这项工作是由对抗癌症，卡罗来纳州为纳米技术卓越试点补助，大学癌症研究基金会和国家卫生研究所的K - 12的职业发展奖中心的高尔夫球手。

1。 PLGA - B - PEG聚合物的合成<ol><li>聚（D，L -乳酸- CO -乙醇酸）（PLGA）终端羧基 - 羧酸（PLGA）是溶解在浓度为5mm的PLGA（材料一节中提到的）在任何溶剂。 PLGA可以在这个浓度轻轻搅拌溶解。 </li><li>国民保健服务（分子量115.09）和EDC（分子量191.7）溶解在PLGA溶液浓度为25mm。 （EDC和NHS在添加了5次计量过剩相比，以PLGA）。 PLGA -羧酸转化为PLGA - NHS轻轻搅拌约1小时的PLGA - 羧酸溶液加入EDC和NHS。 </li><li>反应产物PLGA - NHS加入甲醇洗涤液沉淀出来。约10倍体积的甲醇过量添加到解决方案。解决的办法是在2000年的离心XG沉淀出PLGA - NHS和上清液（EDC和NHS删除的痕迹，这与甲醇洗涤过程至少重复3次。 </li><li> PLGA - NHS颗粒是下一个30分钟的洗涤液中取出任何痕迹的真空干燥。 </li><li>现在重新PLGA - NHS沉淀溶解在相同的溶剂，在相同浓度最初是用来溶解的PLGA。异型聚乙二醇（胺 - 聚乙二醇 - 羧酸），然后加入浓度为5mm的PLGA溶液（配比1:1）。混合溶液中孵育24小时并不断搅拌。 </li><li> 24小时后，反应产物PLGA - B - PEG嵌段共聚物沉淀，加入过量的洗涤液甲醇。重复三次以上所述的洗衣机和离心过程。这将删除所有多余的未反应的聚乙二醇。 </li><li> PLGA - B - PEG嵌段共聚物，是在真空条件下干燥。 </li></ol> 2。 PLGA - B - PEG纳米粒子的制备这些嵌段共聚物PLGA的核心与PEG在表面覆盖的纳米粒子可以准备。在这种纳米粒子，可封装各种不同的疏水性药物。荧光化合物可以被封装在纳米粒子或可结合的PLGA，因此，这些纳米粒子可以被用于荧光成像。 Nanoprecipitation方法是用来制造纳米粒子，尤其是当封装所需的货物是高度疏水的性质。 <ol><li> PLGA - B - PEG嵌段共聚物和药物/货运（封装）溶于任何溶剂溶解的PLGA。 PLGA的可溶于许多共同的溶剂，包括乙腈，DCM，四氢呋喃，丙酮或乙酸乙酯。溶剂的选择是至关重要的，因为它影响纳米粒子的性质。因此，应该用适当的溶剂，在这一步。 </li><li>聚合物/药物混合，然后滴加3-5卷搅拌水，给人一种最终聚合物浓度约300毫克/毫升。 （图2） </li><li>搅拌是持续了2小时减压，让纳米粒子自组装形成的有机溶剂中删除的痕迹。 </li><li>采伐和净化：纳米粒子，然后集中在2700 X克离心10分钟使用Amicon过滤器（截留分子量20KDa），洗净，在PBS重组。这将删除所有联合国包埋药物/货物。可以进行基本的生物物理表征，如大小，表面电荷，和载药效率，更好地了解纳米粒子的性质。 </li></ol> 3。存储冷冻干燥是一种常用的方法来存储纳米粒子 11 。冷冻干燥将保持长期稳定的12纳米粒子的物理和化学特性。冷冻干燥过程中可能会导致粒子的压力和破坏制定，所以常用的冷冻保护剂（保护冻结应力）和lyo防护剂（保护干燥应力）。这些防护剂的选择是由所需长度的存储时间 13 。 <ol><li>在冷冻干燥，有样品中总凝固，冻结它的Tg以下。 </li><li>在干燥的步骤，删除冰升华。应该优化，实现了高效的冷冻干燥过程的温度和压力。 </li></ol> 4。代表性的成果： PLGA - B - PEG迪嵌段共聚物的表征不同的技术可以被用来确认成功的共轭聚合物。 PLGA - B - PEG的组成，可以使用一个400 MHz 1H核磁共振（NMR）的特点。凝胶渗透色谱（GPC），形成产品的分子量（PLGA - B - PEG）可以验证。 PLGA - B - PEG分子量distributioN曲线和洗脱时间应仅PLGA和PEG不同。结合，这些技术特点而形成的产品，并确定是否成功的共轭反应。 PLGA - B - PEG表征可以测量动态光散射粒度和粒度分布。不同的参数，在nanoprecipitation过程中影响粒子的大小。所使用的最初的聚合物（PLGA和PEG）分子量也影响粒径分布。过渡电子显微镜（TEM）也可以用来确认图3可以看出，纳米粒子尺寸分布和结构。粒度范围一般是在纳米范围内。大颗粒大小，粒度分布不均匀，可能表明在共轭反应或nanoprecipitation方法需要优化的错误。此外，表面zeta电位可以衡量的ZetaPALS。 药物/货物装卸效率是可以量化的标准高效液相色谱法。 可以执行的颗粒溶解在有机溶剂和高效液相色谱法来衡量药物/货物（图4）的吸收。可以做药物释放动力学研究，其中已知的固定数量的纳米粒子在30滑动阿仪的微型透析单位透析。在固定的时间间隔，在透析单元的内容是收集和添加有机溶剂等体积溶解的纳米颗粒。高效液相色谱法对这些样本进行量化药物/货物内容。 <img alt="图1" src="/files/ftp_upload/3398/3398fig1.jpg" /> 图1。 EDC / NHS的化学 <img alt="图2" src="/files/ftp_upload/3398/3398fig2.jpg" /> 图2。 Nanoprecipitation准备聚合物纳米粒子的方法，含有PEG - PLGA嵌段和药物或货物装载到粒子溶剂（乙腈或DCM）的有机溶液滴加3-5毫升搅拌H 2 O <img alt="图3" src="/files/ftp_upload/3398/3398fig3.jpg" /> 图3。传输的nanopartices电子显微镜 TEM图像的PEG - PLGA纳米粒的wortamin。磷钨酸作为造影剂。 <img alt="图4" src="/files/ftp_upload/3398/3398fig4.jpg" /> 图4。从纳米粒子药物控释纳米粒子在PBS透析后释放紫杉醇。指出，粒子被拆除透析录音带和乙腈solublized。解决的办法是用HPLC法测定。纳米粒子的两个单独的地段进行了比较。 <div id="myEventWatcherDiv"></div><div id="myEventWatcherDiv"></div>

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没有利益冲突的声明。

nanoprecipitation方法使用嵌段共聚物是一种简单，快速的方法，工程师聚合物纳米粒子。由此产生的纳米粒子组成一个疏水核心，可用于难溶化合物的交付使用。表面的亲水层，使优良的水溶性，同时提供了可能进一步靶向配体共轭基团。 有许多纳米颗粒平台，包括脂质体，聚合物纳米粒子，树枝状，金属粒子，量子点14。在这些平台中，聚合物纳米粒子的平台是一个?...

<table border="1"><tbody><tr><td> 试剂 </td><td> 公司 </td><td> 目录编号 </td><td> 评论 </td></tr><tr><td>东区区议会</td><td> Thermo Scientific的</td><td> 22980 </td><td>共轭试剂</td></tr><tr><td> NHS的</td><td> Thermo Scientific的</td><td> 24500 </td><td>共轭试剂</td></tr><tr><td>胺聚乙二醇 - 羧酸</td><td> Laysan生物公司</td><td> NH2 - PEG - CM - 5000 </td><td>聚合物（可以使用任何的PEG兆瓦，5000这里列出） </td></tr><tr><td> PLGA - carbxylate </td><td> Lactel </td><td> B6013 - 2 </td><td>聚合物</td></tr><tr><td>二氯甲烷（DCM） </td><td> Sigma - Aldrich公司</td><td> 34856 </td><td>溶剂</td></tr><tr><td>乙腈&gt;纯度达99％ </td><td> Sigma - Aldrich公司</td><td> 34851 </td><td>溶剂</td></tr><tr><td>甲醇&gt;纯度达99％ </td><td> Sigma - Aldrich公司</td><td> 34860 </td><td>洗</td></tr></tbody></table>

formulation of diblock polymeric nanoparticles through nanoprecipitation technique

纳米技术是一项相对较新的科学的分支，它涉及到利用规模的纳米颗粒（纳米颗粒）的独特性能。纳米粒子可以被设计在一个精确的时尚，他们的规模，组成和表面化学可仔细控制。这使前所未有的自由，他们的货物的基本特性，如溶解度，扩散，生物分布，释放特性和免疫原性，修改一些。自创建以来，已经利用纳米粒子科学和医学的许多领域，包括药物递送，成像和细胞生物学 1-4 。但是，它并没有得到充分利用“纳米技术实验室”由于感知技术壁垒之外。在这篇文章中，我们描述了一个简单的方法合成的聚合物为基础的纳米颗粒平台，具有广泛的应用潜力。 第一步是合成一个嵌段共聚物，既是一个疏水区和亲水域。使用模型聚合物PLGA和PEG，我们描述了共轭反应，使用EDC / NHS的化学 5（图1） 。我们还讨论了聚合物的净化过程。合成的嵌段共聚物可以自组装在nanoprecipitation过程中通过疏水 - 亲水相互作用成​​纳米粒子。 所描述的聚合物纳米粒子是非常灵活。可以利用纳米颗粒的疏水核心，进行药物输送experiments6难溶性药物。此外，纳米粒子可以克服难溶性的分子生物学试剂，如渥曼青霉素，这就需要一个像DMSO溶剂，有毒溶剂的问题。然而，二甲基亚砜细胞毒性，干扰实验。这些难溶性药物和试剂，可以有效地交付使用聚合物纳米粒子的毒性极小。高分子纳米粒子也可以被载入与荧光染料和细胞内贩卖的研究利用。最后，这些聚合物纳米粒子可以通过表面的PEG目标配体的结合。这些有针对性的纳米粒子，可以利用标签或细胞中的7-10的特定抗原决定簇。

Watch this Scientific Journal Video about Formulation of Diblock Polymeric Nanoparticles through Nanoprecipitation Technique at JoVE.com

Formulation of Diblock Polymeric Nanoparticles through Nanoprecipitation Technique

本文介绍了nanoprecipitation方法使用嵌段共聚物的合成聚合物为基础的纳米粒子。我们将讨论合成的嵌段共聚物，nanoprecipitation技术，和潜在的应用。

制定通过Nanoprecipitation技术嵌段聚合物纳米粒子

Nanotechnology is a relatively new branch of science that involves harnessing the unique properties of particles that are nanometers in scale ...

formulation-diblock-polymeric-nanoparticles-through-nanoprecipitation

Research

JoVE Journal

Bioengineering

37.1K 次观看.  University of North Carolina School of Medicine. 本文介绍了nanoprecipitation方法使用嵌段共聚物的合成聚合物为基础的纳米粒子。我们将讨论合成的嵌段共聚物，nanoprecipitation技术，和潜在的应用。

视频: Formulation of Diblock Polymeric Nanoparticles through Nanoprecipitation Technique

Evaluation of Polymeric Gene Delivery Nanoparticles by Nanoparticle Tracking Analysis and High-throughput Flow Cytometry

Non-viral gene delivery using polymeric nanoparticles has emerged as an attractive approach for gene therapy to treat genetic diseases1 and as a technology for regenerative medicine2. Unlike viruses, which have significant safety issues, polymeric nanoparticles can be designed to be non-toxic, non-immunogenic, non-mutagenic, easier to synthesize, chemically versatile, capable of carrying larger nucleic acid cargo and biodegradable and/or environmentally responsive. Cationic polymers self-assemble with negatively charged DNA via electrostatic interaction to form complexes on the order of 100 nm that are commonly termed polymeric nanoparticles. Examples of biomaterials used to form nanoscale polycationic gene delivery nanoparticles include polylysine, polyphosphoesters, poly(amidoamines)s and polyethylenimine (PEI), which is a non-degradable off-the-shelf cationic polymer commonly used for nucleic acid delivery1,3 . Poly(beta-amino ester)s (PBAEs) are a newer class of cationic polymers4 that are hydrolytically degradable5,6 and have been shown to be effective at gene delivery to hard-to-transfect cell types such as human retinal endothelial cells (HRECs)7, mouse mammary epithelial cells8, human brain cancer cells9 and macrovascular (human umbilical vein, HUVECs) endothelial cells10.
A new protocol to characterize polymeric nanoparticles utilizing nanoparticle tracking analysis (NTA) is described. In this approach, both the particle size distribution and the distribution of the number of plasmids per particle are obtained11. In addition, a high-throughput 96-well plate transfection assay for rapid screening of the transfection efficacy of polymeric nanoparticles is presented. In this protocol, poly(beta-amino ester)s (PBAEs) are used as model polymers and human retinal endothelial cells (HRECs) are used as model human cells. This protocol can be easily adapted to evaluate any polymeric nanoparticle and any cell type of interest in a multi-well plate format.

A protocol for nanoparticle tracking analysis (NTA) and high-throughput flow cytometry to evaluate polymeric gene delivery nanoparticles is described. NTA is utilized to characterize the nanoparticle particle size distribution and the plasmid per particle distribution. High-throughput flow cytometry enables quantitative transfection efficacy evaluation for a library of gene delivery biomaterials.

高分子基因传递纳米粒子的纳米粒子跟踪分析和高通量流式细胞仪评价

Non-viral gene delivery using polymeric nanoparticles has emerged as an attractive approach for gene therapy to treat genetic diseases1 and as a ...

科研

生物工程

Programming Stem Cells for Therapeutic Angiogenesis Using Biodegradable Polymeric Nanoparticles

Controlled vascular growth is critical for successful tissue regeneration and wound healing, as well as for treating ischemic diseases such as stroke, heart attack or peripheral arterial diseases. Direct delivery of angiogenic growth factors has the potential to stimulate new blood vessel growth, but is often associated with limitations such as lack of targeting and short half-life in vivo. Gene therapy offers an alternative approach by delivering genes encoding angiogenic factors, but often requires using virus, and is limited by safety concerns. Here we describe a recently developed strategy for stimulating vascular growth by programming stem cells to overexpress angiogenic factors in situ using biodegradable polymeric nanoparticles. Specifically our strategy utilized stem cells as delivery vehicles by taking advantage of their ability to migrate toward ischemic tissues in vivo. Using the optimized polymeric vectors, adipose-derived stem cells were modified to overexpress an angiogenic gene encoding vascular endothelial growth factor (VEGF). We described the processes for polymer synthesis, nanoparticle formation, transfecting stem cells in vitro, as well as methods for validating the efficacy of VEGF-expressing stem cells for promoting angiogenesis in a murine hindlimb ischemia model.

We describe the method of programming stem cells to overexpress therapeutic factors for angiogenesis using biodegradable polymeric nanoparticles. Processes described include polymer synthesis, transfecting adipose-derived stem cells in vitro, and validating the efficacy of programmed stem cells to promote angiogenesis in a murine hindlimb ischemia model.

编程干细胞的治疗性血管新生使用可生物降解高分子纳米微球

Controlled vascular growth is critical for successful tissue regeneration and wound healing, as well as for treating ischemic diseases such as stroke, ...

Microfluidic Fabrication of Polymeric and Biohybrid Fibers with Predesigned Size and Shape

A &#8220;sheath&#8221; fluid passing through a microfluidic channel at low Reynolds number can be directed around another &#8220;core&#8221; stream and used to dictate the shape as well as the diameter of a core stream.&#160;Grooves in the top and bottom of a microfluidic channel were designed to direct the sheath fluid and shape the core fluid.&#160;By matching the viscosity and hydrophilicity of the sheath and core fluids, the interfacial effects are minimized and complex fluid shapes can be formed.&#160;Controlling the relative flow rates of the sheath and core fluids determines the cross-sectional area of the core fluid.&#160;Fibers have been produced with sizes ranging from 300 nm to ~1 mm, and fiber cross-sections can be round, flat, square, or complex as in the case with double anchor fibers. Polymerization of the core fluid downstream from the shaping region solidifies the fibers.&#160;Photoinitiated click chemistries are well suited for rapid polymerization of the core fluid by irradiation with ultraviolet light.&#160;Fibers with a wide variety of shapes have been produced from a list of polymers including liquid crystals, poly(methylmethacrylate), thiol-ene and thiol-yne resins, polyethylene glycol, and hydrogel derivatives. Minimal shear during the shaping process and mild polymerization conditions also makes the fabrication process well suited for encapsulation of cells and other biological components.

Two adjacent fluids passing through a grooved microfluidic channel can be directed to form a sheath around a prepolymer core; thereby&#160;determining both shape and cross-section. Photoinitiated polymerization, such as thiol click chemistry, is well suited for rapidly solidifying the core fluid into a microfiber with predetermined size and shape.

预设计尺寸和形状的聚合物纤维和生物贿赂纤维的微流体制造

A &#8220;sheath&#8221; fluid passing through a microfluidic channel at low Reynolds number can be directed around another &#8220;core&#8221; stream and ...

Polymeric Microneedle Array Fabrication by Photolithography

This manuscript describes the fabrication of polymeric microneedle (MN) arrays by photolithography. It involves a simple mold-free process by using a photomask consisting of embedded micro-lenses. Embedded micro-lenses were found to influence MN geometry (sharpness). Robust MN arrays with tip diameters ranging between 41.5 &#181;m &#177; 8.4 &#181;m and 71.6 &#181;m &#177; 13.7 &#181;m, with two different lengths (1,336 &#181;m &#177; 193 &#181;m and 957 &#181;m &#177; 171 &#181;m) were fabricated. These MN arrays may provide potential applications in delivery of low molecular and macromolecular therapeutic agents through skin.

Here, we present a protocol describing a mold-free fabrication process of the polymeric microneedles by photolithography.

聚合物微针阵列制造用光刻

This manuscript describes the fabrication of polymeric microneedle (MN) arrays by photolithography. It involves a simple mold-free process by using a ...

Fabricating Superhydrophobic Polymeric Materials for Biomedical Applications

Superhydrophobic materials, with surfaces possessing permanent or metastable non-wetted states, are of interest for a number of biomedical and industrial applications. Here we describe how electrospinning or electrospraying a polymer mixture containing a biodegradable, biocompatible aliphatic polyester (e.g., polycaprolactone and poly(lactide-co-glycolide)), as the major component, doped with a hydrophobic copolymer composed of the polyester and a stearate-modified poly(glycerol carbonate) affords a superhydrophobic biomaterial. The fabrication techniques of electrospinning or electrospraying provide the enhanced surface roughness and porosity on and within the fibers or the particles, respectively. The use of a low surface energy copolymer dopant that blends with the polyester and can be stably electrospun or electrosprayed affords these superhydrophobic materials. Important parameters such as fiber size, copolymer dopant composition and/or concentration, and their effects on wettability are discussed. This combination of polymer chemistry and process engineering affords a versatile approach to develop application-specific materials using scalable techniques, which are likely generalizable to a wider class of polymers for a variety of applications.

Two- and three-dimensional superhydrophobic polymeric materials are prepared by electrospinning or electrospraying biodegradable polymers blended with a lower surface energy polymer of similar composition.

制造超疏水高分子材料生物医学应用

Superhydrophobic materials, with surfaces possessing permanent or metastable non-wetted states, are of interest for a number of biomedical and industrial ...

Predicting Gene Silencing Through the Spatiotemporal Control of siRNA Release from Photo-responsive Polymeric Nanocarriers

New materials and methods are needed to better control the binding vs. release of nucleic acids for a wide range of applications that require the precise regulation of gene activity. In particular, novel stimuli-responsive materials with improved spatiotemporal control over gene expression would unlock translatable platforms in drug discovery and regenerative medicine technologies. Furthermore, an enhanced ability to control nucleic acid release from materials would enable the development of streamlined methods to predict nanocarrier efficacy a priori, leading to expedited screening of delivery vehicles. Herein, we present a protocol for predicting gene silencing efficiencies and achieving spatiotemporal control over gene expression through a modular photo-responsive nanocarrier system. Small interfering RNA (siRNA) is complexed with mPEG-b-poly(5-(3-(amino)propoxy)-2-nitrobenzyl methacrylate) (mPEG-b-P(APNBMA)) polymers to form stable nanocarriers that can be controlled with light to facilitate tunable, on/off siRNA release. We outline two complementary assays employing fluorescence correlation spectroscopy and gel electrophoresis for the accurate quantification of siRNA release from solutions mimicking intracellular environments. Information gained from these assays was incorporated into a simple RNA interference (RNAi) kinetic model to predict the dynamic silencing responses to various photo-stimulus conditions. In turn, these optimized irradiation conditions allowed refinement of a new protocol for spatiotemporally controlling gene silencing. This method can generate cellular patterns in gene expression with cell-to-cell resolution and no detectable off-target effects. Taken together, our approach offers an easy-to-use method for predicting dynamic changes in gene expression and precisely controlling siRNA activity in space and time. This set of assays can be readily adapted to test a wide variety of other stimuli-responsive systems in order to address key challenges pertinent to a multitude of applications in biomedical research and medicine.

We present a novel method that uses photo-responsive block copolymers for more efficient spatiotemporal control of gene silencing with no detectable off-target effects. Additionally, changes in gene expression can be predicted using straightforward siRNA release assays and simple kinetic modeling.

通过从光响应聚合物纳米载体的siRNA释放的时空控制预测基因沉默

New materials and methods are needed to better control the binding vs. release of nucleic acids for a wide range of applications that require the precise ...

Rapid, Scalable Assembly and Loading of Bioactive Proteins and Immunostimulants into Diverse Synthetic Nanocarriers Via Flash Nanoprecipitation

Nanomaterials present a wide range of options to customize the controlled delivery of single and combined molecular payloads for therapeutic and imaging applications. This increased specificity can have significant clinical implications, including decreased side effects and lower dosages with higher potency. Furthermore, the in situ targeting and controlled modulation of specific cell subsets can enhance in vitro and in vivo investigations of basic biological phenomena and probe cell function. Unfortunately, the required expertise in nanoscale science, chemistry and engineering often prohibit laboratories without experience in these fields from fabricating and customizing nanomaterials as tools for their investigations or vehicles for their therapeutic strategies. Here, we provide protocols for the synthesis and scalable assembly of a versatile non-toxic block copolymer system amenable to the facile formation and loading of nanoscale vehicles for biomedical applications. Flash nanoprecipitation is presented as a methodology for rapid fabrication of diverse nanocarriers from poly(ethylene glycol)-bl-poly(propylene sulfide) copolymers. These protocols will allow laboratories with a wide range of expertise and resources to easily and reproducibly fabricate advanced nanocarrier delivery systems for their applications. The design and construction of an automated instrument that employs a high-speed syringe pump to facilitate the flash nanoprecipitation process and to allow enhanced control over the homogeneity, size, morphology and loading of polymersome nanocarriers is described.

Nanomaterials provide versatile mechanisms of controlled therapeutic delivery for both basic science and translational applications, but their fabrication often requires expertise that is unavailable in most biomedical laboratories. Here, we present protocols for the scalable fabrication and therapeutic loading of diverse self-assembled nanocarriers using flash nanoprecipitation.

快速, 可伸缩的组装和负载的生物活性蛋白和免疫增强剂到不同的合成 Nanocarriers 通过闪光 Nanoprecipitation

Nanomaterials present a wide range of options to customize the controlled delivery of single and combined molecular payloads for therapeutic and imaging ...

Protein Kinase C-delta Inhibitor Peptide Formulation using Gold Nanoparticles

Protein kinase C-delta inhibitor (PKC&#948;i) is a promising drug to prevent ischemia-reperfusion-induced organ injury. It is usually conjugated to a cell-penetrating peptide, TAT, for intracellular delivery. However, TAT has shown non-specific biological activities. Gold nanoparticles (GNPs) can be used as drug delivery carriers without recognized toxicity. Therefore, we have used a GNP/peptide hybrid to deliver PKC&#948;i. Two short peptides (P2: CAAAAE and P4: CAAAAW), at a 95:5 ratio, were used to modify the surface properties of GNP. GNPs conjugated with PKC&#948;i (GNP/PKCi) are stable in distilled water, 0.9% NaCl, and phosphate-buffered saline (PBS) containing bovine serum albumin or fetal bovine serum. Intravenous injection of GNP-PKCi was previously shown to prevent ischemia-reperfusion injury of the lung. This article outlines a protocol to formulate GNP/PKCi and assess the physiochemical properties of GNP/PKCi. We have used similar methods to formulate other peptide-based drugs with GNP. This article will hopefully draw more attention to this novel intracellular drug delivery technology and its applications in vivo.

We have previously used a gold nanoparticle peptide hybrid to intravenously deliver a synthetic peptide, protein kinase C-delta inhibitor, which reduced ischemia-reperfusion-induced acute lung injury. Here we show the detailed protocol of the drug formulation. Other intracellular peptides can be formulated similarly.

金纳米粒子的蛋白质激酶 c-三角洲抑制剂肽配方

Protein kinase C-delta inhibitor (PKC&#948;i) is a promising drug to prevent ischemia-reperfusion-induced organ injury. It is usually conjugated to a ...

Flash NanoPrecipitation for the Encapsulation of Hydrophobic and Hydrophilic Compounds in Polymeric Nanoparticles

The formulation of a therapeutic compound into nanoparticles (NPs) can impart unique properties. For poorly water-soluble drugs, NP formulations can improve bioavailability and modify drug distribution within the body. For hydrophilic drugs like peptides or proteins, encapsulation within NPs can also provide protection from natural clearance mechanisms. There are few techniques for the production of polymeric NPs that are scalable. Flash NanoPrecipitation (FNP) is a process that uses engineered mixing geometries to produce NPs with narrow size distributions and tunable sizes between 30 and 400 nm. This protocol provides instructions on the laboratory-scale production of core-shell polymeric nanoparticles of a target size using FNP. The protocol can be implemented to encapsulate either hydrophilic or hydrophobic compounds with only minor modifications. The technique can be readily employed in the laboratory at milligram scale to screen formulations. Lead hits can directly be scaled up to gram- and kilogram-scale. As a continuous process, scale-up involves longer mixing process run time rather than translation to new process vessels. NPs produced by FNP are highly loaded with therapeutic, feature a dense stabilizing polymer brush, and have a size reproducibility of &#177; 6%.

Flash NanoPrecipitation (FNP) is a scalable approach to produce polymeric core-shell nanoparticles. Lab-scale formulations for the encapsulation of hydrophobic or hydrophilic therapeutics are described.

闪速纳米沉淀法包聚物纳米粒子中疏水亲水化合物的作用

The formulation of a therapeutic compound into nanoparticles (NPs) can impart unique properties. For poorly water-soluble drugs, NP formulations can ...

Preparation of Neutrally-charged, pH-responsive Polymeric Nanoparticles for Cytosolic siRNA Delivery

The success of siRNA as a targeted molecular medicine is dependent upon its efficient cytosolic delivery to cells within the tissue of pathology. Clinical success for treating previously &#8216;undruggable&#8217; hepatic disease targets with siRNA has been achieved. However, efficient tumor siRNA delivery necessitates additional pharmacokinetic design considerations, including long circulation time, evasion of clearance organs (e.g., liver and kidneys), and tumor penetration and retention. Here, we describe the preparation and in vitro physicochemical/biological characterization of polymeric nanoparticles designed for efficient siRNA delivery, particularly to non-hepatic tissues such as tumors. The siRNA nanoparticles are prepared by electrostatic complexation of siRNA and the diblock copolymer poly(ethylene glycol-b-[2-(dimethylamino)ethyl methacrylate-co-butyl methacrylate]) (PEG-DB) to form polyion complexes (polyplexes) where siRNA is sequestered within the polyplex core and PEG forms a hydrophilic, neutrally-charged corona. Moreover, the DB block becomes membrane-lytic as vesicles of the endolysosomal pathway acidify (&#60; pH 6.8), triggering endosomal escape and cytosolic delivery of siRNA. Methods to characterize the physicochemical characteristics of siRNA nanoparticles such as size, surface charge, particle morphology, and siRNA loading are described. Bioactivity of siRNA nanoparticles is measured using luciferase as a model gene in a rapid and high-throughput gene silencing assay. Designs which pass these initial tests (such as PEG-DB-based polyplexes) are considered appropriate for translation to preclinical animal studies assessing the delivery of siRNA to tumors or other sites of pathology.

Methods to prepare and characterize the physicochemical properties and bioactivity of neutrally-charged, pH-responsive siRNA nanoparticles are presented. Criteria for successful siRNA nanomedicines such as size, morphology, surface charge, siRNA loading, and gene silencing are discussed.

用于细胞质 siRNA 传递的带电反应性聚合物纳米粒子的制备

The success of siRNA as a targeted molecular medicine is dependent upon its efficient cytosolic delivery to cells within the tissue of pathology. Clinical ...