Name: an aptamer-based sensor for unchelated gadolinium(iii)
Uploaded: 2017-01-09T15:00:00.000+00:00
Duration: 5 min 15 s
Description: Watch this Scientific Journal Video about An Aptamer-based Sensor for Unchelated GadoliniumIII at JoVE.com

We would like to gratefully acknowledge Dr. Milan N Stojanovic from Columbia University, New York, NY for valuable scientific input. This work is supported by funding from the California State University East Bay (CSUEB) and the CSUEB Faculty Support Grant-Individual Researcher. O.E., T.C., and A.L. were supported by the CSUEB Center for Student Research (CSR) Fellowship.

注:分子生物学级水在所有的缓冲和解决方案的准备工作时使用。所有一次性管（离心及PCR）和枪头是DNase-和无RNA酶的。请咨询使用前的所有化学品的材料安全数据表（MSDS）。强烈推荐合适的个人防护装备（PPE）的使用。 1.适体母液的制备<ol><li>购买2股多脱氧核苷酸的市场。通过高效液相色谱法（HPLC）命令与纯化两条链。 串1（GD-适体）: 5&#39; - / 56-FAM / AGGCTCTCGGGACGACCAGTTGGTCCCGCTTTATGTGTCCCGAG-3&#39; 链2（QS）: 5&#39;-GTCCCGAGAGCCT / 3Dab / -3&#39; </li><li>溶解每条链在水中使钆适体和的QS100μM的个别储备溶液。 </li><li>存放于-20°C这些解决方案。该溶液是稳定的迄今，为3年。 </li><li>为了尽量减少FREEZ电子解冻周期，存储解决方案，股价在10微升等分。 </li></ol> 2. 2倍的Gd-传感器溶液的制备<ol><li>制备测定缓冲液（20mM HEPES，2mM的MgCl 2的，150毫摩尔NaCl，5mM的KCl）中。调整用NaOH和HCl pH值至7.4〜，并通过一次性无菌瓶顶过滤器具有0.2微米PES膜过滤器。存放在无菌瓶中。如果过滤，并妥善保存（在常温下），缓冲稳定到目前为止，2年。 </li><li>稀释钆适体原液（从步骤1.2）在测定缓冲液497微升和QS原液（从步骤1.2）的2微升1微升。拌匀，用旋涡。其在2×钆传感器溶液浓度是200和400nm，分别。 注意:在此步骤中制备的2倍的Gd-传感器溶液的体积为500微升，这是足以测试6 - 7不同浓度的Gd 3+的校准曲线和造影剂溶液（步骤3）。每个样品将在384孔板给复孔。 <ol><li>根据需要被测试的Gd 3+解的数目调整2倍的Gd-传感器溶液的体积。 </li></ol></li><li>转移2×钆传感器溶液到9的PCR管，用50微升到每个管中。放置管在热循环仪。 </li><li>在热循环仪设置程序以加热在管至95℃的溶液中，保持5分钟，然后经〜15分钟缓慢地冷却该溶液至25℃（以速率〜0.05 - 0.1℃/ S）。加热和冷却循环是确保钆适体和QS之间最佳杂交。部分杂交结果不完全淬火和传感器的一个更高的背景荧光。如果热循环仪是不可用，进行利用热水浴代替这个过程。 </li><li>一旦冷却到25℃，立即使用该溶液，或保持在热循环仪（最高达约2小时），直到准备好是用过的。当水浴用于加热，离开该管在浴中当水慢慢冷却到室温。 </li></ol> 3.构建荧光校准曲线和钆造影剂的溶液检测未螯合GD的存在3+ <ol><li>溶解GdCl 3在测定缓冲液固体（相同的缓冲液如在步骤2.1）。 </li><li>通过连续稀释，在最终所需浓度的校准曲线（2×溶液）的双制备各6个不同的Gd 3+在微量离心管溶液100微升。 <ol><li>例如，为了构建一个校准为0，50，100，200，400（仅无GdCl 3缓冲液），和钆3+为800nm，制备含有0，100，200，400，800，和1600纳米的解决方案的离子。确保始终包括"空白"0纳米的Gd 3+作为阴性对照。 </li></ol></li><li>溶解造影剂是测试编在测定缓冲液中。准备2个或3个不同的浓度通过连续稀释的造影剂的解决方案。 注:测试3种不同浓度造影剂溶液的建议。这是为了确保这些浓度的线性范围内。如果测试没有显示线性关系的样品中，减少造影剂的浓度使用。 </li><li>服用含有从步骤2.5 2倍的Gd-传感器解决方案出来的热循环仪的PCR管。 </li><li>添加来自步骤3.2的每个的Gd 3+溶液的50微升到含有2倍的Gd-传感器溶液9 PCR管6。吹打向上和向下混合。每个PCR管现在包含钆3+的所需浓度进行测试，100nM的钆适体，和200nM的QS。 </li><li>到含有2倍的Gd-传感器溶液剩余的PCR管，从步骤3.3添加造影剂溶液的50微升。通过上下吹打几次拌匀。 </li><li>孵育在Solutions在PCR管在室温下约5分钟。它们可以被放置长达30分钟。 </li><li>传送每个管的45微升到384孔板中。每个PCR管会给重复孔中。 </li><li>记录的每个孔中的荧光在平板阅读器。在GD-传感器设计中使用的荧光团（FAM），分别有495和520 nm的激发和发射最大值，对供应商的网站上所列出。选择取决于在板阅读器是否monochromator-或基于过滤器的合适的激发和发射波长或过滤器。 </li><li>绘制荧光任意荧光单位（AFU）对钆3+的浓度曲线图。 </li><li>画出图作为对抗的Gd 3+的荧光浓度倍数变化。由"空白"解决方案的AFU（0纳米的Gd 3+）将每个浓度的AFU计算荧光倍数变化。荧光倍数变化将使n个结果ormalization，应AFU显示一些周期性的（不同天等 ）的变化。 </li><li>比较含有造影剂的溶液和"空白"，它是含0纳米GdCl 3（仅缓冲）该溶液的荧光发射。 注:GBCA溶液的更高的荧光意味着未螯合的Gd 3+的存在，这可能需要造影剂的进一步纯化。未螯合的Gd 3+的存在量可以利用在步骤3.10或3.11构建的校准曲线来估算。 </li></ol>

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The authors declare that they have no competing financial interests.

使用基于适体的Gd-传感器，荧光发射的增加正比于观察未螯合的Gd 3+的浓度。为了最大限度地减少样品的使用量，该测定可以在384孔微量板以每孔45微升总样品体积运行。在此设计中，荧光素（FAM）和DABCYL（DAB）的选择主要基于试剂的成本;修改发射波长的荧光团不同的配对和猝灭剂，可以使用11。 要注意的是，以获得与该传感器最好的结果是很重要的，过程的...

在临床诊断中，这是由该技术的固有敏感性限制磁共振成像（MRI）的重要性日益增加，已导致在研究的快速增长到新的基于钆的造影剂（GBCAs）1的开发。 GBCAs是被给药，以提高图像质量的分子，并且它们通常具有配位的多齿配体的三价钆离子（钆3+）的化学结构。这络合作为未螯合的Gd 3+是有毒至关重要;它已经牵涉于肾源性系统纤维化的一些患者的肾疾病或衰竭2的开发。因此，检测该水性自由离子是确保GBCAs的安全工具。未螯合的Gd 3+的GBCA溶液存在往往是在配体和离子的复合物的解离，或者displacemen之间不完全反应的结果其他生物金属阳离子3吨 。 在目前用于确定的Gd 3+，那些在通用性和适用性4方面依靠色谱法和/或光谱法秩最高的存在下的几种技术。在他们的优势是较高的灵敏度和准确度，分析各种样品基质（包括人血清5，尿液和头发6，废水7和造影剂配方8），和多个钆3+络合物的同时定量（一个列表的能力的研究之前，2013是在全面审查Telgmann 等人 ）4所述。唯一的缺点是，一些这些方法需要仪器仪表（如电感耦合等离子体质谱）4，一些实验室可能无法访问。在小说GBCA发现在研究和论证的概念层面，AR上下文elatively更方便，快速和成本有效的基于分光法（如紫外吸收或荧光）可以作为一种有价值的替代。考虑到这些应用中，水的Gd 3+的荧光基于适体的传感器被开发9。 适体（钆适体）是与通过指数富集（SELEX）9的配体的系统进化的过程中分离出的碱基序列特异性的44碱基长的单链DNA分子。适应适体为荧光传感器，荧光团被连接到线料，然后将其与经由13互补碱基（ 图1）淬火链（QS）杂交的5&#39;末端。了QS标记有在3&#39;末端的暗淬灭剂分子。在不存在的Gd 3+，所述传感器（钆传感器）的，由1:分别钆适体和QS的2摩尔比，将具有最小的荧光发射是由于吨从荧光基团淬灭Ø能量转移。在加入含水的Gd 3+的将从钆适体置换的QS，从而增加荧光发射。 <img alt="图1" src="/files/ftp_upload/55216/55216fig1.jpg" /> 图 1. 传感器（GD-传感器），由标有荧光素（荧光基团），并标有DABCYL 13个碱基长的淬链（QS）的44个碱基长的适体（GD-适体）（暗淬灭剂）的。在不存在未螯合的Gd 3+的，传感器的荧光是最小的。与另外的Gd 3+的，发生的QS位移和增加的荧光发射是观察。 <a href="http://ecsource.jove.com/files/ftp_upload/55216/55216fig1large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> 有目前，人们常用的基于分光法检测ING水的Gd 3+。该测定使用了分子二甲酚橙，其经历在其最大吸收波长从433至573纳米时螯合离子10的换档。这两个最大吸收的比率可以用来量化未螯合的Gd 3+的量。适体传感器是一种替代（也可以是互补的），以在二甲酚橙法，因为这两种方法具有不同的反应条件（如pH值和所使用的缓冲溶液的组合物），目标的选择性，定量的线性范围，和检测方式9。

<table><tbody><tr><td>Gd-aptamer</td><td>IDTDNA</td><td>Input sequence and fluorophore modification in the order form</td><td>A fluorophore with a different emission wavelength may be used. The aptamer may also be ordered from another company.</td></tr><tr><td>Quenching strand</td><td>IDTDNA</td><td>Input sequence and quencher modification in the order form</td><td>A different quencher for optimal energy transfer from the fluorophore may be used. The aptamer may also be ordered from another company.</td></tr><tr><td>Molecular biology grade water</td><td></td><td></td><td>No specific manufacturer, both DEPC or non-DEPC treated work equally well</td></tr><tr><td>Gadolinium(III) chloride anhydrous</td><td>Strem</td><td>936416</td><td>Toxic</td></tr><tr><td>HEPES</td><td>Fisher Scientific</td><td>BP310-500</td><td></td></tr><tr><td>Magnesium chloride anhydrous</td><td>MP Biomedicals</td><td>0520984480 - 100 g</td><td></td></tr><tr><td>Sodium Chloride</td><td>Acros Organics</td><td>327300025</td><td></td></tr><tr><td>Potassium chloride</td><td>Fisher Scientific</td><td>P333-500</td><td></td></tr><tr><td>Sodium hydroxide, pellets</td><td>Fisher Scientific</td><td>BP359</td><td>Corrosive</td></tr><tr><td>Hydrochloric acid</td><td>Fisher Scientific</td><td>SA49</td><td>Toxic and corrosive</td></tr><tr><td>384-well low flange black flat bottom polystyrene NBS plates</td><td>Corning</td><td>3575</td><td>Plates which are suitable for fluorescence reading are required.</td></tr><tr><td>Nalgene Rapid-Flow sterile disposable bottle top filter</td><td>Thermo Scientific</td><td>5680020</td><td>The bottle top is fitted with 0.2 micron PES membrane</td></tr><tr><td>Disposable sterile bottles 250 mL</td><td>Corning</td><td>430281</td><td>A larger or smaller bottle may be used</td></tr><tr><td>1.5 mL microcentrifuge tubes</td><td></td><td></td><td>No specific manufacturer, as long as they are DNAse and RNAse-free</td></tr><tr><td>0.2 mL PCR tubes</td><td></td><td></td><td>No specific manufacturer, as long as they are DNAse and RNAse-free</td></tr><tr><td>Micropipets</td><td></td><td></td><td>No specific manufacturer</td></tr><tr><td>Pipet tips (non filter) of appropriate sizes</td><td></td><td></td><td>No specific manufacturer, as long as they are DNAse and RNAse-free</td></tr><tr><td>&lt;strong&gt;Equipment&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Plate reader</td><td>Biotek Synergy H1</td><td></td><td>Plate readers from other manufacturers would work equally well</td></tr></tbody></table>

an aptamer-based sensor for unchelated gadolinium(iii)

A method for determining the presence of unchelated trivalent gadolinium ion (Gd3+) in aqueous solution is demonstrated. Gd3+ is often present in samples of gadolinium-based contrast agents as a result of incomplete reactions between the ligand and the ion, or as a dissociation product. Since the ion is toxic, its detection is of critical importance. Herein, the design and usage of an aptamer-based sensor (Gd-sensor) for Gd3+ are described. The sensor produces a fluorescence change in response to increasing concentrations of the ion, and has a limit of detection in the nanomolar range (~100 nM with a signal-to-noise ratio of 3). The assay may be run in an aqueous buffer at ambient pH (~7 - 7.4) in a 384-well microplate. The sensor is relatively unreactive toward other physiologically relevant metal ions such as sodium, potassium, and calcium ions, although it is not specific for Gd3+ over other trivalent lanthanides such as europium(III) and terbium(III). Nevertheless, the lanthanides are not commonly found in contrast agents or the biological systems, and the sensor may therefore be used to selectively determine unchelated Gd3+ in aqueous conditions.

在未螯合的Gd 3+的存在下的Gd-传感器溶液的典型荧光变化示于图2。发射可被绘制为荧光倍数变化（ 图2A）或在原始荧光读数（ 图2B）以任意单位（AFU）。两条曲线在&gt; 3μM收率低于1μM和饱和信号的线性范围为钆浓度3+非常相似的校准曲线。检测限为〜100纳米带3的信噪比。 ...

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An Aptamer-based Sensor for Unchelated GadoliniumIII

The use of polydeoxynucleotide (44-mer aptamer) molecules for sensing unchelated gadolinium(III) ion in an aqueous solution is described. The presence of the ion is detected via an increase in the fluorescence emission of the sensor.

对于未螯合钆基于核酸适配体传感器（III）

A method for determining the presence of unchelated trivalent gadolinium ion (Gd3+) in aqueous solution is demonstrated. Gd3+ is often present in samples ...

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An Aptamer-based Sensor for Unchelated Gadolinium(III)

7.2K 次观看.  California State University, East Bay. The use of polydeoxynucleotide (44-mer aptamer) molecules for sensing unchelated gadolinium(III) ion in an aqueous solution is described. The presence of the ion is detected via an increase in the fluorescence emission of the sensor. 

视频: An Aptamer-based Sensor for Unchelated GadoliniumIII

Investigations on the Ga(III) Complex of EOB-DTPA and Its 68Ga Radiolabeled Analogue

We demonstrate a method for the isolation of EOB-DTPA (3,6,9-triaza-3,6,9-tris(carboxymethyl)-4-(ethoxybenzyl)-undecanedioic acid) from its Gd(III) complex and protocols for the preparation of its novel non-radioactive, i.e., natural Ga(III) as well as radioactive 68Ga complex. The ligand as well as the Ga(III) complex were characterized by nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry and elemental analysis. 68Ga was obtained by a standard elution method from a 68Ge/68Ga generator. Experiments to evaluate the 68Ga-labeling efficiency of EOB-DTPA at pH 3.8&#8211;4.0 were performed. Established analysis techniques radio TLC (thin layer chromatography) and radio HPLC (high performance liquid chromatography) were used to determine the radiochemical purity of the tracer. As a first investigation of the 68Ga tracers&#39; lipophilicity the n-octanol/water distribution coefficient of 68Ga species present in a pH 7.4 solution was determined by an extraction method. In vitro stability measurements of the tracer in various media at physiological pH were performed, revealing different rates of decomposition.

Investigations on the GaIII Complex of EOB-DTPA and Its 68Ga Radiolabeled Analogue

A procedure for the isolation of EOB-DTPA and subsequent complexation with natural Ga(III) and 68Ga is presented herein, as well as a thorough analysis of all compounds and investigations on labeling efficiency, in vitro stability and the n-octanol/water distribution coefficient of the radiolabeled complex.

在EOB-DTPA和镓（三）复杂的调查其<sup&gt; 68</sup&gt;嘎放射性标记模拟

We demonstrate a method for the isolation of EOB-DTPA (3,6,9-triaza-3,6,9-tris(carboxymethyl)-4-(ethoxybenzyl)-undecanedioic acid) from its Gd(III) ...

科研

化学

Preparation and In Vitro Characterization of Dendrimer-based Contrast Agents for Magnetic Resonance Imaging

Paramagnetic complexes of gadolinium(III) with acyclic or macrocyclic chelates are the most commonly used contrast agents (CAs) for magnetic resonance imaging (MRI). Their purpose is to enhance the relaxation rate of water protons in tissue, thus increasing the MR image contrast and the specificity of the MRI measurements. Current clinically approved contrast agents are low molecular weight molecules that are rapidly cleared from the body. The use of dendrimers as carriers of paramagnetic chelators can play an important role in the future development of more efficient MRI contrast agents. Specifically, the increase in local concentration of the paramagnetic species results in a higher signal contrast. Furthermore, this CA provides a longer tissue retention time due to its high molecular weight and size. Here, we demonstrate a convenient procedure for the preparation of macromolecular MRI contrast agents based on poly(amidoamine) (PAMAM) dendrimers with monomacrocyclic DOTA-type chelators (DOTA &#8211; 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetate). The chelating unit was appended by exploiting the reactivity of the isothiocyanate (NCS) group towards the amine surface groups of the PAMAM dendrimer to form thiourea bridges. Dendrimeric products were purified and analyzed by means of nuclear magnetic resonance spectroscopy, mass spectrometry, and elemental analysis. Finally, high resolution MR images were recorded and the signal contrasts obtained from the prepared dendrimeric and the commercially available monomeric agents were compared.

Preparation and In Vitro Characterization of Dendrimer-based Contrast Agents for Magnetic Resonance Imaging

This protocol describes the preparation and characterization of a dendrimeric magnetic resonance imaging (MRI) contrast agent that carries cyclen-based macrocyclic chelates coordinating paramagnetic gadolinium ions. In a series of MRI experiments in vitro, this agent produced an amplified MRI signal when compared to the commercially available monomeric analogue.

制备及<em&gt;体外</em&gt;磁共振成像基于树状大分子造影剂的表征

Paramagnetic complexes of gadolinium(III) with acyclic or macrocyclic chelates are the most commonly used contrast agents (CAs) for magnetic resonance ...

MR Molecular Imaging of Prostate Cancer with a Small Molecular CLT1 Peptide Targeted Contrast Agent

Tumor extracellular matrix has abundance of cancer related proteins that can be used as biomarkers for cancer molecular imaging. In this work, we demonstrated effective MR cancer molecular imaging with a small molecular peptide targeted Gd-DOTA monoamide complex as a targeted MRI contrast agent specific to clotted plasma proteins in tumor stroma. We performed the experiment of evaluating the effectiveness of the agent for non-invasive detection of prostate tumor with MRI in a mouse orthotopic PC-3 prostate cancer model. The targeted contrast agent was effective to produce significant tumor contrast enhancement at a low dose of 0.03 mmol Gd/kg. The peptide targeted MRI contrast agent is promising for MR molecular imaging of prostate tumor.

To demonstrate MR cancer molecular imaging with a small peptide targeted MRI contrast agent specific to clotted plasma proteins in tumor stroma in a mouse prostate cancer model.

前列腺癌与小分子肽CLT1靶向造影剂的磁共振分子成像

Tumor extracellular  matrix has abundance of cancer related proteins that can be used as biomarkers  for cancer molecular imaging. In this work,  we ...

医学

Biofunctionalized Prussian Blue Nanoparticles for Multimodal Molecular Imaging Applications

Multimodal, molecular imaging allows the visualization of biological processes at cellular, subcellular, and molecular-level resolutions using multiple, complementary imaging techniques. These imaging agents facilitate the real-time assessment of pathways and mechanisms in vivo, which enhance both diagnostic and therapeutic efficacy. This article presents the protocol for the synthesis of biofunctionalized Prussian blue nanoparticles (PB NPs) - a novel class of agents for use in multimodal, molecular imaging applications. The imaging modalities incorporated in the nanoparticles, fluorescence imaging and magnetic resonance imaging (MRI), have complementary features. The PB NPs possess a core-shell design where gadolinium and manganese ions incorporated within the interstitial spaces of the PB lattice generate MRI contrast, both in T1 and T2-weighted sequences. The PB NPs are coated with fluorescent avidin using electrostatic self-assembly, which enables fluorescence imaging. The avidin-coated nanoparticles are modified with biotinylated ligands that confer molecular targeting capabilities to the nanoparticles. The stability and toxicity of the nanoparticles are measured, as well as their MRI relaxivities. The multimodal, molecular imaging capabilities of these biofunctionalized PB NPs are then demonstrated by using them for fluorescence imaging and molecular MRI in vitro.

This protocol describes the synthesis of biofunctionalized Prussian blue nanoparticles and their use as multimodal, molecular imaging agents. The nanoparticles have a core-shell design where gadolinium or manganese ions within the nanoparticle core generate MRI contrast. The biofunctional shell contains fluorophores for fluorescence imaging and targeting ligands for molecular targeting.

Biofunctionalized普鲁士蓝纳米粒子多式联运分子成像应用

Multimodal, molecular imaging allows the visualization of biological processes at cellular, subcellular, and molecular-level resolutions using multiple, ...

生物工程

Custom-designed Laser-based Heating Apparatus for Triggered Release of Cisplatin from Thermosensitive Liposomes with Magnetic Resonance Image Guidance

Liposomes have been employed as drug delivery systems to target solid tumors through exploitation of the enhanced permeability and retention (EPR) effect resulting in significant reductions in systemic toxicity. Nonetheless, insufficient release of encapsulated drug from liposomes has limited their clinical efficacy. Temperature-sensitive liposomes have been engineered to provide site-specific release of drug in order to overcome the problem of limited tumor drug bioavailability. Our lab has designed and developed a heat-activated thermosensitive liposome formulation of cisplatin (CDDP), known as HTLC, to provide triggered release of CDDP at solid tumors. Heat-activated delivery in vivo was achieved in murine models using a custom-built laser-based heating apparatus that provides a conformal heating pattern at the tumor site as confirmed by MR thermometry (MRT). A fiber optic temperature monitoring device was used to measure the temperature in real-time during the entire heating period with online adjustment of heat delivery by alternating the laser power. Drug delivery was optimized under magnetic resonance (MR) image guidance by co-encapsulation of an MR contrast agent (i.e.,&#160;gadoteridol) along with CDDP into the thermosensitive liposomes as a means to validate the heating protocol and to assess tumor accumulation. The heating protocol consisted of a preheating period of 5 min prior to administration of HTLC and 20 min heating post-injection. This heating protocol resulted in effective release of the encapsulated agents with the highest MR signal change observed in the heated tumor in comparison to the unheated tumor and muscle. This study demonstrated the successful application of the laser-based heating apparatus for preclinical thermosensitive liposome development and the importance of MR-guided validation of the heating protocol for optimization of drug delivery.

AN MRI-compatible&#160;custom-designed laser-based heating apparatus has been developed to provide local heating of subcutaneous tumors in order to activate release of agents from thermosensitive liposomes specifically at the tumor region.

定制设计的基于激光的加热装置顺铂触发释放的热敏脂质体与磁共振成像制导

Liposomes have been employed as drug delivery systems to target solid tumors through exploitation of the enhanced permeability and retention (EPR) effect ...

Magnetic Levitation Coupled with Portable Imaging and Analysis for Disease Diagnostics

Currently, many clinical diagnostic procedures are complex, costly, inefficient, and inaccessible to a large population in the world. The requirements for specialized equipment and trained personnel require that many diagnostic tests be performed at remote, centralized clinical laboratories. Magnetic levitation is a simple yet powerful technique and can be applied to levitate cells, which are suspended in a paramagnetic solution and placed in a magnetic field, at a position determined by equilibrium between a magnetic force and a buoyancy force. Here, we present a versatile platform technology designed for point-of-care diagnostics which uses magnetic levitation coupled to microscopic imaging and automated analysis to determine the density distribution of a patient's cells as a useful diagnostic indicator. We present two platforms operating on this principle: (i) a smartphone-compatible version of the technology, where the built-in smartphone camera is used to image cells in the magnetic field and a smartphone application processes the images and to measures the density distribution of the cells and (ii) a self-contained version where a camera board is used to capture images and an embedded processing unit with attached thin-film-transistor (TFT) screen measures and displays the results. Demonstrated applications include: (i) measuring the altered distribution of a cell population with a disease phenotype compared to a healthy phenotype, which is applied to sickle cell disease diagnosis, and (ii) separation of different cell types based on their characteristic densities, which is applied to separate white blood cells from red blood cells for white blood cell cytometry. These applications, as well as future extensions of the essential density-based measurements enabled by this portable, user-friendly platform technology, will significantly enhance disease diagnostic capabilities at the point of care.

We present a magnetic levitation technique coupled with automated imaging and analysis in both a smartphone-compatible device and a device with embedded imaging and processing. This is applied to measure the density distribution of cells with two demonstrated biomedical applications: sickle cell disease diagnosis and separating white and red blood cells.

磁悬浮加上便携式成像与分析诊断疾病

Currently, many clinical diagnostic procedures are complex, costly, inefficient, and inaccessible to a large population in the world. The requirements for ...

<meta charset="utf8"></head><body>用于检测新型铁螯合剂的荧光检测

Quantifiable and Inexpensive Cell-Free Fluorescent Method to Confirm the Ability of Novel Compounds to Chelate Iron

Cancer cells require large amounts of iron to maintain their proliferation. Iron metabolism is considered a hallmark of cancer, making iron a valid target for anti-cancer approaches. The development of novel compounds and the identification of leads for further modification requires that proof of mechanism assays be carried out. There are many assays to evaluate the impact on proliferation; however, the ability to chelate iron is an important and sometimes overlooked end-point measure due to the high costs of equipment and the challenge to quickly and reproducibly quantify the strength of chelation. Here, we describe a quantifiable and inexpensive cell-free fluorescent method to confirm the ability of novel compounds to chelate iron. Our assay relies on the commercially available inexpensive fluorescent dye Calcein, whose fluorescence can be quantified on most fluorescent microtiter plate readers. Calcein is a weak iron chelator, and its fluorescence is quenched when it binds Fe2+/3+; fluorescence is restored when a novel chelator outcompetes Calcein for bound Fe2+/3+. The removal of fluorescent quenching and the resulting increase in fluorescence allows the chelation ability of a novel putative chelator to be determined. Therefore, we offer an inexpensive, high-throughput assay that allows the rapid screening of novel candidate chelator compounds.

<meta charset="utf8"></head><body>作者聚焦:评估新型铁螯合剂对癌细胞代谢的影响

We describe a fluorescent assay that can quickly and inexpensively confirm the ability of novel compounds to chelate iron. The assay measures the ability of compounds to outcompete the iron binding activity of the weak iron chelating fluorescent probe Calcein, resulting in a quantifiable increase in fluorescence when chelation occurs.

可量化且廉价的无细胞荧光方法，用于确认新型化合物螯合铁的能力 

Cancer cells require large amounts of iron to maintain their proliferation. Iron metabolism is considered a hallmark of cancer, making iron a valid target ...

癌症研究

Automated Preparation of [68Ga]Ga-3BP-3940 on a Synthesis Module for PET Imaging of the Tumor Microenvironment

A fast, efficient method has been developed on the GAIA synthesis module for automated gallium-68 radiolabeling of 3BP-3940, a molecular imaging probe targeting the fibroblast activation protein for positron emission tomography imaging of the tumor microenvironment. The reaction conditions involved acetate buffer (final concentration: 0.1 M), methionine as an anti-radiolysis agent (final concentration: 5.4 mg/mL), and 30 &#181;g of 3BP-3940, with heating for 8 min at 98 &#176;C. A final purification step on a C18 cartridge was necessary to obtain a radiolabeled product of high purity. In contrast, the generator-produced 68Ga was used directly without a concentration step on a cation exchange cartridge. The production of three validation batches confirmed the method&#39;s reliability, allowing the synthesis of [68Ga]Ga-3BP-3940 in 22.3 &#177; 0.6 min with high radiochemical purity (RCP), as determined by both radio-HPLC (99.1% &#177; 0.1%) and radio-TLC (99.2% &#177; 0.1%). The average radiochemical yield, based on RCP values measured by radio-HPLC, was 74.4% &#177; 3.3%. The stability of the radiolabeled product was demonstrated for up to 4 h after preparation. This protocol provides a reliable, rapid, and efficient methodology for the preparation of [68Ga]Ga-3BP-3940, which can easily be transposed to a clinical setting.

This research describes the automated process for [68Ga]Ga-3BP-3940 production with the GAIA V2 synthesizer, for PET imaging of fibroblast activation protein. The results of quality control tests performed on three test batches are also presented.

A fast, efficient method has been developed on the GAIA synthesis module for automated gallium-68 radiolabeling of 3BP-3940, a molecular imaging probe ...

对于未螯合钆基于核酸适配体传感器（III）

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此视频中的章节

在EOB-DTPA和镓（三）复杂的调查其

制备及

前列腺癌与小分子肽CLT1靶向造影剂的磁共振分子成像

Biofunctionalized普鲁士蓝纳米粒子多式联运分子成像应用

定制设计的基于激光的加热装置顺铂触发释放的热敏脂质体与磁共振成像制导

磁悬浮加上便携式成像与分析诊断疾病

可量化且廉价的无细胞荧光方法，用于确认新型化合物螯合铁的能力

可量化且廉价的无细胞荧光方法，用于确认新型化合物螯合铁的能力

可量化且廉价的无细胞荧光方法，用于确认新型化合物螯合铁的能力

Automated Preparation of [⁶⁸Ga]Ga-3BP-3940 on a Synthesis Module for PET Imaging of the Tumor Microenvironment