两栖金纳米粒子的合成与表征
Summary
两栖金纳米粒子可用于许多生物应用。提出了一种合成用配体二元混合物涂覆的金纳米粒子的协议,并给出了这些粒子的详细表征。
Abstract
金纳米粒子覆盖着1-辛他醇(OT)和11-mercapto-1-undecane硫酸(MUS)的混合物,因为它们与细胞膜、脂质双层和病毒的相互作用得到了广泛的研究。亲水配体使这些颗粒在水溶液中胶体稳定,与疏水配体结合产生一种两栖粒子,可以装载疏水药物,与脂质膜融合,并抵抗非特异性蛋白质吸附。其中许多特性取决于纳米颗粒大小和配体壳的组成。因此,拥有可重复的合成方法和可靠的表征技术,以便确定纳米粒子特性和配体壳组合物至关重要。这里介绍了一个一相化学还原,然后彻底纯化以合成直径在5nm以下的纳米颗粒。纳米粒子表面两个配体之间的比率可以通过合成过程中使用的化学计量比进行调优。我们演示了各种常规技术,如透射电子显微镜 (TEM)、核磁共振 (NMR)、热重力分析 (TGA) 和紫外线可见 (UV-Vis) 光谱仪如何综合结合描述纳米粒子的物理化学参数。
Introduction
金纳米粒子的配体外壳可以设计成表现出几个不同的特性,可用于应对生物医学1、2、3、4的挑战。这种多功能性可以控制纳米粒子和生物分子5、6、7之间的分子间相互作用。疏水性和电荷起决定性作用,以及其他影响纳米粒子与生物分子5、8、9相互作用的表面参数。为了调整纳米粒子的表面特性,根据所寻求的特性,选择构成配体壳的硫酸盐分子提供了无数的可能性。例如,具有疏水性和亲水性(例如,带电)端群的配体分子混合物通常用于生成两栖纳米粒子10、11。
这种类型的纳米粒子的一个突出例子是由OT和MUS(下称MUS:OT纳米粒子)的混合物保护,这些混合物已被证明具有许多相关特性12,13,14。首先,配体壳的组成为66%MUS(下同66:34 MUS:OT),纳米粒子的胶体稳定性高,在脱离子水以及磷酸盐缓冲盐水(1x,4 mM磷酸盐,150 mM NaCl)中,其重量高达33%。此外,这些颗粒不会以相对较低的pH值沉淀:例如,在pH2.3和盐浓度为1M NaCl15时,这些纳米粒子在胶体上稳定数月。配体壳上两个分子之间的化学计量比很重要,因为它决定了具有高离子强度16的溶液中的胶体稳定性。
这些粒子已被证明通过能量无关的通路1,12穿过细胞膜而不对细胞膜进行波刺。这些粒子和脂质双层之间的自发融合是它们通过细胞膜17扩散的基础。这种相互作用背后的机制是尽量减少接触疏水性溶剂可访问的表面面积和水分子与脂质双层融合18。与全MUS纳米粒子(外壳上只有MUS配体的纳米粒子)相比,混合MUS:OT纳米粒子(例如,在66:34 MUS:OT成分下)的水文性较高,增加了芯直径的跨度,而该直径可以与脂质融合双层18。配体壳的不同自组装组织与66:34 MUS:OT纳米粒子与各种蛋白质(如白蛋白和泛性蛋白)的不同结合模式相关,而全MUS粒子19。最近,据报道,66:34 MUS:OT纳米粒子可用作广谱抗病毒剂,由于MUS配体多价静电结合和OT配体非局部联结联结到辣椒,可不可逆转地消灭病毒。蛋白质14.在所有这些情况下,已经发现疏水性含量以及纳米粒子的核心大小决定了这些生物-纳米相互作用是如何发生的。这些不同特性的MUS:OT纳米粒子已经促使许多计算机模拟研究,旨在澄清支持MUS:OT粒子和各种生物结构,如脂质双层20之间的相互作用的机制。
MUS:OT 保护的Au纳米粒子的制备带来了一些挑战。首先,带电配体(MUS)和疏水配体(OT)是不可混用的。因此,在整个合成过程中以及表征过程中,都需要考虑到纳米粒子和配体的溶解度。此外,MUS配体分子的纯度——特别是起始材料中无机盐的含量——影响纳米颗粒的质量、可重复性以及短期和长期胶体稳定性。
这里概述了由MUS和OT混合物保护的这种两栖金纳米粒子的详细合成和表征。报告用于合成带负电荷的MUS配体的协议,以确保不同纳米粒子合成物的纯度,从而保证其可重复性。然后,详细报道了基于常见的一相合成,然后彻底纯化这些纳米粒子的生成过程。各种必要的表征技术21,如TEM,UV-Vis,TGA和NMR,已经结合,以获得任何进一步的生物实验所需的所有参数。
Protocol
Representative Results
Discussion
该协议首先描述了MUS配体的合成,然后描述了两栖MUS:OT金纳米粒子的合成和表征。在纳米粒子合成过程中,以最小的盐含量合成MUS,使配体之间的化学计量比具有更好的可靠性,这是可重复合成MUS:OT纳米粒子的关键因素,具有目标疏水性内容 (图 8)。甲醇作为MUS和OT的常见溶剂,以及乙醇中颗粒的合成,可以可靠地合成MUS:OT金纳米粒子。这里介绍的表征方法构成获得足够纳米粒子信息?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
Z.P.G.和F.S.感谢瑞士国家科学基金会,特别是NCCR”分子系统工程”。Z.L.和F.S.感谢瑞士国家科学基金会第二部分赠款的支持。所有作者感谢曲昂富有成果的讨论和校对手稿。
Materials
| 11-bromo-1-undecene | Sigma Aldrich | 467642-25 ml | |
| Sodium Sulfite | Sigma Aldrich | S0505-250 g | |
| Benzyltriethyl-ammonium bromide | Sigma Aldrich | 147125-25 g | |
| Methanol | VWR | BDH1135-2.5 LP | |
| DI water | Millipore | ZRXQ003WW | Deionized water |
| 1 L round bottom flask | DURAN | 24 170 56 | |
| Diethyl ether | Sigma Aldrich | 1.00930 EMD Millipore | |
| Stirring bar | Sigma Aldrich | Z329207, | |
| Dow Corning High Vacuum Grease | Sigma Aldrich | Z273554 ALDRICH | |
| Filtering flask | DURAN | 20 201 63 | |
| Filtering Buchner Funnel | FisherSci | 11707335 | |
| Ethanol >99.8%, ACS, Reagent | VWR | 2081.321DP | |
| Deuterium dioxide | Sigma Aldrich | 151882 ALDRICH | |
| Thioacetic acid 96% | Sigma Aldrich | T30805 ALDRICH | |
| Carbon black | Sigma Aldrich | 05105-1KG | |
| Celite | Sigma Aldrich | D3877 SIGMA-ALDRICH | Filtration medium |
| Condenser | Sigma Aldrich | Z531154 | |
| Hydrochloric acid, ACS reagent 37% | Sigma Aldrich | 320331 SIGMA-ALDRICH | |
| Sodium Hydroxide, BioXtra, pellets (anhydrous) | Sigma Aldrich | S8045 SIGMA-ALDRICH | |
| Centrifuge tubes | VWR | 525-0155P | |
| 250 mL round bottom flask | DURAN | 24 170 37 | |
| 500 mL round bottom flask | DURAN | 24 170 46 | |
| Nitric acid, fACS reagent 70% | Sigma Aldrich | 438073 SIGMA-ALDRICH | |
| Gold(III) chloride trihydrate >99.9% trace metal basis | Sigma Aldrich | 520918 ALDRICH | |
| 1-octanethiol >98.5% | Sigma Aldrich | 471836 ALDRICH | |
| Sodium Borohydride purum p.a.>96% | Sigma Aldrich | 71320 ALDRICH | |
| Separatory funnel | SIgma Aldrich | Z330655 SIGMA | |
| Funnel | DURAN | 21 351 46 | |
| 2V folded filtering papers | Whatman | 1202-150 | |
| Amicon filters | Merck | UFC903024 | |
| Iodine, ACS reagent, >99.8%, solid | Sigma Aldrich | 207772 SIGMA-ALDRICH | |
| 5 mm NMR-Tubes, Type 5HP (high precision) | Armar | 32210.503 | Length 178 mm |
| Methanol-d4 99.8 atom%D | Armar | 16400.2035 | |
| TGA crucible | Thepro | 9095-9270.45 | |
| 400 mesh carbon supported copper grid | Electron Microscopy Science | CF400-Cu | |
| quartz cuvette | Hellma Analytics | 100-1-40 |
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