective assessment of the VWF multimeric profile.
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  • 文章类型: Journal Article
    单个细胞内的细胞器异质性和细胞器间接触导致当前细胞器分离技术的灵敏度有限。从而阻碍细胞器亚群的表征。这里,我们使用基于直流绝缘体的介电电泳(DC-iDEP)作为无偏分离方法,并通过从INS-1E胰岛素瘤细胞中鉴定胰岛素囊泡的不同分布模式来证明其能力。已应用了具有更大范围和灵敏度的多电压DC-iDEP策略,并且分化因子(电动与介电泳迁移率的比率)已用于表征胰岛素囊泡分布模式的特征。我们观察到从葡萄糖刺激细胞中分离的胰岛素囊泡相对于未刺激细胞的分布模式存在显着差异,根据葡萄糖刺激时囊泡的成熟。我们将分布模式的差异解释为表明囊泡亚群的高分辨率分离。DC-iDEP为未来表征任何生物系统中细胞器亚群的细微生化差异提供了途径。
    Organelle heterogeneity and inter-organelle contacts within a single cell contribute to the limited sensitivity of current organelle separation techniques, thus hindering organelle subpopulation characterization. Here, we use direct current insulator-based dielectrophoresis (DC-iDEP) as an unbiased separation method and demonstrate its capability by identifying distinct distribution patterns of insulin vesicles from INS-1E insulinoma cells. A multiple voltage DC-iDEP strategy with increased range and sensitivity has been applied, and a differentiation factor (ratio of electrokinetic to dielectrophoretic mobility) has been used to characterize features of insulin vesicle distribution patterns. We observed a significant difference in the distribution pattern of insulin vesicles isolated from glucose-stimulated cells relative to unstimulated cells, in accordance with maturation of vesicles upon glucose stimulation. We interpret the difference in distribution pattern to be indicative of high-resolution separation of vesicle subpopulations. DC-iDEP provides a path for future characterization of subtle biochemical differences of organelle subpopulations within any biological system.
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  • 文章类型: Journal Article
    背景:离子电渗疗法(IPT)是一种非侵入性技术,它使用电脉冲将带电分子输送到皮肤中以进行受控和靶向的药物输送。IPT已被探索为Peyronie病(PD)的非侵入性治疗选择,但是目前这方面的文献仍然很少。
    目的:我们旨在系统回顾当前有关IPT在PD管理中应用的文献,以提供对该主题的全面评估和整体展望。
    方法:在以下数据库中实施了全面的搜索策略,以检索研究文章:PubMed(MEDLINE),Scopus,和WebofScience。谷歌学者也被手动搜索。根据预定义的纳入标准,将搜索结果导入Rayyan参考管理中进行评估。文章的质量由适当的JBI检查表(即,根据研究设计),并使用JBI推荐等级对证据进行分级.
    结果:系统搜索产生451种出版物,其中11项符合本系统审查的标准。结果表明,IPT,通常使用维拉帕米和地塞米松,在治疗PD方面已显示出有希望的结果。这些方法可以减轻疼痛,斑块大小,和阴茎弯曲,同时改善性功能和生活质量,无严重不良事件。然而,大多数研究有中等到低质量,表明对某种健康管理策略的推荐力度较弱。
    结论:根据现有文献,目前没有足够的证据支持使用IPT来管理PD.将其放在研究的最前沿可以进一步促进PD的管理选择,考虑到它的治疗潜力。
    BACKGROUND: Iontophoresis therapy (IPT) is a noninvasive technique that uses electrical impulses to deliver charged molecules into the skin for controlled and targeted drug delivery. IPT has been explored as a noninvasive treatment option for Peyronie\'s disease (PD), but the current literature in this regard is still scarce.
    OBJECTIVE: We aimed to systematically review the current literature on the application of IPT in the management of PD to provide a comprehensive evaluation and holistic outlook on the subject.
    METHODS: A comprehensive search strategy was implemented in the following databases to retrieve research articles: PubMed (MEDLINE), Scopus, and Web of Science. Google Scholar was also manually searched. The search results were imported into Rayyan reference management for assessment based on the predefined inclusion criteria. The quality of the articles was evaluated by the proper JBI checklist (ie, per the study design), and the JBI grades of recommendation were used for grading the evidence.
    RESULTS: A systematic search yielded 451 publications, 11 of which met the criteria to be included in this systematic review. The results demonstrated that IPT, usually with verapamil and dexamethasone, has shown promising results in treating PD. These methods can reduce pain, plaque size, and penile curvature while improving sexual function and quality of life with no serious adverse events. However, most studies had moderate to low quality, indicating a weak recommendation for a certain health management strategy.
    CONCLUSIONS: Based on the extant literature, there is currently insufficient evidence to support the use of IPT for the management of PD. Placing it in the forefront of research can facilitate the management choices for PD even further, given its therapeutic potential.
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  • 文章类型: Journal Article
    转移性癌细胞的扩散对癌症治疗提出了重大挑战,使早期检测和诊断的创新方法至关重要。介电泳阻抗谱(DEPIS),一个强大的细胞分析工具,结合介电泳(DEP)和阻抗谱(IS)来分离,排序,细胞并分析其介电特性。在这项研究中,我们开发并构建了平面外喷墨打印的城堡状阵列,以绘制MDA-MB-231乳腺癌细胞亚型在其转移潜力中的介电特性。这是通过调节连接蛋白43(Cx43)的表达来实现的,与乳腺癌预后不良和转移增加相关的标志物。我们采用了基于DEP的诱捕,其次是对大量细胞群体的EIS测量,用于根据癌细胞的转移状态快速捕获和分化。我们的结果表明,各种MDA-MB-231转移亚型与其各自的介电泳和介电特性之间存在显着相关性。值得注意的是,具有最高转移潜能的细胞表现出最高的膜电容16.88±3.24mFm-2,其次是具有低于14.3±2.54mFm-2的膜电容的较少转移细胞亚型。此外,与转移较少的亚型(≥27±1kHz)相比,高转移性细胞表现出更低的交叉频率(25±1kHz),细胞分选的重要特征。最后,EIS测量显示转移亚组之间在1kHz处不同的双层电容(CDL)值,确认与细胞转移状态相关的独特介电和介电泳特性。我们的发现强调了DEPIS作为一种非侵入性和快速分析工具的潜力,提供对癌症生物学的见解,并促进针对不同转移阶段的个性化治疗干预措施的开发。
    The spread of metastatic cancer cells poses a significant challenge in cancer treatment, making innovative approaches for early detection and diagnosis essential. Dielectrophoretic impedance spectroscopy (DEPIS), a powerful tool for cell analysis, combines dielectrophoresis (DEP) and impedance spectroscopy (IS) to separate, sort, cells and analyze their dielectric properties. In this study, we developed and built out-of-plane inkjet-printed castellated arrays to map the dielectric properties of MDA-MB-231 breast cancer cell subtypes across their metastatic potential. This was realized via modulating the expression of connexin 43 (Cx43), a marker associated with poor breast cancer prognosis and increased metastasis. We employed DEP-based trapping, followed by EIS measurements on bulk cell population, for rapid capture and differentiation of the cancer cells according to their metastatic state. Our results revealed a significant correlation between the various MDA-MB-231 metastatic subtypes and their respective dielectrophoretic and dielectric properties. Notably, cells with the highest metastatic potential exhibited the highest membrane capacitance 16.88 ± 3.24 mF m-2, followed by the less metastatic cell subtypes with membrane capacitances below 14.3 ± 2.54 mF m-2. In addition, highly metastatic cells exhibited lower crossover frequency (25 ± 1 kHz) compared to the less metastatic subtypes (≥27 ± 1 kHz), an important characteristic for cell sorting. Finally, EIS measurements showed distinct double layer capacitance (CDL) values at 1 kHz between the metastatic subgroups, confirming unique dielectric and dielectrophoretic properties correlated with the metastatic state of the cell. Our findings underscore the potential of DEPIS as a non-invasive and rapid analytical tool, offering insights into cancer biology and facilitating the development of personalized therapeutic interventions tailored to distinct metastatic stages.
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  • 文章类型: Journal Article
    目标:通过一个大的参数空间,电场可以调整胶体相互作用和力,导致不同的静态和动态结构。到目前为止,然而,场驱动的相互作用仅限于偶极-偶极和流体动力学贡献。尽管如此,在这项工作中,我们建议在适当的条件下,电场也可以诱导基于局部化学场和扩散泳流的相互作用。
    方法:这里,我们提出了一种在电场下生成和测量3D化学梯度的策略。在这种方法中,电极上的法拉第反应会引起全局pH梯度,从而通过电扩散电泳驱动长距离传输。同时,电场通过驱动颗粒的双层远离平衡来诱导局部pH梯度。
    结果:因此,而全局pH梯度导致2D聚焦远离电极,局部pH梯度在第三维度诱导聚集。证据指出了基于扩散电泳的相互作用机制。粒子间相互作用显示出对表面化学的强烈依赖性,zeta电位和颗粒直径。此外,通过调节电场的电压和频率可以容易地调节pH梯度。对于大Péclet数字,我们观察到粒子的集体趋化崩溃。值得注意的是,这种崩塌在粒子表面没有反应的情况下发生。通过混合不同大小的颗粒,我们还证明,通过实验和布朗动力学模拟,非互惠互动的出现,小颗粒更容易被大颗粒吸引。
    OBJECTIVE: Through a large parameter space, electric fields can tune colloidal interactions and forces leading to diverse static and dynamical structures. So far, however, field-driven interactions have been limited to dipole-dipole and hydrodynamic contributions. Nonetheless, in this work, we propose that under the right conditions, electric fields can also induce interactions based on local chemical fields and diffusiophoretic flows.
    METHODS: Herein, we present a strategy to generate and measure 3D chemical gradients under electric fields. In this approach, faradaic reactions at electrodes induce global pH gradients that drive long-range transport through electrodiffusiophoresis. Simultaneously, the electric field induces local pH gradients by driving the particle\'s double layer far from equilibrium.
    RESULTS: As a result, while global pH gradients lead to 2D focusing away from electrodes, local pH gradients induce aggregation in the third dimension. Evidence points to a mechanism of interaction based on diffusiophoresis. Interparticle interactions display a strong dependence on surface chemistry, zeta potential and diameter of particles. Furthermore, pH gradients can be readily tuned by adjusting the voltage and frequency of the electric field. For large Péclet numbers, we observed a collective chemotactic-like collapse of particles. Remarkably, such collapse occurs without reactions at a particle\'s surface. By mixing particles with different sizes, we also demonstrate, through experiments and Brownian dynamics simulations, the emergence of non-reciprocal interactions, where small particles are more drawn towards large ones.
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  • 文章类型: Journal Article
    通过脱脂核桃粉(WF)获得高蛋白核桃粉(HPWF),这是石油工业的副产品。这项研究的目的是HPWF的化学和技术功能表征。Composition,氨基酸含量,蛋白质二级结构,测量蛋白质溶解度和热转变。此外,技术功能特性,乳液活性和稳定性,以及持水和吸油能力,对HPWF进行了评估。此外,通过电泳和共聚焦扫描激光显微镜评估变性条件下蛋白质的分子质量和HPWF的微观结构,分别。HPWF的蛋白质含量为55.4%,总膳食纤维为21.5%。在HPWF氨基酸组成方面,限制性氨基酸是硫酸化的半胱氨酸和蛋氨酸。通过FTIR分析,主要二级结构是β-折叠(49%),其次是α-螺旋(24%);两种结构都被认为是有序的。同样,HPWF可溶性蛋白在碱性pH下增加,并且通过电泳将HPWF蛋白分离为11条条带,分子量范围为97kDa至18kDa。关于技术功能特性,HPWF具有良好的乳液活性(51%)和高的热乳液稳定性(46%)。此外,HPWF保留了571%和242%的水和油重量,分别。最后,显微照片显示了蛋白质结构和纤维碎片的优势,和少量脂质的存在,大部分被捕获。这些结果表明,HPWF是一种有趣的植物性蛋白质来源,核桃粉可用于从非传统来源获得高蛋白成分。
    A high protein walnut flour (HPWF) was obtained by defatting walnut flour (WF), which is a by-product of the oil industry. The objective of this study was the chemical and techno-functional characterization of HPWF. Composition, amino acid content, protein secondary structure, protein solubility and thermal transitions were measured. Besides, the techno-functional properties, emulsion activity and stability, and water holding and oil absorption capacities, of HPWF were evaluated. Also, the molecular mass of proteins under denaturing conditions and the microstructure of HPWF were evaluated by electrophoresis and confocal scanning laser microscopy, respectively. HPWF had 55.4% protein content and 21.5% total dietary fibre. In terms of HPWF amino acid composition, the limiting amino acids were the sulphurated cysteine and methionine. By FTIR analysis, the main secondary structures were β-sheet (49%) followed by α-helix (24%); both structures are considered to be ordered. Likewise, HPWF soluble proteins increased at basic pH and HPWF proteins were separated in 11 bands with molecular masses ranging from 97 kDa to 18 kDa by electrophoresis. With respect to techno-functional properties, HPWF presented good emulsion activity (51%) and high thermal emulsion stability (46%). In addition, HPWF retained 571% and 242% of water and oil by weight, respectively. Finally, the micrograph showed the predominance of protein structures and fibre fragments, and the presence of few lipids mostly trapped. These results showed that HPWF is an interesting source of plant-based proteins and walnut flour can be used to obtain high protein ingredients from non-traditional sources.
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  • 文章类型: Journal Article
    背景:年龄相关性黄斑变性(AMD)是一种常见的眼部病理,主要影响老年人群。AMD的特征是进行性视网膜色素上皮(RPE)细胞变性,主要由抗氧化防御受损引起。AMD的治疗方法之一是将健康的RPE细胞注入视网膜下空间,需要纯净,健康的RPE细胞悬浮液。这项研究旨在电表征RPE细胞,以证明使用模拟通过介电电泳将健康RPE细胞与健康/氧化细胞的混合物分离的可能性。
    方法:将BPEI-1大鼠RPE细胞暴露于过氧化氢以创建体外AMD细胞模型。使用各种方法评估细胞活力,包括显微成像,基于阻抗的实时细胞分析,和MTS测定。健康和氧化的细胞通过记录它们的介电泳光谱来表征,和电池参数(交叉频率,膜电导率和介电常数,和细胞质电导率)进行计算。使用这些参数在理论的基于微流体的介电泳分离芯片上进行了COMSOL模拟。
    结果:增加过氧化氢浓度使第一个交叉频率向较低值移动,细胞膜介电常数逐渐增加。这些变化归因于进行性膜过氧化,当在用抗氧化剂N-乙酰半胱氨酸处理的细胞上测量时,它们减少了。交叉频率的变化足以有效分离健康细胞,如模拟所示。
    结论:该研究表明,介电电泳可用于根据其电特性将健康的RPE细胞与氧化的RPE细胞分离。这种方法可能是一种可行的方法,用于AMD治疗程序的健康RPE细胞悬浮液。
    BACKGROUND: Age-related macular degeneration (AMD) is a prevalent ocular pathology affecting mostly the elderly population. AMD is characterized by a progressive retinal pigment epithelial (RPE) cell degeneration, mainly caused by an impaired antioxidative defense. One of the AMD therapeutic procedures involves injecting healthy RPE cells into the subretinal space, necessitating pure, healthy RPE cell suspensions. This study aims to electrically characterize RPE cells to demonstrate a possibility using simulations to separate healthy RPE cells from a mixture of healthy/oxidized cells by dielectrophoresis.
    METHODS: BPEI-1 rat RPE cells were exposed to hydrogen peroxide to create an in-vitro AMD cellular model. Cell viability was evaluated using various methods, including microscopic imaging, impedance-based real-time cell analysis, and the MTS assay. Healthy and oxidized cells were characterized by recording their dielectrophoretic spectra, and electric cell parameters (crossover frequency, membrane conductivity and permittivity, and cytoplasm conductivity) were computed. A COMSOL simulation was performed on a theoretical microfluidic-based dielectrophoretic separation chip using these parameters.
    RESULTS: Increasing the hydrogen peroxide concentration shifted the first crossover frequency toward lower values, and the cell membrane permittivity progressively increased. These changes were attributed to progressive membrane peroxidation, as they were diminished when measured on cells treated with the antioxidant N-acetylcysteine. The changes in the crossover frequency were sufficient for the efficient separation of healthy cells, as demonstrated by simulations.
    CONCLUSIONS: The study demonstrates that dielectrophoresis can be used to separate healthy RPE cells from oxidized ones based on their electrical properties. This method could be a viable approach for obtaining pure, healthy RPE cell suspensions for AMD therapeutic procedures.
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