This research was supported by the Natural Sciences and Engineering Research Council (NSERC) of Canada, the Canada Research Chairs Program, and Simon Fraser University. This work made use of 4D LABS shared facilities supported by the Canada Foundation for Innovation (CFI), British Columbia Knowledge Development Fund (BCKDF) and Simon Fraser University.

1. Synthesis of the NaYF4/Yb3+/Er3+ Upconverting Nanoparticles (UCNP)
<ol>
	<li>Set up the apparatus as followed:
	<ol>
		<li>Place a 250 ml heating mantle on a regular stirring plate and plug the mantle onto the thermal couple.</li>
		<li>Place a 250 ml round bottom flask equipped with a magnetic stir bar onto the heating mantle with proper clamping.</li>
		<li>Attach an air adapter to the left neck of the round bottom flask and connect this air adapter to the Schlenk line with plastic tubing.</li>
		<li>Attach a glass adapter to the right neck of the round bottom flask and fix a thermometer adapter onto the glass adapter. Insert the temperature probe into the flask through the thermometer adapter and plug this into the thermocouple.</li>
		<li>Attach a distillation head to the middle neck of the round bottom flask. Place a stopper on top of the distillation head. Connect the head to a condenser, followed by a vacuum distillation adapter and a 50 ml round bottom flask. Connect the vacuum distillation adapter to a bubbler through plastic tubing.&#160;</li>
	</ol>
	</li>
	<li>Weigh 1.17 g (3.9 mmol) of yttrium acetate, 0.439 g of ytterbium acetate and 0.0727 g (0.1 mmol) of erbium acetate and place them in the reaction round bottom flask.</li>
	<li>Add 30 ml of oleic acid and 75 ml of octadecene to the flask using a graduated cylinder.</li>
	<li>Rinse the side of the reaction round bottom flask using 5 ml of methanol to make sure that no oleic acid and octadecene is stuck to the sides of the reaction flask.</li>
	<li>Connect the reaction flask to a double manifold Schlenk line and turn the corresponding valve to keep the reaction flask connected to the nitrogen line.</li>
	<li>Turn on the thermocouple, set the temperature to 80&#160;&#176;C, and gradually heat the system to this temperature. At 80&#160;&#176;C and after all starting materials are dissolved, remove the heating mantle and allow the reaction to cool down to 30 &#176;C.</li>
	<li>When temperature reaches 30 &#176;C, take off the distillation head, switch the air adapter from the left neck to the middle neck and close off the left neck with a stopper. Slowly introduce vacuum to the reaction flask by turning the valve on the Schlenk line from the nitrogen line to the vacuum line. All of the low boiling point components will be pulled out from the reaction at this point.</li>
	<li>When the solution stops bubbling, raise up the temperature to 115 &#176;C in a speed of 5 &#176;C/min.</li>
	<li>Once the temperature reaches 115 &#176;C, maintain this temperature for 15 min, then remove the heating mantle and cool down the reaction to 50 &#176;C. Afterwards, quickly switch the set up back to the original form by reattaching the distillation head to the middle neck and the air adapter to the left head.</li>
	<li>Weigh out 0.74 g (12.5 mmol) of NaOH and 0.50 g (20.0 mmol) of NH4F during the cooling process, and dissolve them in 50 ml of methanol by sonication.</li>
	<li>After sonication, pour the solution into the reaction round bottom flask and rinse the sides of the flask with 5 ml of MeOH.</li>
	<li>Leave the solution stirring at 50 &#176;C for 30 min.</li>
	<li>Increase the temperature to 75 &#176;C to distill the methanol.</li>
	<li>During the distillation, empty the collection flask when necessary. After the distillation is finished, heat up the reaction to 300 &#176;C under nitrogen protection as fast as possible.</li>
	<li>Once the temperature reaches 300 &#176;C, maintain this temperature for 1 hr. If needed, cover the setup with aluminum foil to help maintain the temperature. Then remove heat source and allow the reaction to cool down to room temperature.</li>
	<li>Once it is cooled down to room temperature, split the solution evenly into three centrifugation tubes (50 ml tubes, roughly 35 ml solution per each tube), and top up the tube to the 50 ml scale using anhydrous ethanol. Centrifuge all the tubes at 3,400 x g for 15 min. After centrifugation, the UCNPs should be observed on the side of the tubes as a white precipitate.</li>
	<li>Discard the supernatant and redisperse the UCNPs pellets in hexanes (7.5 ml of hexanes per each tube), then top up the tube with ethanol to the 50 ml scale. Centrifuge tubes again at 3,400 x g for 15 min.</li>
	<li>Once the centrifugation is complete, discard the supernatant and redisperse the solid UCNPs in 30 ml of CHCl3 for further use.</li>
</ol>
2. Assembly of Water-dispersible Nanoassemblies Containing Organic Dye Molecules and Upconverting Nanoparticles
<ol>
	<li>Dissolve 25 mg (0.0147 mmol) of poly(styrene-alt-maleic anhydride) (PSMA) in 3 ml of CHCl3 into a scintillation vial equipped with a magnetic stir bar. This quantity is an optimized quantity after multiple trials.</li>
	<li>Add 250 &#181;l (47 mg/ml) of the upconverting nanoparticles chloroform stock solution to the scintillation vial.</li>
	<li>Cap the vial and place it on the magnetic stirring plate, and stir the solution at room temperature for 2 hr.</li>
	<li>Weigh 160 mg (0.0773 mmol) of polyether amine 2070, and dissolve it in 1 ml of CHCl3. Then add this solution to the scintillation vial in one portion using a pipette. The solution will turn to pale yellow indicating the reaction of polyether amine 2070 with the anhydride groups on the PSMA.</li>
	<li>Continue to stir the solution overnight at room temperature.</li>
	<li>Measure the appropriate quantity of organic dye molecules then dispense it into the scintillation vial in one portion, stir the resulting solution for 1 hr.
	<ol>
		<li>For the sample TPP-NP (nanoassembly containing polymer shell, tetraphenyl porphyrin and upconverting nanoparticles), directly add 1 mg of tetraphenyl porphyrin to the scintillation vial. For the sample DAE-UCNP (nanoassembly containing polymer shell, diarylethene molecules and upconverting nanoparticles), the quantity of each diarylethene molecules is 2 &#215; 10-7 mol. Add the two diarylethene molecules into the reaction solution. The volumes for the two diarylethene molecules are: DAE-1o (1.8 mM), 111 &#181;l and DAE-2o (1.6 mM), 125 &#181;l.</li>
	</ol>
	</li>
	<li>Remove the CHCl3 solvent under reduced pressure using a rotary evaporator, then add 3 ml of 0.001 M aqueous NaOH (pH &#8776; 11) to the scintillation vial, then sonicate the vial until a milky suspension is formed.</li>
	<li>Place the vial back on the rotary evaporator, and carefully remove the remaining CHCl3 until the suspension has turned to a clear solution.</li>
	<li>Transfer the solution from the scintillation vial to two 1.5 ml conical centrifugation tubes, then centrifuge the solution at 20,600 x g for 25 min.</li>
	<li>Discard the supernatant, then add a total of 3 ml of deionized H2O into the two tubes (1.5 ml per tube), sonicate the tubes to redisperse the pellets in the deionized H2O.</li>
	<li>Centrifuge the two tubes again at 20,600 x g for 25 min.</li>
	<li>Discard the supernatant, then add a total of 3 ml of deionized H2O into the two tubes (1.5 ml per tube). Sonicate the tubes to redisperse the pellets in the deionized H2O.</li>
	<li>Filter the aqueous nanoparticles dispersion sample through a 0.2 &#181;m syringe filter to obtain the final sample for further testing.</li>
</ol>

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The authors have nothing to disclose.

The nanoparticles synthesized according to this protocol have a size distribution from 20 to 25 nm centered at around 22.5 nm.26,27 They can be classified as spherical particles with a &#945;-NaYF4 host lattice structure. There are two critical steps in this protocol. In the UCNP synthesis, it is crucial to maintain the heating temperature and time as precise as possible to assure a narrow distribution of particle size. Simultaneous addition of NaOH and NH4F along with the addition of lan...

Today there is still an urgent need to develop new types of bio-imaging agents. Many novel fluorescent probes have been well documented.1-6 However, substantial improvements in the image resolution remains a challenge.7 One practical method is to directly modulate the fluorescence probes between a &#8216;light&#8217; emissive state and a &#8216;dark&#8217; quenched state.8-12 This particular method has been applied to develop technologies such as stimulated emission depletion (STED) microscopy13 and stochastic optical reconstruction microscopy (STORM).14
Another approach to modulate fluorescence is to couple photoresponsive chromophores together with fluorescent probes.15,16 Toggling the photoresponsive chromophore between two isomers where only one of the isomers can act as an efficient energy-transfer acceptor, allows control over quenching of the fluorescence from the probe through F&#246;rster Resonance Energy Transfer (FRET) and other mechanisms. The result is the creation of an emissive state and a quenched state that can be alternated by exposure of the photoresponsive chromophore to different wavelengths of light.
Photoresponsive diarylethene chromophores can be reversibly toggled between a colorless ring-open isomer and a colored ring-closed isomer upon irradiation with UV and visible light.17-19 The thermal stability of the two isomers and tunable absorption spectra of the ring-closed isomer make diarylethenes very good candidates as controllable FRET acceptors.20-23 Lanthanide-doped NaYF4 upconverting nanoparticles are useful for bio-imaging.24 These nanoparticles absorb near-infrared light and emit light in several regions of the visible spectrum. Examples of fluorescence modulation by combining photoresponsive diarylethene chromophores and nanoparticles have been previously reported by our group.25-27 However, the systems described in each example required an additional synthetic modification to attach the diarylethenes to the surface of the nanoparticles, which complicates the development of more diverse systems.
Herein we demonstrate a simple &#8216;plug-and-play&#8217; method to prepare water-dispersible organic dye molecules and photoresponsive upconverting nanoparticles using a self-assembly strategy. The choice of polymers; poly(styrene-alt-maleic anhydride) and polyether amine 2070 provide both a hydrophobic and hydrophilic environment. The hydrophobic sections of the polymer help to hold the normally water insoluble organic molecules and upconverting nanoparticles together, whereas the hydrophilic region of the polymer is critical for maintaining the water-solubility. We will first demonstrate synthesis of the upconverting nanoparticles by the thermal nucleation method. Then, we will prove how the organic molecules and upconverting nanoparticles are encapsulated within hydrophobic regions of the polymer shell and remain stable in aqueous media by simply co-stirring a solution of the upconverting nanoparticles, polymer and different organic dye molecules, followed by a convenient work-up procedure. We also demonstrate how to modulate fluorescence emission of the assemblies using external light irradiation. We anticipate the scope of using this &#8216;plug-and-play&#8217; method to make water-dispersible nanoassemblies will continue to expand.

<table><tbody><tr><td>Yttrium acetate</td><td>Sigma</td><td>326046</td><td>Yttrium(III) acetate hydrate</td></tr><tr><td>Ytterbium acetate</td><td>Sigma</td><td>544973</td><td>Ytterbium(III) acetate hydrate</td></tr><tr><td>Erbium acetate</td><td>Sigma</td><td>325570</td><td>Erbium(III) acetate hydrate</td></tr><tr><td>Oleic acid</td><td>Sigma</td><td>75096</td><td>analytical standard</td></tr><tr><td>Octadecene</td><td>Sigma</td><td>O806</td><td>Technical grade</td></tr><tr><td>NaOH</td><td>Sigma</td><td>S5881</td><td>reagent grade</td></tr><tr><td>NH&lt;sub&gt;4&lt;/sub&gt;F</td><td>Sigma</td><td>216011</td><td>ACS reagent</td></tr><tr><td>Poly(styrene-co-maleic anhydride)</td><td>Sigma</td><td>442399</td><td>Average Mn = 1700</td></tr><tr><td>JeffAmine 2070</td><td>Huntsman</td><td>M-2070</td><td></td></tr><tr><td>Varian Carry 300</td><td>Agilent</td><td></td><td></td></tr><tr><td>JDSU NIR laser</td><td>JSDU</td><td>L4-9897510-100M</td><td>980 nm diode laser</td></tr></tbody></table>

a 'plug and play' method to create water-dispersible nanoassemblies containing an amphiphilic polymer, organic dyes and upconverting nanoparticles

In this protocol, we first describe a procedure to synthesize lanthanide doped upconverting nanoparticles (UCNPs). We then demonstrate how to generate amphiphilic polymers in situ, and describe a protocol to encapsulate the prepared UCNPs and different organic dye molecules (porphyrins and diarylethenes) using polymer shells to form stable water-dispersible nanoassemblies. The nanoassembly samples containing both the UCNPs and the diarylethene organic dyes have interesting photochemical and photophysical properties. Upon 365 nm UV irradiation, the diarylethene group undergoes a visual color change. When the samples are irradiated with visible light of another specific wavelength, the color fades and the samples return to the initial colorless state. The samples also emit visible light from the UCNPs upon irradiation with 980 nm near-infrared light. The emission intensity of the samples can be tuned through alternate irradiation with UV and visible light. Modulation of fluorescence can be performed for many cycles without observable degradation of the samples. This versatile encapsulation procedure allows for the transfer of hydrophobic molecules and nanoparticles from an organic solvent to an aqueous medium. The polymer helps to maintain a lipid-like microenvironment for the organic molecules to aid in preservation of their photochemical behavior in water. Thus this method is ideal to prepare water-dispersible photoresponsive systems. The use of near-infrared light to activate upconverting nanoparticles allows for lower energy light to be used to activate photoreactions instead of more harmful ultraviolet light.

Absorption spectra and photoluminescence spectra were collected for sample DAE-UCNP. The absorption spectra are used for comparing the spectral overlap between the closed diarylethene chromophores and the upconverting nanoparticles. Photographs of the samples (both TPP-UCNP and DAE-UCNP) were also included to demonstrate successful encapsulation of organic dye molecules and upconverting nanoparticles, which are located within the amphiphilic polymer shells in the aqueous phase. The modulation of photochemistry and fluore...

Watch this Scientific Journal Video about A 'Plug and Play' Method to Create Water-dispersible Nanoassemblies Containing an Amphiphilic Polymer, Organic Dyes and Upconverting Nanoparticles at JoVE.com

A 'Plug and Play' Method to Create Water-dispersible Nanoassemblies Containing an Amphiphilic Polymer, Organic Dyes and Upconverting Nanoparticles

Organic dye molecules and oleic acid coated upconverting nanoparticles are not water-soluble. This protocol describes a &#8216;plug and play&#8217; method that enables the transfer of organic dye molecules and upconverting particles from their initial hydrophobic solvent to water.

In this protocol, we first describe a procedure to synthesize lanthanide doped upconverting nanoparticles (UCNPs). We then demonstrate how to generate ...

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9.8K Views.  Simon Fraser University. Organic dye molecules and oleic acid coated upconverting nanoparticles are not water-soluble. This protocol describes a ‘plug and play’ method that enables the transfer of organic dye molecules and upconverting particles from their initial hydrophobic solvent to water. 

Video: A 'Plug and Play' Method to Create Water-dispersible Nanoassemblies Containing an Amphiphilic Polymer, Organic Dyes and Upconverting Nanoparticles

Controlling the Size, Shape and Stability of Supramolecular Polymers in Water

For aqueous based supramolecular polymers, the simultaneous control over shape, size and stability is very difficult1. At the same time, the ability to do so is highly important in view of a number of applications in functional soft matter including electronics, biomedical engineering, and sensors. In the past, successful strategies to control the size and shape of supramolecular polymers typically focused on the use of templates2,3, end cappers4 or selective solvent techniques5. 
Here we disclose a strategy based on self-assembling discotic amphiphiles that leads to the control over stack length and shape of ordered, chiral columnar aggregates. By balancing electrostatic repulsive interactions on the hydrophilic rim and attractive non-covalent forces within the hydrophobic core of the polymerizing building block, we manage to create small and discrete spherical objects6,7. Increasing the salt concentration to screen the charges induces a sphere-to-rod transition. Intriguingly, this transition is expressed in an increase of cooperativity in the temperature-dependent self-assembly mechanism, and more stable aggregates are obtained. 
For our study we select a benzene-1,3,5-tricarboxamide (BTA) core connected to a hydrophilic metal chelate via a hydrophobic, fluorinated L-phenylalanine based spacer (Scheme 1). The metal chelate selected is a Gd(III)-DTPA complex that contains two overall remaining charges per complex and necessarily two counter ions. The one-dimensional growth of the aggregate is directed by &pi;-&pi; stacking and intermolecular hydrogen bonding. However, the electrostatic, repulsive forces that arise from the charges on the Gd(III)-DTPA complex start limiting the one-dimensional growth of the BTA-based discotic once a certain size is reached. At millimolar concentrations the formed aggregate has a spherical shape and a diameter of around 5 nm as inferred from 1H-NMR spectroscopy, small angle X-ray scattering, and cryogenic transmission electron microscopy (cryo-TEM). The strength of the electrostatic repulsive interactions between molecules can be reduced by increasing the salt concentration of the buffered solutions. This screening of the charges induces a transition from spherical aggregates into elongated rods with a length &gt; 25 nm. Cryo-TEM allows to visualise the changes in shape and size. In addition, CD spectroscopy permits to derive the mechanistic details of the self-assembly processes before and after the addition of salt. Importantly, the cooperativity -a key feature that dictates the physical properties of the produced supramolecular polymers- increases dramatically upon screening the electrostatic interactions. This increase in cooperativity results in a significant increase in the molecular weight of the formed supramolecular polymers in water.

The goal of this experiment is to determine and control the size, shape and stability of self-assembled discotic amphiphiles in water. For aqueous based supramolecular polymers such level of control is very difficult. We apply a strategy using both repulsive and attractive interactions. The experimental techniques applied to characterize this system are broadly applicable.

For aqueous based supramolecular polymers, the simultaneous control over shape, size and stability is very difficult1. At the same time, the ability to do ...

Engineering

A Technique to Functionalize and Self-assemble Macroscopic Nanoparticle-ligand Monolayer Films onto Template-free Substrates

This protocol describes a self-assembly technique to create macroscopic monolayer films composed of ligand-coated nanoparticles1,2. The simple, robust and scalable technique efficiently functionalizes metallic nanoparticles with thiol-ligands in a miscible water/organic solvent mixture allowing for rapid grafting of thiol groups onto the gold nanoparticle surface. The hydrophobic ligands on the nanoparticles then quickly phase separate the nanoparticles from the aqueous based suspension and confine them to the air-fluid interface. This drives the ligand-capped nanoparticles to form monolayer domains at the air-fluid interface.&#160; The use of water-miscible organic solvents is important as it enables the transport of the nanoparticles from the interface onto template-free substrates.&#160; The flow is mediated by a surface tension gradient3,4 and creates macroscopic, high-density, monolayer nanoparticle-ligand films.&#160; This self-assembly technique may be generalized to include the use of particles of different compositions, size, and shape and may lead to an efficient assembly method to produce low-cost, macroscopic, high-density, monolayer nanoparticle films for wide-spread applications.

A simple, robust and scalable technique to functionalize and self-assemble macroscopic nanoparticle-ligand monolayer films onto template-free substrates is described in this protocol.

This protocol describes a self-assembly technique to create macroscopic monolayer films composed of ligand-coated nanoparticles1,2. The simple, robust and ...

Formulation of Diblock Polymeric Nanoparticles through Nanoprecipitation Technique

Nanotechnology is a relatively new branch of science that involves harnessing the unique properties of particles that are nanometers in scale (nanoparticles). Nanoparticles can be engineered in a precise fashion where their size, composition and surface chemistry can be carefully controlled. This enables unprecedented freedom to modify some of the fundamental properties of their cargo, such as solubility, diffusivity, biodistribution, release characteristics and immunogenicity. Since their inception, nanoparticles have been utilized in many areas of science and medicine, including drug delivery, imaging, and cell biology1-4. However, it has not been fully utilized outside of "nanotechnology laboratories" due to perceived technical barrier. In this article, we describe a simple method to synthesize a polymer based nanoparticle platform that has a wide range of potential applications. 
The first step is to synthesize a diblock co-polymer that has both a hydrophobic domain and hydrophilic domain. Using PLGA and PEG as model polymers, we described a conjugation reaction using EDC/NHS chemistry5 (Fig 1). We also discuss the polymer purification process. The synthesized diblock co-polymer can self-assemble into nanoparticles in the nanoprecipitation process through hydrophobic-hydrophilic interactions.
The described polymer nanoparticle is very versatile. The hydrophobic core of the nanoparticle can be utilized to carry poorly soluble drugs for drug delivery experiments6. Furthermore, the nanoparticles can overcome the problem of toxic solvents for poorly soluble molecular biology reagents, such as wortmannin, which requires a solvent like DMSO. However, DMSO can be toxic to cells and interfere with the experiment. These poorly soluble drugs and reagents can be effectively delivered using polymer nanoparticles with minimal toxicity. Polymer nanoparticles can also be loaded with fluorescent dye and utilized for intracellular trafficking studies. Lastly, these polymer nanoparticles can be conjugated to targeting ligands through surface PEG. Such targeted nanoparticles can be utilized to label specific epitopes on or in cells7-10.

This article describes a nanoprecipitation method to synthesize polymer-based nanoparticles using diblock co-polymers. We will discuss the synthesis of diblock co-polymers, the nanoprecipitation technique, and potential applications.

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

Bioengineering

Using Polystyrene-block-poly(acrylic acid)-coated Metal Nanoparticles as Monomers for Their Homo- and Co-polymerization

We present a template-free method for &#8220;polymerizing&#8221; nanoparticles into long chains without side branches. A variety of nanoparticles are encapsulated in polystyrene-block-poly(acrylic acid) (PSPAA) shells and then used as monomers for their self-assembly. Spherical PSPAA micelles upon acid treatment are known to assemble into cylindrical micelles. Exploiting this tendency, the core-shell nanoparticles are induced to aggregate, coalesce, and then transform into long chains. When more than one type of nanoparticles are used, random and block &#8220;copolymers&#8221; of nanoparticles can be obtained. Detailed procedures are reported for the PSPAA encapsulation of nanoparticles, homo- and co-polymerization of the core-shell nanoparticles, separation and purification of the resulting nanoparticle chains. Transformations of single-line chains into double- and triple-line chains are also presented. The synergy between the polymer shell and the embedded nanoparticles leads to an unusual chain-growth polymerization mode, giving long nanoparticle chains that are distinct from the products of the traditional step-growth aggregation process.

Using Polystyrene-block-polyacrylic acid-coated Metal Nanoparticles as Monomers for Their Homo- and Co-polymerization

We report protocols for &#8220;polymerizing&#8221; various types of polymer-encapsulated metal nanoparticles into long chains of &#8220;homo-&#8220; and &#8220;co-polymers&#8221;.

We present a template-free method for &#8220;polymerizing&#8221; nanoparticles into long chains without side branches. A variety of nanoparticles are ...

Facile Synthesis of Worm-like Micelles by Visible Light Mediated Dispersion Polymerization Using Photoredox Catalyst

Presented herein is a protocol for the facile synthesis of worm-like micelles by visible light mediated dispersion polymerization. This approach begins with the synthesis of a hydrophilic poly(oligo(ethylene glycol) methyl ether methacrylate) (POEGMA) homopolymer using reversible addition-fragmentation chain-transfer (RAFT) polymerization. Under mild visible light irradiation (&#955; = 460 nm, 0.7 mW/cm2), this macro-chain transfer agent (macro-CTA) in the presence of a ruthenium based photoredox catalyst, Ru(bpy)3Cl2 can be chain extended with a second monomer to form a well-defined block copolymer in a process known as Photoinduced Electron Transfer RAFT (PET-RAFT). When PET-RAFT is used to chain extend POEGMA with benzyl methacrylate (BzMA) in ethanol (EtOH), polymeric nanoparticles with different morphologies are formed in situ according to a polymerization-induced self-assembly (PISA) mechanism. Self-assembly into nanoparticles presenting POEGMA chains at the corona and poly(benzyl methacrylate) (PBzMA) chains in the core occurs in situ due to the growing insolubility of the PBzMA block in ethanol. Interestingly, the formation of highly pure worm-like micelles can be readily monitored by observing the onset of a highly viscous gel in situ due to nanoparticle entanglements occurring during the polymerization. This process thereby allows for a more reproducible synthesis of worm-like micelles simply by monitoring the solution viscosity during the course of the polymerization. In addition, the light stimulus can be intermittently applied in an ON/OFF manner demonstrating temporal control over the nanoparticle morphology.

This article describes a process for producing polymeric self-assembled nanoparticles using visible light mediated dispersion polymerization. Using low energy visible light to control the polymerization allows for the reproducible formation of self-assembled worm-like micelles at high solids content.

Presented herein is a protocol for the facile synthesis of worm-like micelles by visible light mediated dispersion polymerization. This approach begins ...

Fabrication of Spherical and Worm-shaped Micellar Nanocrystals by Combining Electrospray, Self-assembly, and Solvent-based Structure Control

Micellar nanocrystals (micelles with encapsulated nanocrystals) have become an emerging major class of nanobiomaterials. We describe a method of fabricating micellar nanocrystals based on combining top-down electrospray, bottom-up self-assembly, and solvent-based structure control. This method involves first using electrospray to generate uniform ultrafine liquid droplets, each of which functions as a micro-reactor in which self-assembly reaction occurs forming micellar nanocrystals, with the structures (micelle shape and nanocrystal encapsulation) controlled by the organic solvent used. This method is largely continuous and produces high quality micellar nanocrystal products with an inexpensive structure control approach. By using a water-miscible organic solvent tetrahydrofuran (THF), worm-shaped micellar nanocrystals can be produced due to solvent-induced/facilitated micelle fusion. Compared with the common spherical micellar nanocrystals, worm-shaped micellar nanocrystals can offer minimized non-specific cellular uptake, thus enhancing biological targeting. By co-encapsulating multiple nanocrystals into each micelle, multifunctional or synergistic effects can be achieved. Current limitations of this fabrication method, which will be part of the future work, primarily include imperfect encapsulation in the micellar nanocrystal product and the incompletely continuous nature of the process.

The present work describes a method to fabricate micellar nanocrystals, an emerging major class of nanobiomaterials. This method combines top-down electrospray, bottom-up self-assembly, and solvent-based structure control. The fabrication method is largely continuous, can produce high quality products, and possesses an inexpensive means of structure control.

Micellar nanocrystals (micelles with encapsulated nanocrystals) have become an emerging major class of nanobiomaterials. We describe a method of ...

Synthesis of Monodisperse Cylindrical Nanoparticles via Crystallization-driven Self-assembly of Biodegradable Block Copolymers

The production of monodisperse cylindrical micelles is a significant challenge in polymer chemistry. Most cylindrical constructs formed from diblock copolymers are produced by one of three techniques: thin film rehydration, solvent switching or polymerization-induced self-assembly, and produce only flexible, polydisperse cylinders. Crystallization-driven self-assembly (CDSA) is a method which can produce cylinders with these properties, by stabilizing structures of a lower curvature due to the formation of a crystalline core. However, the living polymerization techniques by which most core-forming blocks are formed are not trivial processes and the CDSA process may yield unsatisfactory results if carried out incorrectly. Here, the synthesis of cylindrical nanoparticles from simple reagents is shown. The drying and purification of reagents prior to a ring-opening polymerization of &#949;-caprolactone catalyzed by diphenyl phosphate is described. This polymer is then chain extended by methyl methacrylate (MMA) followed by N,N-dimethyl acrylamide (DMA) using reversible addition&#8722;fragmentation chain-transfer (RAFT) polymerization, affording a triblock copolymer that can undergo CDSA in ethanol. The living CDSA process is outlined, the results of which yield cylindrical nanoparticles up to 500 nm in length and a length dispersity as low as 1.05. It is anticipated that these protocols will allow others to produce cylindrical nanostructures and elevate the field of CDSA in the future.

Crystallization-driven self-assembly (CDSA) displays the unique ability to fabricate cylindrical nanostructures of narrow length distributions. The organocatalyzed ring-opening polymerization of &#949;-caprolactone and subsequent chain extensions of methyl methacrylate and N,N-dimethyl acrylamide are demonstrated. A living CDSA protocol that produces monodisperse cylinders up to 500 nm in length is outlined.

The production of monodisperse cylindrical micelles is a significant challenge in polymer chemistry. Most cylindrical constructs formed from diblock ...

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.

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

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.

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

Facile Preparation and Photoactivation of Prodrug-Dye Nanoassemblies

Self-assembly is a simple yet reliable method for constructing nanoscale drug delivery systems. Photoactivatable prodrugs enable controllable drug release from nanocarriers at target sites modulated by light irradiation. In this protocol, a facile method for fabricating photoactivatable prodrug-dye nanoparticles via molecular self-assembly is presented. The procedures for prodrug synthesis, nanoparticle fabrication, physical characterization of the nanoassembly, photocleavage demonstration, and in vitro cytotoxicity verification are described in detail. A photocleavable boron-dipyrromethene-chlorambucil (BC) prodrug was first synthesized. BC and a near-infrared dye, IR-783, at an optimized ratio, could self-assemble into nanoparticles (IR783/BC NPs). The synthesized nanoparticles had an average size of 87.22 nm and a surface charge of -29.8 mV. The nanoparticles disassembled upon light irradiation, which could be observed by transmission electronic microscopy. The photocleavage of BC was completed within 10 min, with a 22% recovery efficiency for chlorambucil. The nanoparticles displayed enhanced cytotoxicity under light irradiation at 530 nm compared with the non-irradiated nanoparticles and irradiated free BC prodrug. This protocol provides a reference&#160;for the construction and evaluation of photoresponsive drug delivery systems.

This protocol describes the fabrication and characterization of a photoresponsive prodrug-dye nanoassembly. The methodology for drug release from the nanoparticles by light-triggered disassembly, including the light irradiation setup, is explicitly described. The drugs released from the nanoparticles following light irradiation exhibited excellent anti-proliferation effects on human colorectal tumor cells.

Self-assembly is a simple yet reliable method for constructing nanoscale drug delivery systems. Photoactivatable prodrugs enable controllable drug release ...

A 'Plug and Play' Method to Create Water-dispersible Nanoassemblies Containing an Amphiphilic Polymer, Organic Dyes and Upconverting Nanoparticles

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