作者感谢H2020资助项目的财政支持：ACEnano（赠款协议号720952）。这项工作还部分得到了韩国贸易、工业和能源部资助的国际合作研发方案（赠款号N05310009，"Horizon2020 Kor-EU合作研发ACEnano工具箱"），使韩国合作伙伴能够参与 Horizon 2020 ACEnano 项目财团。

此处描述的方法用于第三轮实验室间比较。
1. 样品准备
<ol><li>通过 0.02 μm 注射器过滤器过滤水。水过滤是必要的，以消除任何污染颗粒之前，使用它的样品稀释。</li>
	<li>要分析新鲜制备的样品，请在过滤的超纯水中稀释 60 nm 黄金胶体分散体积的 50 倍。NTA 分析的建议浓度为每 mL 1 x 107 - 1 x10 9 颗粒。</li>
</ol>2. 执行测量
<ol><li>打开系统<ol>
<li>连接 NTA 仪器、注射器泵和计算机。打开硬件和软件。相关软件（见 材料表）可确保所有硬件通信运行，并显示实时温度读出。</li>
		<li>从 NTA 中取出激光模块，使用组织和压缩空气完全干燥玻璃表面和低体积流动电池 （LVFC） 内部通道、管子和流体端口。</li>
	</ol>
</li>
	<li>撬动管子<ol>
<li>用超纯水冲洗入口流体管，以去除任何颗粒，减少气泡干扰测量的可能性。对于冲洗，仪器外壳内的进水管末端放置在一个废物容器中。</li>
		<li>将过滤水的 1 mL 注射器（无针头）插入 Luer 端口，并尽可能快地将约 900μL 的液体推入进水管。离开装有剩余液体的注射器，以防止任何回音。</li>
	</ol>
</li>
	<li>注射器泵管连接<ol>
<li>将LVFC组装到激光模块上，创建 图1中看到的样品室。将出口管连接到 LVFC 的右侧端口。 注：入口和出口管的直径不同，入口直径小于出口。交换入出口管连接可能会导致对流电池施加压力和泄漏。</li>
		<li>将注射器与入入口管断开，换上含有 1 mL 过滤水的新注射器，确保液体与液体的接触。将入口管连接到 LVFC 的左端口。慢慢将约500μL的液体引入样品室。请注意，确保装载过程中不会引入气泡。最终的管子配置显示在 图 2中。</li>
	</ol>
</li>
</ol><img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/61741/61741fig1v2.jpg"> 图1：安装在激光模块上的低体积流细胞组件。 
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/61741/61741fig02.jpg"> 图2：低体积流管配置。<a href="https://www.jove.com/files/ftp_upload/61741/61741fig02large.jpg" target="_blank">请点击这里查看此数字的较大版本。</a>
<ol start="4"><li>激光模块加载和系统检查<ol>
<li>将充满水的LVFC插入激光模块，然后锁定到位。</li>
		<li>将注射器放入注射器泵摇篮并牢固。通过单击软件界面中的 "启动相机"来 初始化摄像机。在界面的 硬件 选项卡中，单击 散射 以移动参考位置。</li>
		<li>将相机水平设置为 16，并手动调整对焦，以检查任何颗粒的 diluent。通过左键单击主查看窗口并使用鼠标上下拖动以检查任何粒子来调整视场位置。如果视野中有三个以上的颗粒，这意味着净水或清洁过程存在问题，因此，清洁过程需要重复，或者需要更换或过滤水。</li>
		<li>从仪器中取出激光模块。</li>
		<li>将注射器与入口管断开，仅用充满空气的注射器更换。慢慢地将空气引入样品室，以去除里面的液体。从激光模块中取出 LVFC 并断开管子。用水清洁激光模块的LVFC玻璃表面和光学玻璃，用纸巾和压缩空气干燥。用压缩空气干燥管子。将 LVFC 重新组装到激光模块上并连接管子，以便进行样品加载。 注：此步骤并不总是需要的，但是，在这种情况下，它被添加为额外的预防措施，以进一步减少任何可能的变化。</li>
	</ol>
</li>
	<li>加载样本<ol>
<li>重复步骤2.2.2。将第 1.1 步中分散的 60 nm 金纳米粒子中的 1 mL 的注射器连接到 Luer 端口。通过入口管将样品的 750 μL 缓慢注入 LVFC，并在仪器外观看激光模块，以确保不会引入气泡。</li>
		<li>将激光模块加载回 NTA 仪器，然后通过单击软件界面中的 "启动相机"来 初始化相机。在界面的 硬件 选项卡中，单击 散射 以移动到参考对焦位置，检查是否正确设置以提供粒子的清晰图像。</li>
		<li>检查视场是否与激光束位置集中设置。通过左键单击软件中的主要查看窗口和鼠标上下拖动来相应地进行调整。</li>
		<li>运行 自动设置 功能，自动优化对焦和摄像机水平，确保实现最佳图像质量。 注：自动摄像头和对焦参数允许不同实验室之间更加一致，因为这是用户独立。</li>
	</ol>
</li>
	<li>样本分析<ol>
<li>在标准测量（SOP选项卡）中创建一个测量脚本，以获得 5 个 60 年代慢速下的重复视频（粒子应在大约 10 s 时从屏幕的一侧传到另一侧）和恒定流（补充文件 1）。 注：建议流，以确保更好地表示整体样本的测量。当样品上传递缓慢的流时，浓度测量的精度和可重复性显著提高，以确保更多的新粒子流经测量区并在实验期间进行分析。视频长度取决于配置文件分布以及分析时间的可变性。60s 的 5 个视频被视为典型的测量持续时间。</li>
		<li>为数据设置实验文件名称和位置并开始运行。概述程序之后的分析是由地平线2020 ACEnano项目17的七个实验室进行的。</li>
	</ol>
</li>
</ol>3. 数据分析
注：所有数据分析均在 v 3.4 软件内完成（见 材料表），不使用其他手动转换或计算。粒子大小数据以原始形式作为直方图分布呈现，并使用斯托克斯-爱因斯坦方程从粒子位置的测量变化中计算。该软件确定 x 和 y 平面中每个粒子移动的平均距离。此值允许确定粒子扩散系数 （D），如果已知样品温度 T 和溶剂粘度η，则可以计算粒子的等效球形流动力学半径 RH。样本的温度由 NTA 自动记录。软件使用的默认样品粘度为水，并包含在上面所示的测量脚本中，但当在测量之前或之后使用不同的样品 diluent 时，用户可以修改粘度。
<ol><li>通过拖动滑块条或在检测阈值下单击软件中的 + 和 按钮 来设置 检测阈值（DT），这是在 2 到 20 之间对可视化粒子进行最佳跟踪的分析参数。确保所选的DT值识别和跟踪尽可能多的可见粒子（在软件图像屏幕上自动标记为红色十字）。<ol>
<li>作为设置检测阈值的指导，图像中已识别的粒子数应在大约 30-80 之间，其中不超过 10 个红十字应与观察者不被视为粒子的站点相对应。观察到的蓝色十字架不应超过 5 个（表示噪音）。</li>
	</ol>
</li>
</ol><img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/61741/61741fig03.jpg"> 图3：阈值设置观察。坏（左）和好（右）检测阈值设置观察。 <a href="https://www.jove.com/files/ftp_upload/61741/61741fig03large.jpg" target="_blank">请点击这里查看此数字的较大版本。</a>
<ol start="2"><li>通过按下软件中的 "过程 "按钮自动处理粒子跟踪分析视频。将所有处理参数设置为自动并导出数据，作为.csv格式结果文件，并附上完整的粒子大小分布和描述测量设置的其他元数据。要验证测量质量，请查看软件中的 分析 选项卡或检查.csv输出文件以查看任何警告消息或警报。PDF 结果报告的示例显示在 补充文件 2 中。
</li>
	<li>阅读模式大小结果和与 PDF 报告相关的标准偏差。 注：模式大小结果用于比较在七个实验室中获得的大小，并在第 5 节中显示和讨论。</li>
</ol>4. 清洁和干燥
<ol><li>使用后，用干净的水彻底冲洗系统，以清除管道和光学表面的所有样品痕迹。通过观察视野中的颗粒量，可以监测清洁效果。</li>
	<li>从 NTA 仪器中取出激光模块。</li>
	<li>通过系统加载空气注射器以清空管道和 LVFC。</li>
</ol>

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作者乔·沙利文、阿格涅什卡·西乌帕、宝琳·卡内尔-莫里斯和米歇尔·卡比尼是马尔文泛光有限公司的员工，该公司生产本文中使用的仪器。

从第 1 轮 ILC 获得的结果不一致，突出表明需要对较旧的系统进行仪器健康检查，以及开发更详细的 SOP、需要进行实践培训以及更好地了解测量和分析设置，以确保不同实验室的结果更加一致。事实上，Hole等人发现，缺乏关于如何使用NTA系统和准备样品的共享指南，导致实验室的变异性，即使是相对单分散的样品16。因此，所有ILC参与者都参加了一个培训讲习班，讲习班涵盖系统操作和测量条件的最佳做法，以及特定NTA仪器的清洁和维护指导。所有参与者还在自己的实验室中用同一仪器进行测量，以进行随后的ILC回合。该过程首先涉及在每个实验室中通过在乳胶标准样品（ILC 第 2 轮）上运行 ILC 对系统进行本地测试的一轮测试，然后由合作伙伴用于重复黄金测量（ILC 第 3 轮）。通过 NTA 测量这些黄金样品的目的是在纳米材料风险评估方法和做法中引入信心和清晰度，以影响纳米安全指导协议。
NTA是一种技术，可以测量粒子的流体球当量直径，并可用于粒子粒子，实时视觉分析的多分散系统范围从10 nm - 50nm，到约1000nm的大小（取决于样本属性和仪器配置）。需要最少的样品准备。尽管样品准备最少，但这一步骤对协议至关重要，在稀释样品和选择稀释剂时应小心谨慎。形状可能是NTA的一个限制因素，因为球形等效大小测量获得，非球形粒子将有一个不太准确的大小值。
对于 NTA 技术，由于仅从整个样本中观察到具有代表性的样本，因此始终预期会有一些结果变异。无论如何，所有结果都符合ISO 19430粒子尺寸标准。提供的最佳浓度通常在 30-60 秒分析时间内约为 108 颗粒/毫升。对于颗粒浓度较低的样品，需要更长的分析时间，以确保可重复的结果。对于颗粒浓度大于 109 颗粒/mL 的样品，跟踪问题的可能性更大，需要稀释到适合 NTA 测量的范围。
总体而言，第三轮ILC的结果表明，通过NTA，金纳米粒子测量具有良好的可重复性，提高了准确性和可重复性。所有 NTA 测量均使用自动摄像机级别和对焦设置进行，以调整图像，这是由软件中的自动设置功能选择的。该软件设置的摄像机级别非常一致，在所有情况下设置的摄像机级别为 10 或 11，这表明，正如预期的那样，过程包含的自动化程度越高，实现的一致性就越高。尺寸结果与制造商通过 TEM 获得的量身定做结果相当，表明其结果可重复，但由于 TEM 无法确定水动力直径，预计不同技术的差异将微乎其微。结果一致性显著提高，表明仪器维护、详细 SOP、最佳实践更新、用户知识和 NTA 应用测量自动化的重要性和益处。最后，ILC 验证了这一特定的 NTA SOP，从而为这种特定的纳米材料风险评估方法带来了信心和清晰度。

纳米材料（NMs）在物理和化学特性上都可能有所不同，进而影响其行为、稳定性和毒性1、2、3、4、5。在深入了解NM特性、危害和行为时，主要困难之一是能够获得有关物理和化学纳米材料特征的可重复信息。这些物理特性的例子包括粒子大小和大小分布6，7，8。这些都是重要的参数，因为它们是欧盟委员会（EC）对"纳米"9一词定义的一个关键方面。
除了了解NMs6、10的命运和毒性影响外，实现精确的粒子尺寸测量对于许多不同的工业和研究应用和过程也至关重要。重要的是要有成熟的方法，能够准确，可靠和可重复地测量NM的大小。此外，所报告的信息应深入了解所使用的技术，例如，指示大小参数的类型（例如实际大小或水动力学大小）以及样本条件，例如存在 NM 的特定介质，以及用于在不同介质中可靠执行的方法。为了测量尺寸，可以使用许多技术，包括电子显微镜（EM）、动态光散射（DLS）、单粒子感应耦合等离子体质谱仪（SPICP-MS）、差分离心沉降（DCS）、扫描探针显微镜（SPM）、小角度X射线散射（SAXS）和纳米粒子跟踪分析（NTA）。
NTA 是一项相对较新的技术，近年来取得了长足的进步，并已被证明能够可靠地测量复杂水态介质（如与环境相关的技术，例如淡水系统）中球形 NM 的水动力学直径。水动力学直径是"一个假想的硬球体的大小，以与所测粒子相同的方式扩散"11：在实际操作中，在水介质中，这描述了一个直径大于粒子本身的直径，其中还包括一层分子（主要是水），由弱静电力保持在粒子表面。粒子的水动力学直径会因不同介质而变化，随着测量粒子的离子强度升高而变小。
NTA 技术的另一个重要特点是，它允许分析员实现数字加权大小测量，这是 EC 纳米材料定义所要求的。高分辨率、逐粒子分析使这种技术在粒子吞吐量为10、12的异质试验样品中出现时，更容易受到聚合物或较大颗粒的干扰。
测量过程包括准备一个合适的悬浮样品，这往往需要样品稀释，然后视频记录粒子的布朗运动行为和视频分析。从样品室中，激光束经过，激光束路径中的悬浮粒子通过装有摄像头的光学显微镜散射光，导致其可视化。摄像机从在布朗运动下移动的粒子中捕捉散射激光的视频文件。许多粒子可以单独跟踪以确定其扩散系数，其水动力学直径可以使用斯托克斯-爱因斯坦方程计算：d = kT/3πηD d 是水动力学直径，k 是博尔茨曼常数，T 是温度，η是粘度，D 是扩散系数10。NTA还可用于跟踪通常胶体不稳定的粒子的聚集行为（然而，粒子必须在测量时间尺度上胶体稳定）13，14。NTA 是一种绝对方法，在这项工作中使用的仪器上不需要系统校准。如果用户想要检查系统性能，可以通过按需要频繁测量尺寸标准材料来轻松完成此任务。
NTA 仪器易于操作，分析时间快（每个样品不到 10 分钟）。对于具有良好数据可重复性和可重复性的高质量测量，在样品准备和仪器操作中应考虑若干因素。如果不仔细考虑这些因素，不同实验室和操作员对同一材料的测量可能会受到未知或未量化的不确定性的影响。在 NP 定性中，使用内部开发的 SOP 的最佳实践并不总是保证与其他实验室的一致性，如 Roebben 等人对 DLS 技术15所示。
事实上，不同实验室、用户和仪器之间的早期（第一轮）NTA ILC显示的结果不一致。其中一个主要问题是使用各种没有定期服务或校准检查的旧遗留文书，以及方法解释的差异。霍尔等人的NTA ILC研究发现，由于缺乏关于如何使用系统和准备样品的共同指南，即使对于相对单一分散的样品16，实验室的变异性也会很大。这以及第一轮 ILC 的结果突出了良好的仪器维护以及方法培训和发达的标准操作程序 （SOP） 的必要性。后者是描述和记录遵守良好做法的有力工具。如果详细，标准操作程序 （SOP） 可以提供清晰、解释、理解、标准化和质量保证。
因此，采用ILC研究的建议非常适合开发和测试协议16。ILC 演习旨在验证这一特定的 NTA SOP，从而为这种特定的纳米材料风险评估方法带来了信心和清晰度。它涉及三轮。第 1 轮在训练前分析了每个参与者自己的仪器上的 60 纳米金纳米粒子。第 2 轮涉及使用具有常见配置的新仪器分析 100 nm 乳胶作为简单测试，以确定仪器设置正确，用户对如何使用仪器有良好的了解。第3轮涉及在训练后对新仪器上60纳米金纳米粒子进行分析，并具有共同的配置。ILC的参与者来自七个不同的实验室，都是地平线2020 ACEnano项目17的财团成员。
本文的目的是讨论 NTA 技术第三轮基准测试的方法和结果，在详细培训和 SOP 开发之后，7 个合作伙伴重新分析了 60 nm 黄金 NP。还将比较和参考在ILC第一轮中取得的结果。ILC 第 3 轮的所有分析均使用配备 405 nm 激光器和高灵敏度 sCMOS 摄像机的相同配置的相同仪器（见 材料表）进行。基准评估技术在样品上的性能，从而导致"最佳实践"协议的制定。因此，本文还分享并使本ILC中使用的仪器的NTA方法可供科学界使用，因为它通过按照国际标准对ILC进行和评估而得到协调。

<table>
	<tbody>
		<tr>
			<td>60 nm gold colloid dispersion</td>
			<td>BBI Solutions OEM Ltd.</td>
			<td>Product EM. GC60, Batch number 024650</td>
			<td></td>
		</tr>
		<tr>
			<td>0.02 &#181;m syringe filter - Whatman Anotop 25 Sterile Syringe Filters</td>
			<td>Sigma Aldrich</td>
			<td>WHA68092102</td>
			<td></td>
		</tr>
		<tr>
			<td>NanoSight</td>
			<td>Malvern Panalytical Ltd.</td>
			<td>NS300</td>
			<td></td>
		</tr>
		<tr>
			<td>NanoSight NTA Software v3.4</td>
			<td>Malvern Panalytical Ltd.</td>
			<td>v3.4</td>
			<td></td>
		</tr>
		<tr>
			<td>Syringe PP/PE without needle luer slip tip, centered, capacity 1&#160;mL, graduated, 0.01&#160;mL, sterile</td>
			<td>Sigma Aldrich</td>
			<td>Z230723</td>
			<td></td>
		</tr>
	</tbody>
</table>

nanoparticle tracking analysis of gold nanoparticles in aqueous media through an inter-laboratory comparison

在纳米技术领域，分析性特征在理解纳米材料（NMs）的行为和毒性方面起着至关重要的作用。特征化需要彻底，所选择的技术应非常适合要确定的属性、所分析的材料和存在的介质。此外，仪器操作和方法需要得到用户的充分开发和明确理解，以避免数据收集错误。应用方法或程序的任何差异都可能导致已获得数据的差异和可重复性差。本文旨在通过纳米粒子跟踪分析（NTA）来阐明测量金纳米粒子水动力学直径的方法。这项研究是在七个不同的实验室之间作为实验室间比较（ILC）进行的，以验证标准操作程序的性能和可重复性。ILC 研究的结果揭示了详细标准操作程序 （SOP）、最佳实践更新、用户知识和测量自动化的重要性和益处。

使用各种 NTA 仪器配置的第 1 轮 ILC 结果显示在 图 4中。除实验室 6 外，5 个捕获重复之间的可重复性良好，但多个实验室记录的模式大小高于预期。实验室 6 结果表明，可重复性差，测量的模式大小要高得多。调查后发现，报告最大尺寸变化的系统要么没有按照建议进行维护，要么分析受到样品制备不一致的影响，即稀释步骤可能因不同的管道设备、用户操作和技术以及/或测量设置（包括流细胞不干净、使用错误的摄像机水平、图像未正确对焦点以及设置分析检测阈值）而产生变异。
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/61741/61741fig4v2.jpg"> 图4：ILC第1轮模式大小结果。模式大小来自所有 NTA 基准合作伙伴在不同的 NTA 仪器上进行的第 1 轮 60 nm 金纳米粒子分散（如 x 轴的缩写）。
实施相同 SOP 和仪器设置的所有实验室都提高了第 3 轮的 NTA 结果精度。此 ILC 第 3 轮获得的模式大小结果可在 图 5中看到。所有实验室的平均模式为 62.02 ± 1.97 nm。第 3 轮的所有测量结果都比第一阶段的结果更一致，结果远远低于制造商所述批次 60.5 nm 平均尺寸的 10%。制造商陈述的黄金样品变异系数为≤8%。
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/61741/61741fig5v2.jpg"> 图5：ILC第3轮模式大小结果。模式大小来自所有 NTA 基准合作伙伴的结果，用于在同一 NTA 仪器上分析的 60 nm 黄金 ILC 第 3 轮。所有实验室的平均模式为 62.02 ± 1.97 nm。
为了验证粒子大小，根据制造商提供，传输电子显微镜 （TEM） 分析了少量 （N=82） 粒子。大约 10μl 的未浸出分散物被投在碳涂层 Cu TEM 网格上，在 200 kV 的分析 TEM 成像之前在空气中干燥。像 补充图1 这样的图像取自粒子重叠最小的区域，并使用半自动图像分析过程进行分析。一种自动分水岭方法应用于分离粒子，这个过程的人工制品被排除在外，以及边缘粒子18。平均直径的计算方法是主轴和次小轴的平均值（61±7 nm），或假设球形粒子从测量区域（62±6nm）的转换。粒子似乎大部分是球形的，平均纵横比为1.1。TEM 结果显示直径略高于制造商值 （60.5 nm），但处于公差水平。此外，与 NTA 的流体动力学直径推导值有非常好的协议。
补充图1：60纳米金纳米粒子的TEM图像。<a href="https://cloudflare.jove.com/files/ftp_upload/61741/61741supfig01.jpg" target="_blank">请点击这里下载此数字。</a>
补充文件1：测量脚本。<a href="https://cloudflare.jove.com/files/ftp_upload/61741/Measurement_script.docx" target="_blank">请点击这里下载此文件。</a>
补充文件2：PDF结果报告示例。<a href="https://cloudflare.jove.com/files/ftp_upload/61741/PDF_results_report.pdf" target="_blank">请点击这里下载此文件。</a>

Watch this Scientific Journal Video about Nanoparticle Tracking Analysis of Gold Nanoparticles in Aqueous Media through an Inter-Laboratory Comparison at JoVE.com

Nanoparticle Tracking Analysis of Gold Nanoparticles in Aqueous Media through an Inter-Laboratory Comparison

此处描述的协议旨在通过纳米粒子跟踪分析 （NTA） 在水介质中测量球形纳米粒子（更具体地说是金纳米粒子）的水动力学直径。后者包括跟踪由于布朗运动而产生的粒子的运动，并实施斯托克斯-爱因斯坦方程来获得水动力学直径。

通过实验室间比较，对液态介质中的金纳米粒子进行纳米粒子跟踪分析

In the field of nanotechnology, analytical characterization plays a vital role in understanding the behavior and toxicity of nanomaterials (NMs). ...

nanoparticle-tracking-analysis-gold-nanoparticles-aqueous-media

Research

JoVE Journal

Chemistry

7.2K 次观看.  University of Birmingham. 此处描述的协议旨在通过纳米粒子跟踪分析 （NTA） 在水介质中测量球形纳米粒子（更具体地说是金纳米粒子）的水动力学直径。后者包括跟踪由于布朗运动而产生的粒子的运动，并实施斯托克斯-爱因斯坦方程来获得水动力学直径。

视频: Nanoparticle Tracking Analysis of Gold Nanoparticles in Aqueous Media through an Inter-Laboratory Comparison

Preparation of Silica Nanoparticles Through Microwave-assisted Acid-catalysis

Microwave-assisted synthetic techniques were used to quickly and reproducibly produce silica nanoparticle sols using an acid catalyst with nanoparticle diameters ranging from 30-250 nm by varying the reaction conditions. Through the selection of a microwave compatible solvent, silicic acid precursor, catalyst, and microwave irradiation time, these microwave-assisted methods were capable of overcoming the previously reported shortcomings associated with synthesis of silica nanoparticles using microwave reactors. The siloxane precursor was hydrolyzed using the acid catalyst, HCl. Acetone, a low-tan &#948; solvent, mediates the condensation reactions and has minimal interaction with the electromagnetic field. Condensation reactions begin when the silicic acid precursor couples with the microwave radiation, leading to silica nanoparticle sol formation. The silica nanoparticles were characterized by dynamic light scattering data and scanning electron microscopy, which show the materials&#39; morphology and size to be dependent on the reaction conditions. Microwave-assisted reactions produce silica nanoparticles with roughened textured surfaces that are atypical for silica sols produced by St&#246;ber&#39;s methods, which have smooth surfaces.

Silica nanoparticles were prepared using acid-catalysis of a siloxane precursor and microwave-assisted synthetic techniques resulting in the controlled growth of nanomaterials ranging from 30-250 nm in diameter. The growth dynamics can be controlled by varying the initial silicic acid concentration, time of the reaction, and temperature of reaction.

Microwave-assisted synthetic techniques were used to quickly and reproducibly produce silica nanoparticle sols using an acid catalyst with nanoparticle ...

科研

化学

An Optimized Protocol for the Efficient Radiolabeling of Gold Nanoparticles by Using a 125I-labeled Azide Prosthetic Group

Here, we demonstrate a detailed protocol for the radiosynthesis of a 125I-labeled azide prosthetic group and its application to the efficient radiolabeling of DBCO-group-functionalized gold nanoparticles using a copper-free click reaction. Radioiodination of the stannylated precursor (2) was carried out by using [125I]NaI and chloramine T as an oxidant at room temperature for 15 min. After HPLC purification of the crude product, the purified 125I-labeled azide (1) was obtained with high radiochemical yield (75 &plusmn; 10%, n = 8) and excellent radiochemical purity (&gt;99%). For the synthesis of radiolabeled 13-nm-sized gold nanoparticles, the DBCO-functionalized gold nanoparticles (3) were prepared by using a thiolated polyethylene glycol polymer. A copper-free click reaction between 1 and 3 gave the 125I-labeled gold nanoparticles (4) with more than 95% of radiochemical yield as determined by radio-thin-layer chromatography (radio-TLC). These results clearly indicate that the present radiolabeling method using a strain-promoted copper-free click reaction will be useful for the efficient and convenient radiolabeling of DBCO-group-containing nanomaterials.

An Optimized Protocol for the Efficient Radiolabeling of Gold Nanoparticles by Using a 125I-labeled Azide Prosthetic Group

A detailed procedure for the synthesis of a 125I-labeled azide and the radiolabeling of dibenzocyclooctyne (DBCO)-group-conjugated, 13-nm-sized gold nanoparticles using a copper-free click reaction is described.

一个优化的协议通过使用金纳米粒子的放射性标记效率<sup&gt; 125</sup&gt;我标记的叠氮辅基

Here, we demonstrate a detailed protocol for the radiosynthesis of a 125I-labeled azide prosthetic group and its application to the efficient ...

Liquid-cell Transmission Electron Microscopy for Tracking Self-assembly of Nanoparticles

Drying a nanoparticle dispersion is a versatile way to create self-assembled structures of nanoparticles, but the mechanism of this process is not fully understood. We have traced the trajectories of individual nanoparticles using liquid-cell transmission electron microscopy (TEM) to investigate the mechanism of the assembly process. Herein, we present the protocols used for liquid-cell TEM studies of the self-assembly mechanism. First, we introduce the detailed synthetic protocols used to produce uniformly sized platinum and lead selenide nanoparticles. Next, we present the microfabrication processes used to produce liquid cells with silicon nitride or silicon windows and then describe the loading and imaging procedures of the liquid-cell TEM technique. Several notes are included to provide helpful tips for the entire process, including how to manage the fragile cell windows. The individual motions of nanoparticles tracked by liquid-cell TEM revealed that changes in the solvent boundaries caused by evaporation affected the self-assembly process of nanoparticles. The solvent boundaries drove nanoparticles to primarily form amorphous aggregates, followed by flattening of the aggregates to produce a 2-dimensional (2D) self-assembled structure. These behaviors are also observed for different nanoparticle types and different liquid-cell compositions.

Here we introduce experimental protocols for the real-time observation of a self-assembly process using liquid-cell transmission electron microscopy.

用于跟踪纳米粒子自组装的液体细胞透射电子显微镜

Drying a nanoparticle dispersion is a versatile way to create self-assembled structures of nanoparticles, but the mechanism of this process is not fully ...

Rapid Nanoprobe Signal Enhancement by In Situ Gold Nanoparticle Synthesis

The use of nanoprobes such as gold, silver, silica or iron-oxide nanoparticles as detection reagents in bioanalytical assays can enable high sensitivity and convenient colorimetric readout. However, high densities of nanoparticles are typically needed for detection. The available synthesis-based enhancement protocols are either limited to gold and silver nanoparticles or rely on precise enzymatic control and optimization. Here, we present a protocol to enhance the colorimetric readout of gold, silver, silica, and iron oxide nanoprobes. It was observed that the colorimetric signal can be improved by up to a 10000-fold factor. The basis for such signal enhancement strategies is the chemical reduction of Au3+ to Au0. There are several chemical reactions that enable the reduction of Au3+ to Au0. In the protocol, Good&#39;s buffers and H2O2 are used and it is possible to favor the deposition of Au0 onto the surface of existing nanoprobes, in detriment of the formation of new gold nanoparticles. The protocol consists of the incubation of the microarray with a solution consisting of chloroauric acid and H2O2 in 2-(N-morpholino)ethanesulfonic acid pH 6 buffer following the nanoprobe-based detection assay. The enhancement solution can be applied to paper and glass-based sensors. Moreover, it can be used in commercially available immunoassays as demonstrated by the application of the method to a commercial allergen microarray. The signal development requires less than 5 min of incubation with the enhancement solution and the readout can be assessed by naked eye or low-end image acquisition devices such as a table-top scanner or a digital camera.

Rapid Nanoprobe Signal Enhancement by In Situ Gold Nanoparticle Synthesis

In this work, a protocol for signal enhancement of nanoprobe-based biosensing is presented. The protocol is based on the reduction of chloroauric acid onto the surface of existing nanoprobes that consist of gold, silver, silica or iron-oxide nanoparticles.

原位金纳米粒子合成快速 Nanoprobe 信号增强

The use of nanoprobes such as gold, silver, silica or iron-oxide nanoparticles as detection reagents in bioanalytical assays can enable high sensitivity ...

Heterogeneous Removal of Water-Soluble Ruthenium Olefin Metathesis Catalyst from Aqueous Media Via Host-Guest Interaction

A highly efficient transition metal catalyst removal method is developed. The water-soluble catalyst contains a newly-designed NHC ligand for the catalyst removal via host-guest interactions. The new NHC ligand possesses an adamantyl (guest) tethered linear ethylene glycol units for hydrophobic inclusion into the cavity of a &#946;-cyclodextrin (&#946;-CD) host compound. The new NHC ligand was applied to a Ru-based olefin metathesis catalyst. The Ru catalyst demonstrated excellent activity in representative ring-closing metathesis (RCM) and ring-opening metathesis polymerization (ROMP) reactions in aqueous media as well as organic solvent, CH2Cl2. After the reaction was complete, the lingering Ru residue was removed from the aqueous solution with the efficiency of more than 99% (53 ppm of Ru residue) by simple filtration utilizing a host-guest interaction between insoluble silica-grafted &#946;-CD (host) and the adamantyl moiety (guest) on the catalyst. The new Ru catalyst also demonstrated high removal efficiency via extraction when the reaction is run in organic solvent by partitioning the crude reaction mixture between layers of diethyl ether and water. In this way, the catalyst stays in aqueous layer only. In organic layer, the residual Ru amount was only 0.14 ppm in the RCM reactions of diallyl compounds.

A removable water-soluble N-heterocyclic carbene (NHC) ligand in aqueous media via host-guest interaction has been developed. We demonstrated representative olefin metathesis reactions in water as well as in dichloromethane. Via either host-guest interaction or extraction, the residual ruthenium (Ru) catalyst was as low as 0.14 ppm after the reaction.

水介质中水溶性钌烯烃置换催化剂的异质化去除与寄主交互作用

A highly efficient transition metal catalyst removal method is developed. The water-soluble catalyst contains a newly-designed NHC ligand for the catalyst ...

Synthesis and Characterization of Amphiphilic Gold Nanoparticles

Gold nanoparticles covered with a mixture of 1-octanethiol (OT) and 11-mercapto-1-undecane sulfonic acid (MUS) have been extensively studied because of their interactions with cell membranes, lipid bilayers, and viruses. The hydrophilic ligands make these particles colloidally stable in aqueous solutions and the combination with hydrophobic ligands creates an amphiphilic particle that can be loaded with hydrophobic drugs, fuse with the lipid membranes, and resist nonspecific protein adsorption. Many of these properties depend on nanoparticle size and the composition of the ligand shell. It is, therefore, crucial to have a reproducible synthetic method and reliable characterization techniques that allow the determination of nanoparticle properties and the ligand shell composition. Here, a one-phase chemical reduction, followed by a thorough purification to synthesize these nanoparticles with diameters below 5 nm, is presented. The ratio between the two ligands on the surface of the nanoparticle can be tuned through their stoichiometric ratio used during synthesis. We demonstrate how various routine techniques, such as transmission electron microscopy (TEM), nuclear magnetic resonance (NMR), thermogravimetric analysis (TGA), and ultraviolet-visible (UV-Vis) spectrometry, are combined to comprehensively characterize the physicochemical parameters of the nanoparticles.

Amphiphilic gold nanoparticles can be used in many biological applications. A protocol to synthesize gold nanoparticles coated by a binary mixture of ligands and a detailed characterization of these particles is presented.

两栖金纳米粒子的合成与表征

Gold nanoparticles covered with a mixture of 1-octanethiol (OT) and 11-mercapto-1-undecane sulfonic acid (MUS) have been extensively studied because of ...

Gold Nanoparticle Modified Carbon Fiber Microelectrodes for Enhanced Neurochemical Detection

For over 30 years, carbon-fiber microelectrodes (CFMEs) have been the standard for neurotransmitter detection. Generally, carbon fibers are aspirated into glass capillaries, pulled to a fine taper, and then sealed using an epoxy to create electrode materials that are used for fast scan cyclic voltammetry testing. The use of bare CFMEs has several limitations, though. First and foremost, the carbon fiber contains mostly basal plane carbon, which has a relatively low surface area and yields lower sensitivities than other nanomaterials. Furthermore, the graphitic carbon is limited by its temporal resolution, and its relatively low conductivity. Lastly, neurochemicals and macromolecules have been known to foul at the surface of carbon electrodes where they form non-conductive polymers that block further neurotransmitter adsorption. For this study, we modify CFMEs with gold nanoparticles to enhance neurochemical testing with fast scan cyclic voltammetry. Au3+ was electrodeposited or dipcoated from a colloidal solution onto the surface of CFMEs. Since gold is a stable and relatively inert metal, it is an ideal electrode material for analytical measurements of neurochemicals. Gold nanoparticle modified (AuNP-CFMEs) had a stability to dopamine response for over 4 h. Moreover, AuNP-CFMEs exhibit an increased sensitivity (higher peak oxidative current of the cyclic voltammograms) and faster electron transfer kinetics (lower &#916;EP or peak separation) than bare unmodified CFMEs. The development of AuNP-CFMEs provides the creation of novel electrochemical sensors for detecting fast changes in dopamine concentration and other neurochemicals at lower limits of detection. This work has vast applications for the enhancement of neurochemical measurements. The generation of gold nanoparticle modified CFMEs will be vitally important for the development of novel electrode sensors to detect neurotransmitters in vivo in rodent and other models to study neurochemical effects of drug abuse, depression, stroke, ischemia, and other behavioral and disease states.

In this study, we modify carbon-fiber microelectrodes with gold nanoparticles to enhance the sensitivity of neurotransmitter detection.

用于增强神经化学检测的金纳米粒子改性碳纤维微电极

For over 30 years, carbon-fiber microelectrodes (CFMEs) have been the standard for neurotransmitter detection. Generally, carbon fibers are aspirated into ...

Synthesis of Aptamer-PEI-g-PEG Modified Gold Nanoparticles Loaded with Doxorubicin for Targeted Drug Delivery

Due to drug resistance and toxicity in healthy cells, use of doxorubicin (DOX) has been limited in clinical cancer therapy. This protocol describes the designing of poly(ethylenimine) grafted with polyethylene glycol (PEI-g-PEG) copolymer functionalized gold nanoparticles (AuNPs) with loaded aptamer (AS1411) and DOX through amide reactions. AS1411 is specifically bonded with targeted nucleolin receptors on cancer cells so that DOX targets cancer cells instead of healthy cells. First, PEG is carboxylated, then grafted to branched PEI to obtain a PEI-g-PEG copolymer, which is confirmed by 1H NMR analysis. Next, PEI-g-PEG copolymer coated gold nanoparticles (PEI-g-PEG@AuNPs) are synthesized, and DOX and AS1411 are covalently bonded to AuNPs gradually via amide reactions. The diameter of the prepared AS1411-g-DOX-g-PEI-g-PEG@AuNPs is ~39.9 nm, with a zeta potential of -29.3 mV, indicating that the nanoparticles are stable in water and cell medium. Cell cytotoxicity assays show that the newly designed DOX loaded AuNPs are able to kill cancer cells (A549). This synthesis demonstrates the delicate arrangement of PEI-g-PEG copolymers, aptamers, and DOX on AuNPs that are achieved by sequential amide reactions. Such aptamer-PEI-g-PEG functionalized AuNPs provide a promising platform for targeted drug delivery in cancer therapy.

In this protocol, doxorubicin-loaded AS1411-g-PEI-g-PEG modified gold nanoparticles are synthesized via three-step amide reactions. Then, doxorubicin is loaded and delivered to target cancer cells for cancer therapy.

合成阿普塔默-PEI-g-PEG改性金纳米粒子装载多索鲁比辛用于靶向药物输送

Due to drug resistance and toxicity in healthy cells, use of doxorubicin (DOX) has been limited in clinical cancer therapy. This protocol describes the ...

Preparation of Nanoparticles for ToF-SIMS and XPS Analysis

Nanoparticles have gained increasing attention in recent years due to their potential and application in different fields including medicine, cosmetics, chemistry, and their potential to enable advanced materials. To effectively understand and regulate the physico-chemical properties and potential adverse effects of nanoparticles, validated measurement procedures for the various properties of nanoparticles need to be developed. While procedures for measuring nanoparticle size and size distribution are already established, standardized methods for analysis of their surface chemistry are not yet in place, although the influence of the surface chemistry on nanoparticle properties is undisputed. In particular, storage and preparation of nanoparticles for surface analysis strongly influences the analytical results from various methods, and in order to obtain consistent results, sample preparation must be both optimized and standardized. In this contribution, we present, in detail, some standard procedures for preparing nanoparticles for surface analytics. In principle, nanoparticles can be deposited on a suitable substrate from suspension or as a powder. Silicon (Si) wafers are commonly used as substrate, however, their cleaning is critical to the process. For sample preparation from suspension, we will discuss drop-casting and spin-coating, where not only the cleanliness of the substrate and purity of the suspension but also its concentration play important roles for the success of the preparation methodology. For nanoparticles with sensitive ligand shells or coatings, deposition as powders is more suitable, although this method requires particular care in fixing the sample.

A number of different procedures for preparing nanoparticles for surface analysis are presented (drop casting, spin coating, deposition from powders, and cryofixation). We discuss the challenges, opportunities, and possible applications of each method, particularly regarding the changes in the surface properties caused by the different preparation methods.

用于ToF-SIMS和XPS分析的纳米颗粒的制备

Nanoparticles have gained increasing attention in recent years due to their potential and application in different fields including medicine, cosmetics, ...

Gold Nanoparticle Synthesis

Gold nanoparticles (Au nanoparticles) that are ~12 nm in diameter were synthesized by rapidly injecting a solution of 150 mg (0.15 mmol) of tetrachloroauric acid in 3.0 g (3.7 mmol, 3.6 mL) of oleylamine (technical grade) and 3.0 mL of toluene into a boiling solution of 5.1 g (6.4 mmol, 8.7 mL) of oleylamine in 147 mL of toluene. While boiling and mixing the reaction solution for 2 hours, the color of the reaction mixture changed from clear, to light yellow, to light pink, and then slowly to dark red. The heat was then turned off, and the solution was allowed to gradually cool down to room temperature for 1 hour. The gold nanoparticles were then collected and separated from the solution using a centrifuge and washed three times; by vortexing and dispersing the gold nanoparticles in 10 mL portions of toluene, and then precipitating the gold nanoparticles by adding 40 mL portions of methanol and spinning them in a centrifuge. The solution was then decanted to remove any remaining byproducts and unreacted starting materials. Drying the gold nanoparticles in a vacuum environment produced a solid black pellet; which could be stored for long periods of time (up to one year) for later use, and then redissolved in organic solvents such as toluene.

A protocol for synthesizing ~12 nm diameter gold nanoparticles (Au nanoparticles) in an organic solvent is presented. The gold nanoparticles are capped with oleylamine ligands to prevent agglomeration. The gold nanoparticles are soluble in organic solvents such as toluene.

Methods Article

通过实验室间比较，对液态介质中的金纳米粒子进行纳米粒子跟踪分析