Name: synthesis of aptamer-pei-g-peg modified gold nanoparticles loaded with doxorubicin for targeted drug delivery
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Description: Watch this Scientific Journal Video about Synthesis of Aptamer-PEI-g-PEG Modified Gold Nanoparticles Loaded with Doxorubicin for Targeted Drug Delivery at JoVE.com

本研究由中国国家自然科学基金委员会（31700840）资助：河南省重点科研项目（18B430013，18A150049）。这项研究得到了南湖青年学者项目的支持。作者要感谢来自XYNU生命科学学院的本科生曲泽波的乐于助人的作品。作者希望确认XYNU的分析和测试中心使用他们的设备。

注意：在使用所有化学品之前，请务必查阅所有相关材料安全数据表 （MSDS）。用于制备共聚物和纳米粒子的几种化学物质具有剧毒性。纳米粒子也有潜在的危害。确保使用所有适当的安全做法和个人防护设备，包括手套、实验室外套、头罩、全长裤子和近身鞋。
1. 双碳水化合物聚乙二醇（CT-PEG）合成33
<ol><li>将 1 . 46 克（ 14 . 6 mmol ）的氢化物（ SA ）和 209...

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1H NMR光谱（图2）确认CT-PEG共聚合体和PEI-g-PEG共聚合体的成功合成。PEG和PEI的分子量分别为1，000和1，200。此外，EDC/NHS催化系统还用于通过阿米德反应合成PEI-g-PEG共聚物。需要注意的是，如果PEG和PEI的分子量因合成PEI-g-PEG共聚物而发生变化，则需要重新评估反应时间和催化系统。此外，需要进一步调整AUNPs上PEI-g-PEG共聚合体涂层的反应条件，主要是因为PEG-g-PEI聚合体?...

癌症是世界性的主要公共卫生问题，其广泛特征是治愈率低、复发率高、死亡率高。目前传统的抗癌方法包括手术、化疗和放疗3种，其中化疗是4号诊所癌症患者的主要治疗方法。临床使用的抗癌药物主要包括帕利塔塞尔（PTX）5和多索鲁比辛（DOX）6，7。DOX，一种抗肿瘤药物，由于癌症细胞毒性和抑制癌细胞增殖8，9的优点，已广泛应用于临床化疗。然而，DOX导致心毒性10，11，和DOX的短半生限制其应用在诊所12。因此，需要降解的药物携带者以可控的方式将 DOX 加载并以子等方式释放到目标区域。
纳米粒子已广泛应用于靶向药物输送系统，在癌症治疗方面具有若干优势（即表面与体积比大、体积小、封装各种药物的能力、可调谐表面化学等）。13，14，15.特别是金纳米粒子（AuNPs）已广泛应用于生物和生物医学应用，如光热癌治疗16，17。AuNPs的独特特性，如简体合成和一般表面功能化，在癌症治疗18的临床领域具有良好的前景。此外，AuNP已经被用来识别药物输送策略，诊断肿瘤，并克服阻力在许多研究19，20。
尽管如此，AuNP 需要进一步定制，通过增强渗透和保留 （EPR）（如靶向和辅助功能）在肿瘤病变时高局部释放来克服耐药性。聚合物功能化 AuNP 具有独特的优势，如疏水抗癌药物水溶解性提高，循环时间延长21、22。各种生物相容聚合物已用于 AuNP 涂料，如聚乙二醇 （PEG）、聚乙烯胺 （PEI）、透明质酸、肝素和黄豆胶。然后，AuNP的稳定性和有效载荷得到很好的改善。具体来说，PEI是一种高度分支的聚合物，由许多重复单位的初级，二级和三级胺24组成。PEI 具有极好的溶解度、低粘度和高功能性，适合涂在 AuNPs 上。
另一方面，抗癌药物需要直接输送到癌细胞，提高负荷效率，降低毒性，治疗原发性肿瘤和晚期转移性肿瘤25。靶向配体具有巨大的潜力，抗癌药物靶向输送系统26。其靶分子结合的选择性赋予抗癌药物针对特异性，并增加药物在病组织中的丰富性27。更多的配体包括抗体、多肽和小分子。与其他配体相比，核酸贴合剂可以在体外合成，易于修改。AS1411是一种未经改性26b磷化物寡核苷酸，形成稳定的二元G-四聚氰胺结构，专门与癌细胞28、29、30上过度表达的目标核蛋白受体结合。AS1411抑制许多癌细胞的增殖，但不影响健康细胞31，32的生长。因此，AS1411 被用来制造理想的有针对性的药物输送系统。
在这项研究中，PEI-g-PEG共聚物通过阿米德反应合成，然后制造PEI-g-PEG共聚物涂层金纳米粒子（PEI-g-PEG@AuNPs）。此外，DOX 和 AS1411 与准备好的 PEI-g-PEG@AuNPs有顺序链接，如 图 1所示。此详细的协议旨在帮助研究人员避免与制造装有 DOX 和 AS1411 的新 PEI-g-PEG@AuNPs相关的许多常见陷阱。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>4-Dimethylaminopyridine</td>
			<td>Macklin</td>
			<td>D807273</td>
			<td></td>
		</tr>
		<tr>
			<td>A549 cell</td>
			<td></td>
			<td>ATCC CCL-185TM</td>
			<td></td>
		</tr>
		<tr>
			<td>AS1411</td>
			<td>BBI Life Sciences Corporation</td>
			<td></td>
			<td>5&#39;-d (TTTGGTGGTGGTGGTTGTGGTGGTGGTGG) FL-AS1411 (fluorophore-labeled AS1411)</td>
		</tr>
		<tr>
			<td>Anhydrous Tetrahydrofuran (THF)</td>
			<td>SinoPharm Chemical Reagent Co., Ltd</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Cell counting kit-8 (CCK-8)</td>
			<td>Sigma Aldrich</td>
			<td>96992-500TESTS-F</td>
			<td></td>
		</tr>
		<tr>
			<td>Dichloromethane</td>
			<td>Traditional Chinese medicine</td>
			<td>80047318</td>
			<td></td>
		</tr>
		<tr>
			<td>Diethyl ether (Et2O)</td>
			<td>SinoPharm Chemical Reagent Co., Ltd</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Dimethyl sulfoxide</td>
			<td>Macklin</td>
			<td>D806645</td>
			<td></td>
		</tr>
		<tr>
			<td>Dulbecco&#39;s modified Eagle&#39;s medium (DMEM)</td>
			<td>Sigma Aldrich</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Doxorubicin hydrochloride</td>
			<td>Rhawn</td>
			<td>R017518</td>
			<td></td>
		</tr>
		<tr>
			<td>Ether absolute</td>
			<td>Traditional Chinese medicine</td>
			<td>80059618</td>
			<td></td>
		</tr>
		<tr>
			<td>Field Emission Transmission Electron Microscope</td>
			<td>FEI Company</td>
			<td>Tecnai G2 F 20</td>
			<td></td>
		</tr>
		<tr>
			<td>Gold(III) chloride trihydrate</td>
			<td>Rhawn</td>
			<td>R016035</td>
			<td></td>
		</tr>
		<tr>
			<td>Laser Particle-size Instrument</td>
			<td>Malvern Instruments Ltd</td>
			<td>ZetasizerNanoZS/Masterszer3000E</td>
			<td></td>
		</tr>
		<tr>
			<td>Microplate Reader</td>
			<td>Molecular Devices</td>
			<td>SpectraMax 190</td>
			<td></td>
		</tr>
		<tr>
			<td>N-(3-Dimethylaminopropyl)-N&#39;-ethylcarbodiimide hydrochloride</td>
			<td>Macklin</td>
			<td>N808856</td>
			<td></td>
		</tr>
		<tr>
			<td>N-Hydroxysuccinimide</td>
			<td>Macklin</td>
			<td>H6231</td>
			<td></td>
		</tr>
		<tr>
			<td>NMR software</td>
			<td>Delta 5.2.1</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Nuclear Magnetic Resonance Spectrometer</td>
			<td>JEOL</td>
			<td>JNM-ECZ600R/S3</td>
			<td></td>
		</tr>
		<tr>
			<td>Origin 8.5</td>
			<td>OriginLab</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Penicillin</td>
			<td>Sigma Aldrich</td>
			<td>V900929-100ML</td>
			<td></td>
		</tr>
		<tr>
			<td>Phosphate-buffered saline</td>
			<td>Sigma Aldrich</td>
			<td>P4417-100TAB</td>
			<td></td>
		</tr>
		<tr>
			<td>Poly(ethylene glycol)</td>
			<td>Sigma Aldrich</td>
			<td>81188</td>
			<td>BioUltra, average Mn ~ 1000</td>
		</tr>
		<tr>
			<td>Poly (ethyleneimine) solution</td>
			<td>Sigma Aldrich</td>
			<td>482595</td>
			<td>average Mn ~ 1200, 50 wt.% in H2O</td>
		</tr>
		<tr>
			<td>Sodium borohydride, powder</td>
			<td>Acros</td>
			<td>C18930</td>
			<td></td>
		</tr>
		<tr>
			<td>Streptomycin</td>
			<td>Sigma Aldrich</td>
			<td>85886-10ML</td>
			<td></td>
		</tr>
		<tr>
			<td>Succinic anhydride</td>
			<td>Traditional Chinese medicine</td>
			<td>30171826</td>
			<td></td>
		</tr>
		<tr>
			<td>Tetrahydrofuran</td>
			<td>Traditional Chinese medicine</td>
			<td>40058161</td>
			<td></td>
		</tr>
		<tr>
			<td>Triethylamine</td>
			<td>Traditional Chinese medicine</td>
			<td>80134318</td>
			<td></td>
		</tr>
		<tr>
			<td>UV/VIS/NIR Spectrometer</td>
			<td>Lambda950</td>
			<td>Lambda950</td>
			<td></td>
		</tr>
		<tr>
			<td>X-ray Photoelectron Spectrometer</td>
			<td>Thermo&#8194;Fisher&#8194;Scientific</td>
			<td>K-ALPHA 0.5EV</td>
			<td></td>
		</tr>
	</tbody>
</table>

synthesis of aptamer-pei-g-peg modified gold nanoparticles loaded with doxorubicin for targeted drug delivery

由于健康细胞的耐药性和毒性，多索鲁比辛（DOX）在临床癌症治疗中的使用受到限制。该协议描述了用聚乙二醇（PEI-g-PEG）二甘醇（PEI-g-PEG）共聚聚（乙二烯氨酸）嫁接的聚（乙二烯氨酸）与加载的阿普塔默（AS1411）和DOX通过阿米特反应进行功能化金纳米粒子（AuNPs）的设计。AS1411与癌细胞上的靶向核素受体特别结合，使DOX针对的是癌细胞而不是健康细胞。首先，PEG 是盒装的，然后嫁接到分支 PEI 以获得 PEI-g-PEG 聚合物，这一点由 1H NMR 分析确认。其次，PEI-g-PEG copolymer 涂层金纳米粒子 （PEI-g-PEG@AuNPs） 进行合成，DOX 和 AS1411 通过阿米德反应逐渐与 AuNP 粘合。制备的 AS1411-g-DOX-g-PEI-g-PEG@AuNPs直径为 39.9 nm，Zeta 电位为 -29.3 mV，表明纳米粒子在水和细胞介质中是稳定的。细胞毒性检测表明，新设计的DOX加载的AuNPs能够杀死癌细胞（A549）。这种合成展示了PEI-g-PEG共聚物、贴花和DOX在AuNP上的微妙排列，这些排列是通过连续的阿米德反应实现的。这种阿普塔默-PEI-g-PEG功能化的 AuNP 为癌症治疗中的靶向药物输送提供了一个很有前途的平台。

1H NMR光谱学用于确认CT-PEG聚合物和PEI-g-PEG共聚合物（图2）的成功合成。图2a显示，δ的甲基质子信号=3.61ppm，δ=2.57ppm的纸箱质子信号确认CT-PEG聚合物的成功合成。图2b显示PEG的乙烯质子信号在δ=2.6ppm，PEI的质子信号在δ=1.66ppm确认PEI-g-PEG共聚物的合成。 
紫外线光谱学的进行，...

Watch this Scientific Journal Video about Synthesis of Aptamer-PEI-g-PEG Modified Gold Nanoparticles Loaded with Doxorubicin for Targeted Drug Delivery at JoVE.com

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

在此协议中，多索鲁比辛加载 AS1411-g-PEI-g-PEG 改性金纳米粒子通过三步阿米内反应合成。然后，多索鲁比辛被加载并交付给靶向癌细胞进行癌症治疗。

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 ...

Research

JoVE Journal

Chemistry

Template Directed Synthesis of Plasmonic Gold Nanotubes with Tunable IR Absorbance

的模板导演电浆黄金纳米管的合成与可调谐红外吸收

A nearly parallel array of pores can be produced by anodizing aluminum foils in acidic environments1, 2. Applications of anodic aluminum oxide (AAO) ...

科研

化学

Preparation and Characterization of SDF-1&#945;-Chitosan-Dextran Sulfate Nanoparticles

Preparation and Characterization of SDF-1ÃƒÅ½Ã‚Â±-Chitosan-Dextran Sulfate Nanoparticles

SDF-1α - 壳聚糖硫酸葡聚糖纳米粒子的制备与表征

Chitosan (CS) and dextran sulfate (DS) are charged polysaccharides (glycans), which form polyelectrolyte complex-based nanoparticles when mixed under ...

A Simple Method for the Size Controlled Synthesis of Stable Oligomeric Clusters of Gold Nanoparticles under Ambient Conditions

根据环境条件的金纳米粒子的稳定低聚簇的大小控制合成的一种简便方法

Reducing dilute aqueous HAuCl4 with sodium thiocyanate (NaSCN) under alkaline conditions produces 2 to 3 nm diameter nanoparticles. Stable grape-like ...

Protocol for the Synthesis of Ortho-trifluoromethoxylated Aniline Derivatives

Protocol for the Synthesis of Ortho-trifluoromethoxylated Aniline Derivatives

协议的合成<em&gt;邻</em&gt; -trifluoromethoxylated苯胺类化合物

Molecules bearing trifluoromethoxy (OCF3) group often show desired pharmacological and biological properties. However, facile synthesis of ...

Hydroquinone Based Synthesis of Gold Nanorods

金纳米棒基于对苯二酚合成

Gold nanorods are an important kind of nanoparticles characterized by peculiar plasmonic properties. Despite their widespread use in nanotechnology, the ...

Chemical Precipitation Method for the Synthesis of Nb2O5 Modified Bulk Nickel Catalysts with High Specific Surface Area

Chemical Precipitation Method for the Synthesis of Nb2O5 Modified Bulk Nickel Catalysts with High Specific Surface Area

化学沉淀法合成铌2O5改性高比表面积镍催化剂

We demonstrate a method for the synthesis of NixNb1-xO catalysts with sponge-like and fold-like nanostructures. By varying the Nb:Ni ratio, a series of ...

Rapid Nanoprobe Signal Enhancement by In Situ Gold Nanoparticle Synthesis

Rapid Nanoprobe Signal Enhancement by In Situ Gold Nanoparticle Synthesis

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 ...

Synthesis and Characterization of 1,2-Dithiolane Modified Self-Assembling Peptides

12-Dithiolane 改性自组装肽的合成与表征

This report focuses on the synthesis of an N-terminus 1,2-dithiolane modified self-assembling peptide and the characterization of the resulting ...

Molten-Salt Synthesis of Complex Metal Oxide Nanoparticles

复合金属氧化物纳米粒子的熔融盐合成

The development of feasible synthesis methods is critical for the successful exploration of novel properties and potential applications of nanomaterials. ...

Methionine Functionalized Biocompatible Block Copolymers for Targeted Plasmid DNA Delivery

用于靶向质粒DNA输送的美聚氨酸功能化生物相容性块共聚物

Reversible addition-fragmentation chain transfer (RAFT) polymerization integrates the advantages of radical polymerization and living polymerization. This ...