这项工作是由美国国立卫生研究院（补助08UDE017788-05）的支持。 EBP也承认支持西班牙基金会科学与技术（FECYT）。作者感谢人Shonda T.盖洛德和Pratyusha Mogalisetti的手稿的批判性阅读。

<img alt="图1" width="600px" fo:content-width="5.5in" src="/files/ftp_upload/50726/50726fig1.jpg" /> 图1。 。工作流应用的光纤微球体抗体阵列唾液剖析 （1）微球的内部编码的两种荧光染料，（2）所编码的微球在外部修改与蛋白特异性的单克隆抗体，（3）复用的微球混合，和（4）随机地沉积在微孔蚀刻在光纤束的近端，（5）唾液蛋白是通过夹心免疫捕获的微球，和（6）使用荧光显微镜进行定量。 1。编码微球<ol><li>称取钠铕（Ⅲ）thenoyltrifluoroacetonate三水（欧盟TTA，MW =869.54克/摩尔）在琥珀色玻璃瓶中，并在四氢呋喃（THF）准备一个200毫米原液。用移液枪轻轻混匀;直观地验证染料完全溶解。 </li><li>称取香豆素30（C30，MW =347.41克/摩尔）在琥珀色玻璃瓶中，并准备一个12毫米原液在THF。用移液枪轻轻混匀，直观地验证了染料完全溶解。 </li><li>制备700微升使用储备溶液和THF中以达到表1中列出的最终浓度为每微球型工作液。 </li><li>确保微球（10％重量/体积）深受涡旋暂停，再换乘60微升悬浮液1.5 ml离心管中。 </li><li>加入600微升PBS，以微球悬浮和20倍吹打混匀。离心管在10,000 rpm 3分钟，小心取出上清液。重复此过程，另2个洗球。 </li><li>使用相同的程序，步骤1.5十二烷基硫酸钠洗涤微球沉淀3次。 </li><li>离心微球/ THF悬浮液以10,000 rpm离心3分钟，除去上清液，并重新悬浮于600微升的沉淀工作溶液列于表1。将混合物转移至新的1.5 ml微量离心管中。 </li><li>密封离心管封口膜，并把它放在摇床。摇在3,000 rpm孵育24小时，用一个盒子上面有铝箔覆盖安装避光。 </li><li>离心微球悬浮液在10,000 rpm下3分钟，以形成颗粒。 </li><li>除去上清液染料溶液，冲洗微球6X与600微升甲醇中，然后用600微升PBS含0.01％Tween-20的另一种6倍，如在步骤1.5中描述。 </li><li>存储编码微球在600微升PBS含0.01％吐温20在4°C，直到使用，避光。 </li></ol><table border="1"><tbody><tr><td> 微球型名称： </td><td> 1 </td><td> 2 </td><td> 3 </td><td> 4 </td><td> 5 </td><td> 6 </STRONG&gt; </td><td> 7 </td></tr><tr><td> EU-TTA（MM） </td><td> 100 </td><td> 100 </td><td> 10 </td><td> 100 </td><td> 10 </td><td></td><td></td></tr><tr><td> C30（MM） </td><td></td><td> 1 </td><td> 1 </td><td> 6 </td><td> 6 </td><td> 1 </td><td> 6 </td></tr></tbody></table> 表1中。的Eu-TTA和C30浓度的工作溶液。 2。蛋白质捕获微球的制备<ol><li>制备的MES缓冲液[2 - （N-吗啉）乙磺酸（MES）的0.1M，氯化钠0.9％，SDS 0.01％，pH值为5.7]和PBS / SDS缓冲液（磷酸钠0.01 M，氯化钠0.154 M，SDS 0.01％，pH值7.4）预先在室温下。 </li><li>加入200μl编码微球悬浮液（含〜2毫克微球）到一个安全锁1.5 ml离心管。使用这种类型的离心管中，以避免在步骤2.6中温育过程中泄漏是很重要的。 </li> &lt;LI&gt;清洗微球600微升的MES按照步骤1.5中所述缓冲3倍。加入700μl的MES在微球颗粒和吹打拌匀。 </li><li>准备0.105克/ ml的1 - 乙基-3 - [3 - 二甲基氨基丙基]碳二亚胺盐酸盐（EDC，MW =191.7克/摩尔）和0.163克/毫升磺基-N-羟基硫代琥珀酰亚胺（磺基-NHS，MW =217.13克/ mol）的在MES缓冲溶液中分离管。 </li><li>加入新鲜配制的EDC解决方案， 一滴一滴进入微球悬浮液150微升。 EDC的水解速度非常快，因此，它必须使用新鲜的溶液，以确保固定化效率。紧接在添加EDC后，加入150μl的磺基-NHS溶液的微球，通过移液混合均匀，并验证该悬浮液是均匀的。 </li><li>盖上管，用封口膜密封，并将其放置在摇床。摇在1,500 rpm 4小时，用一个盒子上面有铝箔避光。 </li><li>离心微球suspens离子以10,000 rpm离心3分钟，以形成沉淀物，并除去上清液。如在步骤1.5中所述，用500μL的PBS / SDS缓冲液3X冲洗微球。 </li><li>加入200μl的PBS / SDS缓冲液的颗粒，混合微球以及后转移到300微升含60微克的捕获抗体的PBS / SDS缓冲液。它首先暂停微球的颗粒，然后加入微球悬浮液到抗体溶液中是很重要的。 </li><li>通过摇动在1,500 rpm 4小时在摇床上孵育抗体 - 微球混合物中，用框有铝箔避光。 </li><li>离心微球悬浮液在10,000 rpm下3分钟，以形成沉淀物，并除去上清液。用500μlStartingBlock（TBS）封闭缓冲液3次，如在步骤1.5中所述洗涤微球。 </li><li>混合性微球用1ml TBS的封闭缓冲液孵育微球在1,500 rpm进行1小时的摇床上，用一个框科夫避光红色的有铝箔。 </li><li>离心微球悬浮液在7,000 rpm离心3分钟，以形成沉淀物，并除去上清液。用500μLTBS封闭缓冲液，如在步骤1.5中所述洗涤沉淀3次。 </li><li>离心微球溶液以7,000 rpm离心3分钟，以形成沉淀物，并除去上清液。加入500μlTBS封闭液并吹打混匀。 </li><li>存储修改后的微球悬浮在4°C，直到使用，避光。 </li><li>重复此步骤，以使其他类型使用相应抗体的微球。 </li></ol> 3。光纤芯片组装<ol><li>混合50μl的每种类型的蛋白捕获微球的成新的高压灭菌的离心管中。离心机股票微球池在7000 rpm下3分钟，除去上清，使剩余的体积为约100微升。 </li><li>手工切割的光纤束至大约5厘米的长度和依次用30，15，9,6，3，1，0.5和0.05微米尺寸的金刚石研磨膜的抛光机上抛光的两端。超声处理在去离子水中2分钟，在抛光束以除去任何颗粒。 </li><li>把小电磁搅拌棒加入0.5 ml离心管中，加入400微升的无蛋白质的PBS缓冲液中，并搅拌上磁性搅拌板30分钟以除去气泡。 </li><li>在150秒10,16 0.025 1N HCl溶液蚀刻抛光纤维束的一端。立即浸没和超声处理的蚀刻结束在去离子水中1分钟。干燥该纤维束的压缩空气。 </li><li>装载在纤维束上的纤维托座和阻止蚀刻端在无气泡，无蛋白的PBS缓冲液1小时。 </li><li>关于光纤束的蚀刻端存储的微球悬浮液的存款1微升等份。保护设置从光与铝箔盒，并等待15分钟。悬浮液的量将通过蒸发减少，微球强行进入蚀刻的微孔。重复此步加载一次。 </li><li>使用与样品溶液饱和棉签除去未被困在微孔微球。蛋白质微阵列现在可以使用了。请立刻步骤4.1唾液分析。 </li></ol> 4。唾液样本分析使用微芯片<ol><li>加入200μl样品溶液（100微升唾液上清液稀释StartingBlock T20（PBS）等体积的封闭液。唾液上清液中收集协议先前已公布的10）。入0.5ml的灭菌离心管中。沾在纤维进入溶液中加载的蛋白质微阵列孵育在摇床上以600rpm搅拌2小时，用的是盒覆盖有铝箔避光。 </li><li>准备20毫升洗涤缓冲液，加入200μl的10％BSA溶液和200μl的10％Tween-20的成19.6毫升PBS，英里x深受轻轻晃动。 </li><li>浸蛋白质微阵列到含有200μl的洗涤缓冲液的新高压灭菌的离心管中，并摇动以600rpm持续2分钟。重复此洗3次。 </li><li>孵育微阵列中含有抗-VEGF，IP-10，IL-8，EGF，MMP-9的5μg/ ml的检测抗体鸡尾酒的100微升溶液中，在StartingBlock T20 IL-1β的生物素化的检测抗体（PBS）封闭缓冲液30分钟，在室温下在摇床上以600rpm，从光利用箱用铝箔覆盖保护。 </li><li>按照步骤4.3中所述用洗涤缓冲液清洗芯片3倍。 </li><li>孵育微阵列在200μl的PBS中20μg/ ml的链霉亲和R-藻红蛋白缀合物（SAPE）的溶液在600rpm的摇床上10分钟，用盒子覆盖有铝箔避光。 </li><li>按照步骤4.3中所述用洗涤缓冲液洗芯片5倍。 </li><li>清洁纤维（ 即 ，端部远离的远端从微球），捆绑用无水乙醇饱和棉签。 </li><li>干燥压缩空气的光纤的两端。通过纤维的前端安装的光纤上的落射荧光显微镜和图像。 </li><li>获取对应的Eu-TTA，C30，和SAPE荧光发射强度的微球的图像。光过滤器的设置和曝光时间的Eu-TTA，C30，和SAPE的荧光成像被描述在表2中。 </li></ol><table border="1"><tbody><tr><td> 渠道 </td><td> EU-TTA </td><td> C30 </td><td> SAPE </td></tr><tr><td>激励</td><td> 365/10x </td><td> 350/50x </td><td> 546/10x </td></tr><tr><td>分光镜</td><td> 525DCLP </td><td> 400DCLP </td><td> 560LP </td></tr><tr><td>发射器</td><td> 620/60m </td><td> 460/50m </td><td> 580/30m </td></tr><tr><td>曝光时间</td><td> 1秒</td><td> 0.3秒</td><td> 1秒</td></tr></tbody></table> 表2。参数用于微阵列的三个荧光图像。

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作者宣称，他们有没有竞争的财务权益。

研究者应该格外注意以下步骤：为更好的解码精度，有必要验证微球均匀地悬浮在所有孵育和微球编码过程中的洗涤步骤。此外，经编码的微球需要在整个实验中，以避光。下列正确的编码和储存程序中，我们发现，总体解码准确率为99％以上。该编码微球应保存在4℃保存，避免冷冻和避光。在适当的储存条件下，编码微球是稳定超过六个月。在编码和修改步骤，需要格外小心，以?...

由于报道由Mark Schena并在90年代中期的同事第一芯片，这个强大的工具已被用于生物研究1的许多领域。能够同时检测多种蛋白质在诊断液，如血液的抗体微阵列，具有在临床诊断和生物标志物筛选2-10重要的应用。唾液，含有许多相同的分析物如血，一直被认为是一个优选的替代方案，因为血液唾液收集是安全的，非侵入性的，并且可以进行通过微创医疗人员11-13。目前，使用唾液样本复用蛋白分析是由几个重要的因素，其中包括目标分析物14的低浓度和不同的生物标记物15的宽浓度范围的限制。 。 在此，我们证明6种蛋白的分析：人血管内皮生长因子（VEGF），干扰素γ-诱导蛋白10（IP-10），白细胞介素-8（IL-8），表皮生长因子（EGF），基质金属蛋白酶9（MMP-9），和白细胞介素-1β（IL-1β） 。该方法的性能用标准溶液构成的重组蛋白分析物和封闭缓冲液中最初验证。从不同的慢性呼吸系统疾病的患者以及健康对照真正的唾液样本进行了测试表现令人满意。该协议应当适用于其他蛋白质的分析物和其他基于微球的实验。该平台提供了相当大的优势，在分析化学领域，因为它使低浓度的几种蛋白质具有广泛的动态范围，最小的非特异性相互作用，减少样品消耗和低成本的快速，精确，可重复的和同时分析在比较的类似的酶联免疫吸附试验（ELISA）。

<table border="1">
	<tbody>
		<tr>
			<td>Name of Reagent</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Eu-TTA dye</td>
			<td>Fisher Scientific</td>
			<td>AC42319-0010</td>
			<td></td>
		</tr>
		<tr>
			<td>THF</td>
			<td>Sigma-Aldrich</td>
			<td>34865-100ML</td>
			<td></td>
		</tr>
		<tr>
			<td>Amber glass vial</td>
			<td>Fisher Scientific</td>
			<td>03-339-23B</td>
			<td></td>
		</tr>
		<tr>
			<td>Coumarin 30 dye</td>
			<td>Sigma-Aldrich</td>
			<td>546127-100MG</td>
			<td></td>
		</tr>
		<tr>
			<td>Microspheres</td>
			<td>Bangslabs</td>
			<td>PC05N/6698</td>
			<td></td>
		</tr>
		<tr>
			<td>1.5 ml microcentrifuge tubes</td>
			<td>Fisher Scientific</td>
			<td>05-408-129</td>
			<td></td>
		</tr>
		<tr>
			<td>PBS 10x concentrate</td>
			<td>Sigma-Aldrich</td>
			<td>P5493-1L</td>
			<td></td>
		</tr>
		<tr>
			<td>Water</td>
			<td>Sigma-Aldrich</td>
			<td>W4502-1L</td>
			<td></td>
		</tr>
		<tr>
			<td>Methanol</td>
			<td>Sigma-Aldrich</td>
			<td>34860-100ML</td>
			<td></td>
		</tr>
		<tr>
			<td>Tw-20</td>
			<td>Sigma-Aldrich</td>
			<td>P7949-100 ml</td>
			<td></td>
		</tr>
		<tr>
			<td>BupH MES buffered saline</td>
			<td>Thermo Scientific</td>
			<td>28390</td>
			<td></td>
		</tr>
		<tr>
			<td>SDS</td>
			<td>Sigma-Aldrich</td>
			<td>05030-500ML-F</td>
			<td></td>
		</tr>
		<tr>
			<td>NaOH solution</td>
			<td>Fisher Scientific</td>
			<td>SS256-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Safe-lock microcentrifuge tube</td>
			<td>VWR labshop</td>
			<td>53511-997</td>
			<td></td>
		</tr>
		<tr>
			<td>EDC</td>
			<td>Thermo Scientific</td>
			<td>22980</td>
			<td></td>
		</tr>
		<tr>
			<td>Sulfo-NHS</td>
			<td>Thermo Scientific</td>
			<td>24510</td>
			<td></td>
		</tr>
		<tr>
			<td>Human VEGF capture antibody</td>
			<td>R&#38;D Systems</td>
			<td>MAB293</td>
			<td></td>
		</tr>
		<tr>
			<td>Human IP-10 capture antibody</td>
			<td>R&#38;D Systems</td>
			<td>MAB266</td>
			<td></td>
		</tr>
		<tr>
			<td>Human IL-8 capture antibody</td>
			<td>R&#38;D Systems</td>
			<td>MAB208</td>
			<td></td>
		</tr>
		<tr>
			<td>Human EGF capture antibody</td>
			<td>R&#38;D Systems</td>
			<td>MAB636</td>
			<td></td>
		</tr>
		<tr>
			<td>Human MMP-9 capture antibody</td>
			<td>R&#38;D Systems</td>
			<td>MAB936</td>
			<td></td>
		</tr>
		<tr>
			<td>Human IL-1&#946; capture antibody</td>
			<td>R&#38;D Systems</td>
			<td>MAB601</td>
			<td></td>
		</tr>
		<tr>
			<td>Mouse IgG1 isotype control antibody</td>
			<td>R&#38;D Systems</td>
			<td>MAB002</td>
			<td></td>
		</tr>
		<tr>
			<td>StartingBlock (TBS) buffer</td>
			<td>Thermo Scientific</td>
			<td>37542</td>
			<td></td>
		</tr>
		<tr>
			<td>HCl standard solution 1.0 N</td>
			<td>Sigma-Aldrich</td>
			<td>318949-500 ml</td>
			<td></td>
		</tr>
		<tr>
			<td>0.5 ml microcentrifuge tubes</td>
			<td>Fisher Scientific</td>
			<td>05-408-120</td>
			<td></td>
		</tr>
		<tr>
			<td>Protein-free (PBS) buffer</td>
			<td>Thermo Scientific</td>
			<td>37572</td>
			<td></td>
		</tr>
		<tr>
			<td>Recombinant human VEGF 165</td>
			<td>R&#38;D Systems</td>
			<td>293-VE</td>
			<td></td>
		</tr>
		<tr>
			<td>Recombinant human IP-10</td>
			<td>R&#38;D Systems</td>
			<td>266-IP</td>
			<td></td>
		</tr>
		<tr>
			<td>Recombinant human IL-8</td>
			<td>R&#38;D Systems</td>
			<td>208-IL</td>
			<td></td>
		</tr>
		<tr>
			<td>Recombinant human EGF</td>
			<td>R&#38;D Systems</td>
			<td>236-EG</td>
			<td></td>
		</tr>
		<tr>
			<td>Recombinant human MMP-9</td>
			<td>R&#38;D Systems</td>
			<td>911-MP</td>
			<td></td>
		</tr>
		<tr>
			<td>Recombinant human IL-1&#946;</td>
			<td>R&#38;D Systems</td>
			<td>201-LB</td>
			<td></td>
		</tr>
		<tr>
			<td>StartingBlock T20 (PBS) buffer</td>
			<td>Thermo Scientific</td>
			<td>37539</td>
			<td></td>
		</tr>
		<tr>
			<td>Blocker BSA in PBS</td>
			<td>Thermo Scientific</td>
			<td>37525</td>
			<td></td>
		</tr>
		<tr>
			<td>Biotinylated VEGF detection antibody</td>
			<td>R&#38;D Systems</td>
			<td>BAF293</td>
			<td></td>
		</tr>
		<tr>
			<td>Biotinylated IP-10 detection antibody</td>
			<td>R&#38;D Systems</td>
			<td>BAF266</td>
			<td></td>
		</tr>
		<tr>
			<td>Biotinylated IL-8 detection antibody</td>
			<td>R&#38;D Systems</td>
			<td>BAF208</td>
			<td></td>
		</tr>
		<tr>
			<td>Biotinylated EGF detection antibody</td>
			<td>R&#38;D Systems</td>
			<td>BAF236</td>
			<td></td>
		</tr>
		<tr>
			<td>Biotinylated MMP-9 detection antibody</td>
			<td>R&#38;D Systems</td>
			<td>BAF911</td>
			<td></td>
		</tr>
		<tr>
			<td>Biotinylated IL-1&#946; detection antibody</td>
			<td>R&#38;D Systems</td>
			<td>BAF201</td>
			<td></td>
		</tr>
		<tr>
			<td>Streptavidin, R-phycoerythrin</td>
			<td>Invitrogen</td>
			<td>S-21388</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethanol (200 proof)</td>
			<td>Sigma-Aldrich</td>
			<td>E7023-500ML</td>
			<td></td>
		</tr>
	</tbody>
</table>

multiplexed fluorescent microarray for human salivary protein analysis using polymer microspheres and fiber-optic bundles

本文中，我们描述了一种协议，用于同时测量6蛋白在唾液中使用的光纤基于微球的抗体阵列。所采用的免疫阵列技术结合基于微球的悬浮阵列制造与使用荧光显微镜的优点。如在视频协议描述的，可商购的4.5微米的聚合物微球进行编码成七种不同类型的，由两个荧光染料物理截留在微球内的浓度分化。表面含羧基的编码微球通过EDC / NHS偶联化学进行了修改，用单克隆捕获抗体。组装的蛋白质微阵列，不同类型的编码和官能化的微球混合，并随机地沉积在4.5微米的微孔，从而进行化学蚀刻在一光纤束的近端。光纤束被用来同时作为载体和用于成像的米icrospheres。一旦组装后，该微阵列用于捕获蛋白在从诊所收集唾液上清液。该检测是基于使用生物素化的检测抗体的混合物为不同的分析物与链霉亲和结合的荧光探针，R-藻红蛋白的夹心免疫测定。微阵列是在三个不同的通道，两个对微球的注册和一个用于检测信号成像，荧光显微镜。的荧光显微照片，然后解码，并在MATLAB中使用自制的算法进行分析。

从三个通道表示光纤束的一小部分的荧光图像示于图2A-C。使用MATLAB写的一个算法这些图像进行分析（如在讨论部分更详细地描述）。该分析采用从铕-TTA编码图像（ 图2A）中的信息和C30的编码图像（ 图2B）来解码的微球，并计算信号的图像（ 图2C）在不同的微球的荧光强度。与相关蛋白的标准中获得的校准曲线，计算蛋白的唾液样品中的浓度。 </...

Watch this Scientific Journal Video about Multiplexed Fluorescent Microarray for Human Salivary Protein Analysis Using Polymer Microspheres and Fiber-optic Bundles at JoVE.com

Multiplexed Fluorescent Microarray for Human Salivary Protein Analysis Using Polymer Microspheres and Fiber-optic Bundles

我们描述了使用多路复用基于微球的抗体阵列分析唾液蛋白的过程。单克隆抗体共价连接到荧光染料编码的4.5微米聚合物用碳二亚胺化学性微球。修改后的微球沉积在光纤的微孔用荧光夹心免疫测定蛋白水平在唾液。

复用荧光微阵列人类唾液蛋白质分析用高分子微球和光纤束

Herein, we describe a protocol for simultaneously measuring six proteins in saliva using a fiber-optic microsphere-based antibody array. The immuno-array ...

multiplexed-fluorescent-microarray-for-human-salivary-protein

Research

JoVE Journal

Chemistry

11.6K 次观看.  Tufts University. 我们描述了使用多路复用基于微球的抗体阵列分析唾液蛋白的过程。单克隆抗体共价连接到荧光染料编码的4.5微米聚合物用碳二亚胺化学性微球。修改后的微球沉积在光纤的微孔用荧光夹心免疫测定蛋白水平在唾液。 

视频: Multiplexed Fluorescent Microarray for Human Salivary Protein Analysis Using Polymer Microspheres and Fiber-optic Bundles

Gyroid Nickel Nanostructures from Diblock Copolymer Supramolecules

Nanoporous metal foams possess a unique combination of properties - they are catalytically active, thermally and electrically conductive, and furthermore, have high porosity, high surface-to-volume and strength-to-weight ratio. Unfortunately, common approaches for preparation of metallic nanostructures render materials with highly disordered architecture, which might have an adverse effect on their mechanical properties. Block copolymers have the ability to self-assemble into ordered nanostructures and can be applied as templates for the preparation of well-ordered metal nanofoams. Here we describe the application of a block copolymer-based supramolecular complex - polystyrene-block-poly(4-vinylpyridine)(pentadecylphenol) PS-b-P4VP(PDP) - as a precursor for well-ordered nickel nanofoam. The supramolecular complexes exhibit a phase behavior similar to conventional block copolymers and can self-assemble into the bicontinuous gyroid morphology with two PS networks placed in a P4VP(PDP) matrix. PDP can be dissolved in ethanol leading to the formation of a porous structure that can be backfilled with metal. Using electroless plating technique, nickel can be inserted into the template&#39;s channels. Finally, the remaining polymer can be removed via pyrolysis from the polymer/inorganic nanohybrid resulting in nanoporous nickel foam with inverse gyroid morphology.

This article describes the preparation of well-ordered nickel nanofoams via electroless metal deposition onto nanoporous templates obtained from self-assembled diblock copolymer based supramolecules.

Gyroid镍纳米结构的嵌段共聚物超分子

Nanoporous metal foams possess a unique combination of properties - they are catalytically active, thermally and electrically conductive, and furthermore, ...

科研

化学

Preparation of Highly Porous Coordination Polymer Coatings on Macroporous Polymer Monoliths for Enhanced Enrichment of Phosphopeptides

We describe a protocol for the preparation of hybrid materials based on highly porous coordination polymer coatings on the internal surface of macroporous polymer monoliths. The developed approach is based on the preparation of a macroporous polymer containing carboxylic acid functional groups and the subsequent step-by-step solution-based controlled growth of a layer of a porous coordination polymer on the surface of the pores of the polymer monolith. The prepared metal-organic polymer hybrid has a high specific micropore surface area. The amount of iron(III) sites is enhanced through metal-organic coordination on the surface of the pores of the functional polymer support. The increase of metal sites is related to the number of iterations of the coating process.
The developed preparation scheme is easily adapted to a capillary column format. The functional porous polymer is prepared as a self-contained single-block porous monolith within the capillary, yielding a flow-through separation device with excellent flow permeability and modest back-pressure. The metal-organic polymer hybrid column showed excellent performance for the enrichment of phosphopeptides from digested proteins and their subsequent detection using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The presented experimental protocol is highly versatile, and can be easily implemented to different organic polymer supports and coatings with a plethora of porous coordination polymers and metal-organic frameworks for multiple purification and/or separation applications.

A procedure for the preparation of porous hybrid separation media composed of a macroporous polymer monolith internally coated by a high surface area microporous coordination polymer is presented.

对磷酸增强富集制备高度多孔配位聚合物涂层的孔聚合物石柱的

We describe a protocol for the preparation of hybrid materials based on highly porous coordination polymer coatings on the internal surface of macroporous ...

Functionalization of Single-walled Carbon Nanotubes with Thermo-reversible Block Copolymers and Characterization by Small-angle Neutron Scattering

We demonstrate a protocol for single-walled carbon nanotube functionalization using thermo-sensitive PEO-PPO-PEO triblock copolymers in an aqueous solution. In a carbon nanotube/PEO105-PPO70-PEO105 (poloxamer 407) aqueous solution, the amphiphilic poloxamer 407 adsorbs onto the carbon nanotube surfaces and self-assembles into continuous layers, driven by intermolecular interactions between constituent molecules. The addition of 5-methylsalicylic acid changes the self-assembled structure from spherical-micellar to a cylindrical morphology. The fabricated poloxamer 407/carbon nanotube hybrid particles exhibit thermo-responsive structural features so that the density and thickness of poloxamer 407 layers are also reversibly controllable by varying temperature. The detailed structural properties of the poloxamer 407/carbon nanotube particles in suspension can be characterized by small-angle neutron scattering experiments and model fit analyses. The distinct curve shapes of the scattering intensities depending on temperature control or addition of aromatic additives are well described by a modified core-shell cylinder model consisting of a carbon nanotube core cylinder, a hydrophobic shell, and a hydrated polymer layer. This method can provide a simple but efficient way for the fabrication and in-situ characterization of carbon nanotube-based nano particles with a structure-tunable encapsulation.

A method for the functionalization of carbon nanotubes with structure-tunable polymeric encapsulation layers and structural characterization using small-angle neutron scattering is presented.

单壁碳纳米管与热可逆嵌段共聚物及表征功能化小角度中子散射

We demonstrate a protocol for single-walled carbon nanotube functionalization using thermo-sensitive PEO-PPO-PEO triblock copolymers in an aqueous ...

The Preparation and Properties of Thermo-reversibly Cross-linked Rubber Via Diels-Alder Chemistry

A method for using Diels Alder thermo-reversible chemistry as cross-linking tool for rubber products is demonstrated. In this work, a commercial ethylene-propylene rubber, grafted with maleic anhydride, is thermo-reversibly cross-linked in two steps. The pending anhydride moieties are first modified with furfurylamine to graft furan groups to the rubber backbone. These pendant furan groups are then cross-linked with a bis-maleimide via a Diels-Alder coupling reaction. Both reactions can be performed under a broad range of experimental conditions and can easily be applied on a large scale. The material properties of the resulting Diels-Alder cross-linked rubbers are similar to a peroxide-cured ethylene/propylene/diene rubber (EPDM) reference. The cross-links break at elevated temperatures (&#62; 150 &#176;C) via the retro-Diels-Alder reaction and can be reformed by thermal annealing at lower temperatures (50-70 &#176;C). Reversibility of the system was proven with infrared spectroscopy, solubility tests and mechanical properties. Recyclability of the material was also shown in a practical way, i.e., by cutting a cross-linked sample into small parts and compression molding them into new samples displaying comparable mechanical properties, which is not possible for conventionally cross-linked rubbers.

A simple two-step approach involving rubber modification and cross-linking yields fully reworkable, elastic rubber products.

热可逆交联橡胶通过的Diels-Alder反应的化学制备与性能

A method for using Diels Alder thermo-reversible chemistry as cross-linking tool for rubber products is demonstrated. In this work, a commercial ...

Using Capillary Electrophoresis to Quantify Organic Acids from Plant Tissue: A Test Case Examining Coffea arabica Seeds

Carboxylic acids are organic acids containing one or more terminal carboxyl (COOH) functional groups. Short chain carboxylic acids (SCCAs; carboxylic acids containing three to six carbons), such as malate and citrate, are critical to the proper functioning of many biological systems, where they function in cellular respiration and can serve as indicators of cell health. In foods, organic acid content can have significant impact on taste, with increased SCCA levels resulting in a sour or &#34;acid&#34; taste. Because of this, methods for the rapid analysis of organic acid levels are of particular interest to the food and beverage industries. Unfortunately, however, most methods used for SCCA quantification are dependent on time-consuming protocols requiring the derivatization of samples with hazardous reagents, followed by costly chromatographic and/or mass spectrometric analyses. This method details an alternate method for the detection and quantification of organic acids from plant material and food samples using free zonal capillary electrophoresis (CZE), sometimes simply referred to as capillary electrophoresis (CE). CZE provides a cost-effective method for measuring SCCAs with a low limit of detection (0.005 mg/ml). This article details the extraction and quantification of SCCAs from plant samples. While the method provided focuses on measurement of SCCAs from coffee beans, the method provided can be applied to multiple plant-based food materials.

Using Capillary Electrophoresis to Quantify Organic Acids from Plant Tissue: A Test Case Examining Coffea arabica Seeds

This article presents a method for the detection and quantification of organic acids from plant material using free zonal capillary electrophoresis. An example of the potential application of this method, determining the effects of a secondary fermentation on organic acid levels in coffee seeds, is provided.

利用毛细管电泳从植物组织量化有机酸：一个测试用例检查<em&gt;小粒种咖啡</em&gt;种子

Carboxylic acids are organic acids containing one or more terminal carboxyl (COOH) functional groups. Short chain carboxylic acids (SCCAs; carboxylic ...

Microfluidic Dry-spinning and Characterization of Regenerated Silk Fibroin Fibers

The protocol demonstrates a method for mimicking the spinning process of silkworm. In the native spinning process, the contracting spinning duct enables the silk proteins to be compact and ordered by shearing and elongation forces. Here, a biomimetic microfluidic channel was designed to mimic the specific geometry of the spinning duct of the silkworm. Regenerated silk fibroin (RSF) spinning doped with high concentration, was extruded through the microchannel to dry-spin fibers at ambient temperature and pressure. In the post-treated process, the as-spun fibers were drawn and stored in ethanol aqueous solution. Synchrotron radiation wide-angle X-ray diffraction (SR-WAXD) technology was used to investigate the microstructure of single RSF fibers, which were fixed to a sample holder with the RSF fiber axis normal to the microbeam of the X-ray. The crystallinity, crystallite size, and crystalline orientation of the fiber were calculated from the WAXD data. The diffraction arcs near the equator of the two-dimensional WAXD pattern indicate that the post-treated RSF fiber has a high orientation degree.

A protocol for the microfluidic spinning and microstructure characterization of regenerated silk fibroin monofilament is presented.

微流控干纺及再生丝素纤维的表征

The protocol demonstrates a method for mimicking the spinning process of silkworm. In the native spinning process, the contracting spinning duct enables ...

Electrochemical Impedance Spectroscopy as a Tool for Electrochemical Rate Constant Estimation

Electrochemical impedance spectroscopy (EIS) was used for advanced characterization of organic electroactive compounds along with cyclic voltammetry (CV). In the case of fast reversible electrochemical processes, current is predominantly affected by the rate of diffusion, which is the slowest and limiting stage. EIS is a powerful technique that allows separate analysis of stages of charge transfer that have different AC frequency response. The capability of the method was used to extract the value of charge transfer resistance, which characterizes the rate of charge exchange on the electrode-solution interface. The application of this technique is broad, from biochemistry up to organic electronics. In this work, we are presenting the method of analysis of organic compounds for optoelectronic applications.

Electrochemical impedance spectroscopy (EIS) of species that undergo reversible oxidation or reduction in solution was used for determination of rate constants of oxidation or reduction.

电化学阻抗谱作为电化学速率常数估计的一种工具

Electrochemical impedance spectroscopy (EIS) was used for advanced characterization of organic electroactive compounds along with cyclic voltammetry (CV). ...

Time-resolved Photophysical Characterization of Triplet-harvesting Organic Compounds at an Oxygen-free Environment Using an iCCD Camera

Here, we present a sensible method of the acquisition and analysis of time-resolved photoluminescence using an ultrafast iCCD camera. This system enables the acquisition of photoluminescence spectra covering the time regime from nanoseconds up to 0.1 s. This enables us to follow the changes in the intensity (decay) and emission of the spectra over time. Using this method, it is possible to study diverse photophysical phenomena, such as the emission of phosphorescence, and the contributions of prompt and delayed fluorescence in molecules showing thermally activated delayed fluorescence (TADF). Remarkably, all spectra and decays are obtained in a single experiment. This can be done for solids (thin film, powder, crystal) and liquid samples, where the only limitations are the spectral sensitivity of the camera and the excitation wavelength (532 nm, 355 nm, 337 nm, and 266 nm). This technique is, thus, very important when investigating the excited state dynamics in organic emitters for their application in organic light-emitting diodes and other areas where triplet harvesting is of paramount importance. Since triplet states are strongly quenched by oxygen, emitters with efficient TADF luminescence, or those showing room temperature phosphorescence (RTP), must be correctly prepared in order to remove any dissolved oxygen from solutions and films. Otherwise, no long-lived emission will be observed. The method of degassing solid samples as presented in this work is basic and simple, but the degassing of liquid samples creates additional difficulties and is particularly interesting. A method of minimizing solvent loss and changing the sample concentration, while still enabling to remove oxygen in a very efficient and a repeatable manner, is presented in this work.

Here, we present a method of the spectroscopic characterization of organic molecules by means of time-resolved photoluminescence spectroscopy on the nanosecond-to-millisecond timescale in oxygen-free conditions. Methods to efficiently remove oxygen from the samples and, thus, limit luminescence quenching are also described.

利用 iccd 摄像机对无氧环境下的三重收集有机化合物进行光物理表征

Here, we present a sensible method of the acquisition and analysis of time-resolved photoluminescence using an ultrafast iCCD camera. This system enables ...

Using Cyclic Voltammetry, UV-Vis-NIR, and EPR Spectroelectrochemistry to Analyze Organic Compounds

Cyclic voltammetry (CV) is a technique used in the analysis of organic compounds. When this technique is combined with electron paramagnetic resonance (EPR) or ultraviolet-visible and near-infrared (UV-Vis-NIR) spectroscopies, we obtain useful information such as electron affinity, ionization potential, band-gap energies, the type of charge carriers, and degradation information that can be used to synthesize stable organic electronic devices. In this study, we present electrochemical and spectroelectrochemical methods to analyze the processes occurring in active layers of an organic device as well as the generated charge carriers.

In this article, we describe electrochemical, electron paramagnetic resonance, and ultraviolet-visible and near-infrared spectroelectrochemical methods to analyze organic compounds for application in organic electronics.

利用循环伏安法、紫外-近红外光谱和 EPR 光谱电化学对有机化合物进行分析

Cyclic voltammetry (CV) is a technique used in the analysis of organic compounds. When this technique is combined with electron paramagnetic resonance ...

Sample Preparation in Quartz Crystal Microbalance Measurements of Protein Adsorption and Polymer Mechanics

In this study, we present various examples of how thin film preparation for quartz crystal microbalance experiments informs the appropriate modeling of the data and determines which properties of the film can be quantified. The quartz crystal microbalance offers a uniquely sensitive platform for measuring fine changes in mass and/or mechanical properties of an applied film by observing the changes in mechanical resonance of a quartz crystal oscillating at high frequency. The advantages of this approach include its experimental versatility, ability to study changes in properties over a wide range of experimental time lengths, and the use of small sample sizes. We demonstrate that, based on the thickness and shear modulus of the layer deposited on the sensor, we can acquire different information from the material. Here, this concept is specifically exploited to display experimental parameters resulting in mass and viscoelastic calculations of adsorbed collagen on gold and polyelectrolyte complexes during swelling as a function of salt concentration.

The quartz crystal microbalance can provide accurate mass and viscoelastic properties for films in the micron or submicron range, which is relevant for investigations in biomedical and environmental sensing, coatings, and polymer science. The sample thickness influences which information can be obtained from the material in contact with the sensor.

石英晶体微平衡测量中的蛋白质吸附和聚合物力学的样品制备

In this study, we present various examples of how thin film preparation for quartz crystal microbalance experiments informs the appropriate modeling of ...