Z.P.G. and F.S. thank the Swiss National Science Foundation and, specifically, NCCR &#39;Molecular Systems Engineering&#39;. Z.L. and F.S. thank the support of the Swiss National Science Foundation Division II grant. All authors thank Quy Ong for fruitful discussions and for proofreading the manuscript.

1. Synthesis of 11-mercapto-1-undecanesulfonate (MUS)
NOTE: This protocol can be used at any scale desired. Here, a 10 g scale-of-product is described.
<ol>
	<li>Sodium undec-10-enesulfonate 
	<ol>
		<li>Add 11-bromo-1-undecene (25 mL, 111.975 mmol), sodium sulfite (28.75 g, 227.92 mmol), and benzyltriethylammonium bromide (10 mg) to a mixture of 200 mL of methanol (MeOH) and 450 mL of deionized (DI) water (4:9 v/v MeOH:H2O ratio) in a 1 L round-bottom flask.</li>
		<li>Reflux the reaction mixture at 102 &#176;C for 48 h. Cap the system with a pressure relief mechanism&#8212;for example, a balloon with a needle, or simply a needle. This reaction is not sensitive to atmospheric gases. 
		NOTE: The solution becomes colorless when the reaction is complete.</li>
		<li>Connect the reaction mixture to a rotary evaporator to evaporate MeOH and reduce the volume to approximately 300 mL.</li>
		<li>Transfer the remaining solution to a 1 L addition funnel.</li>
		<li>Extract the remaining aqueous solution 5x with diethyl ether, using the addition funnel. Unreacted 11-bromo-1-undecene stays in the diethyl ether phase and the sulfonated product in H2O. 
		CAUTION: Release any pressure buildup frequently during the extraction, and consult the correct usage of addition funnels.</li>
		<li>Collect the final extracted water solution into a 1 L single-neck round-bottom flask.</li>
		<li>Connect the reaction flask to a rotary evaporator by putting a bit of grease (or Teflon ring strips or any other sealant) between the flask and the trap.</li>
		<li>Decrease the vacuum slowly to evaporate the aqueous phase in a rotary evaporator. Because the product is a surfactant, foaming will occur during the evaporation. To circumvent this problem, follow the instruction in the next step.
		<ol>
			<li>Add ethanol to the mixture to accelerate the evaporation of H2O and prevent foaming. When foaming restarts because of the decrease in ethanol content, stop the evaporation, remove the flask from the rotary evaporator, add more ethanol (about one-third of the total volume), and reconnect the flask to the rotary evaporator. Repeat this process until the solution mixture decreases significantly and does not form bubbles.</li>
		</ol>
		</li>
		<li>Dry the white powder directly by connecting the flask to a high vacuum. The drier the powder, the less inorganic salts will creep into the subsequent steps. 
		NOTE: Heat can be used to dry the product&#8212;for example, by keeping the flask under vacuum in a 60 &#176;C bath and left overnight.</li>
		<li>Suspend the white powder in 400 mL of methanol in a flask. Sonicate to dissolve the maximum amount of product. 
		NOTE: The goal of this step is to dissolve the product but not the inorganic byproducts, such as excess sodium sulfite and sodium bromide, that have limited solubility in methanol. Use methanol with the lowest water content possible, because water in the methanol will increase the solubility of the inorganic byproducts in the solvent.</li>
		<li>To increase the solubility of the product, methanol can be gently heated close to its boiling point (~64 &#176;C). 
		CAUTION: Make sure to work under a fume hood during the heating of the flask. The fumes of the evaporated methanol are dangerous.</li>
		<li>Filter the solution to remove the methanol insoluble inorganic byproducts. Use a filtering flask connected to a vacuum pump and a filtering funnel with quantitative filter paper, or a borosilicate filter. Both the product and the inorganic salts are white powders when dry: the product is soluble in methanol, while the salts are not.</li>
		<li>Transfer the filtered solution from the filtering flask to a 1 L round-bottom flask.</li>
		<li>Connect the flask to a rotary evaporator and evaporate the methanolic solution at 45 &#176;C, redissolve the white powder in methanol, and filter the solution (protocol steps 1.1.7, 1.1.8, and 1.1.9). Repeat this process at least 2x, to decrease the amount of inorganic salt.</li>
		<li>Collect the white, methanol soluble powder (approximately 30 g, at this scale).</li>
		<li>Dissolve approximately 10 mg of product in 500 &#181;L of D2O and transfer the solution to an NMR tube.</li>
		<li>Perform 1H NMR spectrometry on the product in D2O at 400 MHz with 32 scans. 
		NOTE: The peak assignments for 1H NMR (D2O) are 5.97 (m, 1H), 5.09 (m, 2H), 2.95 (t, 2H), 2.10 (m, 2H), 1.77 (q, 2H), 1.44 (br s, 12H).</li>
	</ol>
	</li>
	<li>Sodium 11-acetylthio-undecanesulfonate
	<ol>
		<li>Dissolve the approximate 30 g of sodium undec-10-enesulfonate (the reaction product of section 1.1) in 500 mL of methanol inside a 1 L round-bottom flask. Add a 2.6x excess of thioacetic acid to the solution and stir it in front of a UV lamp (250 W) overnight (~12 h). In case a UV lamp is not available, the reaction can be performed by refluxing using a radical initiator, such as azobisisobutyronitrile (AIBN); however, the use of a UV lamp is strongly recommended. 
		CAUTION: Make sure to work under the fume hood at all times. If the flask needs to be transported to another space where the UV lamp is located, seal the flask to avoid spreading the strong odor of thioacetic acid. Exercise care when operating a UV lamp: completely block the space where the lamp is located and consult the institution&#8217;s safety guidelines on how to operate a UV lamp.</li>
		<li>Monitor the reaction by taking ~2 mL aliquots from the reaction, evaporate solvent, and add deuterated water to check with 1H NMR. Once the peaks corresponding to the double bond disappear, stop the reaction. 
		NOTE: Usually, after 12 h in front of the UV lamp, the reaction is complete. If the reaction mixture becomes turbid, add more MeOH and continue the exposure to the UV light for six additional hours.</li>
		<li>Evaporate all of MeOH in a rotary evaporator until the solid residue becomes orange-red. If left long enough, the product becomes brown to black. 
		CAUTION: Work mindfully because of the strong odors from the thioacetic acid. The strong odors of any thiolate spills can be neutralized using an aqueous solution of bleach (sodium hypochlorite).</li>
		<li>Using a filtering flask, wash the product with diethyl ether to remove any excess thioacetic acid, until no more colored (orange-yellow) substances appear in the diethyl ether supernatant. Dry the solid under high vacuum and, then, dissolve it in methanol, yielding a yellow to orange solution. 
		NOTE:&#160;Add enough methanol to dissolve the product. 
		NOTE: The color may vary at this step.&#160;</li>
		<li>Add 3 g of carbon black to the solution, mix vigorously, and filter the mixture through filtration medium (see Table of Materials) covering two-thirds of a fluted filter paper. 
		NOTE: The porous structure of carbon black captures the colored side-product material (and some of the product). The filtered solution should be clear. If the filtered solution is still colored (yellow), repeat this process.</li>
		<li>Evaporate the solvent completely in a rotary evaporator and collect approximately 35 g of white powder.</li>
		<li>Dissolve ~10 mg of the product in ~500 &#181;L of D2O and transfer the solution to NMR tubes.</li>
		<li>Perform 1H NMR on the product in D2O at 400 MHz with 32 scans. 
		NOTE: The peak assignments for the H NMR (D2O) are 2.93 (t, 4H), 2.40 (s, 3H), 1.77 (m, 2H), 1.62 (m, 2H), 1.45 (br s, 14H).</li>
	</ol>
	</li>
	<li>11-mercapto-1-undecanesulfonate (MUS)
	<ol>
		<li>Reflux sodium 11-acetylthio-undecanesulfonate at 102 &#176;C in 400 mL of 1 M HCl for 12 h to cleave the thioacetate group and obtain a thiol.</li>
		<li>Transfer the product to a 1.5 L or 2 L round-bottom flask. Add 200 mL of 1 M NaOH to the final solution and top it with 400 mL of DI water to have a final volume of 1 L. This will keep the solution acidic and prevent the crystallization of inorganic salts as byproduct. 
		NOTE: A complete neutralization of the solution to pH 7 will result in the crystallization of a product insoluble in methanol.</li>
		<li>Keep the clear solution at 4 &#176;C and it will crystallize overnight. The product crystallizes as fine crystals that are viscous when wet. 
		NOTE: To accelerate the crystallization, add presynthesized MUS to the solution, if available.</li>
		<li>Decant the clear supernatant and centrifuge down the viscous white product in 50 mL centrifuge tubes for 5 min at 4,000 x g.</li>
		<li>Decant the supernatant into another flask and dry the white pellets under high vacuum&#8212;depending on the centrifuge available, this can be 2 - 16 tubes or more. 
		NOTE: Filtering is not advised because of the surfactant nature of the product; excessive foaming will occur and most of the product will be lost.</li>
		<li>Collect approximately 12 g (about 30% yield) of methanol-soluble MUS from this purification step. 
		NOTE: Be mindful that the powder is fine and electrostatic&#8212;it tends to stick to spatulas and the surfaces of containers. Also, more material can be extracted from the supernatant of the centrifugation step by reducing the volume (to about a third of its original value) and keeping it at 4 &#176;C. Decrease the volume even more (by 75%) to increase the yield at this step.</li>
		<li>Dissolve ~10 mg of the product in ~500 &#181;L of D2O and transfer the solution to NMR tubes.</li>
		<li>Perform 1H NMR on the product in D2O at 400 MHz with 32 scans. 
		NOTE: The peak assignments of H NMR (D2O) are 2.93 (t, 4H), 2.59 (t, 3H), 1.78 (m, 2H), 1.65 (m, 2H), 1.44 (br s, 14H). The calculated molar mass (including the sodium counterion) of the product is 290.42 g/mol.</li>
	</ol>
	</li>
</ol>
2. Nanoparticle Synthesis: Preparation of the Reagents
<ol>
	<li>Clean all glassware (one 250 mL and one 500 mL single-neck round-bottom flask, a 100&#160;mL addition funnel, and a small funnel) with fresh aqua regia (three parts hydrochloric acid to one part nitric acid). Rinse the glassware with an excess amount of water inside a fume hood and remove all the fumes. Then, rinse the glassware with ethanol and dry it in a laboratory glassware oven (40 - 60 &#176;C is recommended).</li>
	<li>Weigh 177.2&#160;mg (0.45 mmol) of gold (III) chloride trihydrate (HAuCl4&#8729;3H2O) in a small glass vial (10 or 20 mL clean glass vials, or on weighing paper).</li>
	<li>Weigh 87 mg (0.3 mmol) MUS in a glass vial of 20 mL.</li>
	<li>Add 10 mL of methanol to dissolve the MUS. Sonicate it in an ultrasonic bath until no solid material is visible, to ensure complete dissolution. 
	NOTE: Alternatively, using a heat gun or a warm bath (~60 &#176;C), heat the solution gently. When heated, run cold water through the outside of the flask to bring it back to room temperature.</li>
	<li>Add 26 &#181;L (0.15 mmol) of OT to the methanol solution and agitate it to mix the ligands.</li>
	<li>Weigh 500 mg (13 mmol) of sodium borohydride (NaBH4) and add it to 100 mL of ethanol in the 250 mL round-bottom flask. Stir vigorously using magnetic stirring (600 - 800 rpm). (The NaBH4 takes 10 to 20 min, depending on the grade, to form a clear solution in ethanol.)</li>
</ol>
3. Synthesis of Gold Nanoparticles
<ol>
	<li>Dissolve gold salt in 100 mL of ethanol in the 500 mL round-bottom flask and start stirring at 800 rpm with a magnetic bar on a stirring plate. Make sure the gold salt dissolves completely.</li>
	<li>Place a 100 mL addition funnel above the round-bottom flask. Put a funnel on the top of the addition funnel with a quantitative paper filter inside. When the NaBH4 is dissolved in ethanol, start filtering the solution into the addition funnel through the filter paper in the funnel.</li>
	<li>Add the ligand solution to the reaction mixture. Wait 15 min for the formation of gold-thiolate complex. The color change of the reaction mixture from translucent yellow to turbid yellow indicates the formation of gold-thiolate complex.</li>
	<li>Start adding the filtered NaBH4 solution from the addition funnel dropwise. Adjust the interval time of the drops so that the addition of NaBH4 takes about 1 h.</li>
	<li>After the complete addition of NaBH4, remove the funnel. Keep stirring the reaction for another hour. At the end of the reaction, remove the magnetic stirring bar using a magnet placed on the outside of the flask.</li>
	<li>Use a septum to close the flask and pierce a needle into the septum to release the H2 gas that will evolve after the reaction.</li>
	<li>Keep the reaction mixture inside a laboratory refrigerator (4 &#176;C) to precipitate the nanoparticles overnight.</li>
</ol>
4. Workup of the Synthesis
<ol>
	<li>Decant the supernatant ethanol to reduce the volume.</li>
	<li>Transfer the remaining precipitant to 50 mL centrifuge tubes and centrifuge for 3 min at 4,000 x g.</li>
	<li>Decant the supernatant, disperse the nanoparticles again with ethanol by vortexing, and centrifuge them again. Repeat this washing process 4x.</li>
	<li>Dry the nanoparticles under vacuum to remove the residual ethanol.</li>
	<li>To clean the nanoparticles from free hydrophilic ligands/molecules, dissolve the precipitates in 15 mL of DI water and transfer them to the centrifuge tubes with a filtration membrane of 30 kDa cutoff molecular weight. Dialysis is also amenable for this procedure.</li>
	<li>Centrifuge these tubes for 5 min at 4,000 x g to concentrate the nanoparticle solution.</li>
	<li>Add 15 mL of DI water to this solution and centrifuge to concentrate again. Repeat this cleaning process at least 10x. 
	NOTE: One indication that the water-soluble impurities have been removed is the absence of foaming when agitating the aqueous waste; after all, most of the impurities are disulfides of MUS with itself or with OT (this can be determined by collecting the material and performing 1H NMR).</li>
	<li>After the centrifugation, transfer the concentrated nanoparticles to a 15 mL centrifuge tube. To turn the nanoparticles into a manageable powder, either precipitate them in a solvent such as acetone or freeze-dry the remaining aqueous solution. When freeze-dried, the nanoparticles tend to form a loose powder that sticks to surfaces and may be difficult to manipulate.</li>
</ol>
5. Characterization of the Nanoparticles
<ol>
	<li>Purity
	<ol>
		<li>To check whether the nanoparticles are free from unbound ligands, dissolve 5 mg of dry nanoparticles in 600 &#181;L of D2O and perform a 1H NMR measurement of the particles. If there are no sharp peaks of the ligands, it means the nanoparticles are free from small organic molecules.</li>
	</ol>
	</li>
	<li>Ligand ratio
	<ol>
		<li>Prepare a 20&#160;mg/mL methanol-d4 solution of iodine. Add 600 &#181;L of this solution to the ~5 mg of nanoparticles in a glass vial, to etch the nanoparticles.</li>
		<li>Wrap the cap of the vial with paraffin film and sonicate it in an ultrasonic bath for 20 min. Transfer the solution to an NMR tube and acquire a 1H NMR (400 MHz) spectrum with 32 scans.</li>
	</ol>
	</li>
	<li>Ligand density
	<ol>
		<li>Transfer 2 to 8 mg of nanoparticles to a TGA crucible. Choose a temperature range from 30 &#176;C to 900 &#176;C and a speed of 5 &#176;C per minute under N2 gas.</li>
	</ol>
	</li>
	<li>Size distribution
	<ol>
		<li>TEM
		<ol>
			<li>Prepare 0.1 mg/mL nanoparticle solution in DI water. Drop 5 &#181;L of the prepared solution onto the 400-mesh carbon-supported copper grid. Wait until it dries.</li>
			<li>Transfer the grid in a TEM holder and insert it into the microscope. Acquire 5 - 10 images with a magnification of at least 64,000X, operated at 200 kV. 
			NOTE: To increase the contrast, an objective aperture of 20 nm can be inserted.</li>
		</ol>
		</li>
		<li>UV-Vis spectra
		<ol>
			<li>Prepare a 0.2 mg/mL nanoparticle solution in DI water.</li>
			<li>Put the required amount of this solution in the quartz cuvette and scan from 200 nm to 700 nm.</li>
		</ol>
		</li>
	</ol>
	</li>
</ol>

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The authors have nothing to disclose.

This protocol describes first the synthesis of MUS ligand and, then, the synthesis and characterization of amphiphilic MUS:OT gold nanoparticles. Synthesizing MUS with minimal salt content enables a better reliability of the stoichiometric ratio between the ligands during the nanoparticle synthesis, which is a key factor for the reproducible synthesis of MUS:OT nanoparticles with a target hydrophobic content (Figure 8). The use of methanol as a common solvent for MUS and OT, along with the s...

The ligand shell of gold nanoparticles can be engineered to exhibit several different properties that can be applied to address challenges in biomedicine1,2,3,4. Such versatility allows for the control of the intermolecular interactions between nanoparticles and biomolecules5,6,7. Hydrophobicity and charge play a decisive role, as well as other surface parameters that affect how nanoparticles interact with biomolecules5,8,9. To tune the nanoparticles&#39; surface properties, the choice of thiolate molecules that make up the ligand shell offers a myriad of possibilities, according to the characteristics sought. For example, a mixture of ligand molecules with hydrophobic and hydrophilic (e.g.,&#160;charged) end groups are often used to generate amphiphilic nanoparticles10,11.
One prominent example of this type of nanoparticles is protected by a mixture of OT and MUS (hereafter called MUS:OT nanoparticles) that has been shown to possess many relevant properties12,13,14. First, with a ligand shell composition of 66% MUS (hereafter 66:34 MUS:OT), the colloidal stability of the nanoparticles is high, reaching up to 33% in weight in deionized water, as well as in phosphate-buffered saline (1x, 4 mM phosphate, 150 mM NaCl)15. Moreover, these particles do not precipitate at relatively low pH values: for example, at pH 2.3 and with salt concentrations of 1 M NaCl15, these nanoparticles remain colloidally stable for months.The stoichiometric ratio between the two molecules on the ligand shell is important because it dictates the colloidal stability in solutions with a high ionic strength16.
These particles have been shown to traverse the cell membrane without porating it, via an energy-independent pathway1,12. The spontaneous fusion between these particles and lipid bilayers underlies their diffusivity through cell membranes17. The mechanism behind this interaction is the minimization of contact between a hydrophobic solvent-accessible surface area and water molecules upon fusion with lipid bilayers18. Compared to all-MUS nanoparticles (nanoparticles having only the MUS ligand on their shell), the higher hydrophobicity on mixed MUS:OT nanoparticles (for example, at a 66:34 MUS:OT composition) increases the span of the core diameter that can fuse with lipid bilayers18.Different self-assembly organizations of the ligand shell correlate to distinct binding modes of 66:34 MUS:OT nanoparticles with various proteins, such as albumin and ubiquitin, when compared to all-MUS particles19.Recently, it has been reported that 66:34 MUS:OT nanoparticles can be utilized as a broad-spectrum antiviral agent that irreversibly destroys the viruses because of multivalent electrostatic bindings of MUS ligands and nonlocal couplings of OT ligands to capsid proteins14.In all these cases, it has been found that the hydrophobic content, as well as the core size of the nanoparticles, determines how these bio-nano interactions take place. These diverse properties of MUS:OT nanoparticles have prompted many computer simulation studies that aimed to clarify the mechanisms underpinning the interactions between MUS:OT particles and various biological structures such as lipid bilayers20.
The preparation of MUS:OT-protected Au nanoparticles poses a few challenges. First, the charged ligand (MUS) and the hydrophobic ligand (OT) are immiscible. Thus, the solubility of the nanoparticles and of the ligands needs to be taken into account throughout the synthesis, as well as during characterization. Additionally, the purity of the MUS ligand molecules&#8212;specifically, the content of inorganic salts in the starting material&#8212;influences the quality, reproducibility, as well as the short- and long-term colloidal stability of the nanoparticles.
Here, a detailed synthesis and characterization of this class of amphiphilic gold nanoparticles protected by a mixture of MUS and OT are outlined. A protocol for the synthesis of the negatively charged MUS ligand is reported to ensure the purity and, hence, the reproducibility of different nanoparticle syntheses. Then, the procedure to generate these nanoparticles, based on a common one-phase synthesis, followed by thorough purification, is reported in detail. Various necessary characterization techniques21, such as TEM, UV-Vis, TGA, and NMR, have been combined to obtain all the necessary parameters for any further biological experiments.

<table><tbody><tr><td>11-bromo-1-undecene</td><td>Sigma Aldrich</td><td>467642-25 ml</td><td></td></tr><tr><td>Sodium Sulfite</td><td>Sigma Aldrich</td><td>S0505-250 g</td><td></td></tr><tr><td>Benzyltriethyl-ammonium bromide</td><td>Sigma Aldrich</td><td>147125-25 g</td><td></td></tr><tr><td>Methanol</td><td>VWR</td><td>BDH1135-2.5 LP</td><td></td></tr><tr><td>DI water</td><td>Millipore</td><td>ZRXQ003WW</td><td>Deionized water</td></tr><tr><td>1 L round bottom flask</td><td>DURAN</td><td>24 170 56</td><td></td></tr><tr><td>Diethyl ether</td><td>Sigma Aldrich</td><td>1.00930 EMD Millipore</td><td></td></tr><tr><td>Stirring bar</td><td>Sigma Aldrich</td><td>Z329207,</td><td></td></tr><tr><td>Dow Corning High Vacuum Grease</td><td>Sigma Aldrich</td><td>Z273554 ALDRICH</td><td></td></tr><tr><td>Filtering flask</td><td>DURAN</td><td>20 201 63</td><td></td></tr><tr><td>Filtering Buchner Funnel</td><td>FisherSci</td><td>11707335</td><td></td></tr><tr><td>Ethanol &gt;99.8%, ACS, Reagent</td><td>VWR</td><td>2081.321DP</td><td></td></tr><tr><td>Deuterium dioxide</td><td>Sigma Aldrich</td><td>151882 ALDRICH</td><td></td></tr><tr><td>Thioacetic acid 96%</td><td>Sigma Aldrich</td><td>T30805 ALDRICH</td><td></td></tr><tr><td>Carbon black</td><td>Sigma Aldrich</td><td>05105-1KG</td><td></td></tr><tr><td>Celite</td><td>Sigma Aldrich</td><td>D3877 SIGMA-ALDRICH</td><td>Filtration medium</td></tr><tr><td>Condenser</td><td>Sigma Aldrich</td><td>Z531154</td><td></td></tr><tr><td>Hydrochloric acid, ACS reagent 37%</td><td>Sigma Aldrich</td><td>320331 SIGMA-ALDRICH</td><td></td></tr><tr><td>Sodium Hydroxide, BioXtra, pellets (anhydrous)</td><td>Sigma Aldrich</td><td>S8045 SIGMA-ALDRICH</td><td></td></tr><tr><td>Centrifuge tubes</td><td>VWR</td><td>525-0155P</td><td></td></tr><tr><td>250 mL round bottom flask</td><td>DURAN</td><td>24 170 37</td><td></td></tr><tr><td>500 mL round bottom flask</td><td>DURAN</td><td>24 170 46</td><td></td></tr><tr><td>Nitric acid, fACS reagent 70%</td><td>Sigma Aldrich</td><td>438073 SIGMA-ALDRICH</td><td></td></tr><tr><td>Gold(III) chloride trihydrate &gt;99.9% trace metal basis</td><td>Sigma Aldrich</td><td>520918 ALDRICH</td><td></td></tr><tr><td>1-octanethiol &gt;98.5%</td><td>Sigma Aldrich</td><td>471836 ALDRICH</td><td></td></tr><tr><td>Sodium Borohydride purum p.a.&gt;96%</td><td>Sigma Aldrich</td><td>71320 ALDRICH</td><td></td></tr><tr><td>addition funnel</td><td>SIgma Aldrich</td><td>Z330655 SIGMA</td><td></td></tr><tr><td>Funnel</td><td>DURAN</td><td>21 351 46</td><td></td></tr><tr><td>2V folded filtering papers</td><td>Whatman</td><td>1202-150</td><td></td></tr><tr><td>Amicon filters</td><td>Merck</td><td>UFC903024</td><td></td></tr><tr><td>Iodine, ACS reagent, &gt;99.8%, solid</td><td>Sigma Aldrich</td><td>207772 SIGMA-ALDRICH</td><td></td></tr><tr><td>5 mm NMR-Tubes, Type 5HP (high precision)</td><td>Armar</td><td>32210.503</td><td>Length 178 mm</td></tr><tr><td>Methanol-d4 99.8 atom%D</td><td>Armar</td><td>16400.2035</td><td></td></tr><tr><td>TGA crucible</td><td>Thepro</td><td>9095-9270.45</td><td></td></tr><tr><td>400 mesh carbon supported copper grid</td><td>Electron Microscopy Science</td><td>CF400-Cu</td><td></td></tr><tr><td>quartz cuvette</td><td>Hellma Analytics</td><td>100-1-40</td><td></td></tr></tbody></table>

synthesis and characterization of amphiphilic gold nanoparticles

Gold nanoparticles covered with a mixture of 1-octanethiol (OT) and 11-mercapto-1-undecane sulfonic acid (MUS) have been extensively studied because of their interactions with cell membranes, lipid bilayers, and viruses. The hydrophilic ligands make these particles colloidally stable in aqueous solutions and the combination with hydrophobic ligands creates an amphiphilic particle that can be loaded with hydrophobic drugs, fuse with the lipid membranes, and resist nonspecific protein adsorption. Many of these properties depend on nanoparticle size and the composition of the ligand shell. It is, therefore, crucial to have a reproducible synthetic method and reliable characterization techniques that allow the determination of nanoparticle properties and the ligand shell composition. Here, a one-phase chemical reduction, followed by a thorough purification to synthesize these nanoparticles with diameters below 5 nm, is presented. The ratio between the two ligands on the surface of the nanoparticle can be tuned through their stoichiometric ratio used during synthesis. We demonstrate how various routine techniques, such as transmission electron microscopy (TEM), nuclear magnetic resonance (NMR), thermogravimetric analysis (TGA), and ultraviolet-visible (UV-Vis) spectrometry, are combined to comprehensively characterize the physicochemical parameters of the nanoparticles.

The reaction steps to synthesize MUS are shown in Figure 1. The 1H NMR spectra of the product of each step are represented in Figure 2. The synthesis workflow of the binary MUS:OT amphiphilic gold nanoparticles is described in Figure 3. Following the synthesis, the workup of the nanoparticles consisted of washing the particles several times with ethanol and DI water. Prior to any character...

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Synthesis and Characterization of Amphiphilic Gold Nanoparticles

Amphiphilic gold nanoparticles can be used in many biological applications. A protocol to synthesize gold nanoparticles coated by a binary mixture of ligands and a detailed characterization of these particles is presented.

Gold nanoparticles covered with a mixture of 1-octanethiol (OT) and 11-mercapto-1-undecane sulfonic acid (MUS) have been extensively studied because of ...

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Chemistry

17.4K Views.  École Polytechnique Fédérale de Lausanne. Amphiphilic gold nanoparticles can be used in many biological applications. A protocol to synthesize gold nanoparticles coated by a binary mixture of ligands and a detailed characterization of these particles is presented.

Video: Synthesis and Characterization of Amphiphilic Gold Nanoparticles

Synthesis, Assembly, and Characterization of Monolayer Protected Gold Nanoparticle Films for Protein Monolayer Electrochemistry

Colloidal gold nanoparticles protected with alkanethiolate ligands called monolayer protected gold clusters (MPCs) are synthesized and subsequently incorporated into film assemblies that serve as adsorption platforms for protein monolayer electrochemistry (PME). PME is utilized as the model system for studying electrochemical properties of redox proteins by confining them to an adsorption platform at a modified electrode, which also serves as a redox partner for electron transfer (ET) reactions. Studies have shown that gold nanoparticle film assemblies of this nature provide for a more homogeneous protein adsorption environment and promote ET without distance dependence compared to the more traditional systems modified with alkanethiol self-assembled monolayers (SAM).1-3 In this paper, MPCs functionalized with hexanethiolate ligands are synthesized using a modified Brust reaction4 and characterized with ultraviolet visible (UV-Vis) spectroscopy, transmission electron microscopy (TEM), and proton (1H) nuclear magnetic resonance (NMR). MPC films are assembled on SAM modified gold electrode interfaces by using a "dip cycle" method of alternating MPC layers and dithiol linking molecules. Film growth at gold electrode is tracked electrochemically by measuring changes to the double layer charging current of the system. Analogous films assembled on silane modified glass slides allow for optical monitoring of film growth and cross-sectional TEM analysis provides an estimated film thickness. During film assembly, manipulation of the MPC ligand protection as well as the interparticle linkage mechanism allow for networked films, that are readily adaptable, to interface with redox protein having different adsorption mechanism. For example, Pseudomonas aeruginosa azurin (AZ) can be adsorbed hydrophobically to dithiol-linked films of hexanethiolate MPCs and cytochrome c (cyt c) can be immobilized electrostatically at a carboxylic acid modified MPC interfacial layer. In this report, we focus on the film protocol for the AZ system exclusively. Investigations involving the adsorption of proteins on MPC modified synthetic platforms could further the understanding of interactions between biomolecules and man-made materials, and consequently aid the development of biosensor schemes, ET modeling systems, and synthetic biocompatible materials.5-8

Alkanethiolate stabilized gold colloids known as monolayer protected clusters (MPCs) are synthesized, characterized, and assembled into thin films as an adsorption interface for protein monolayer electrochemistry of simple redox protein like Pseudomonas aeruginosa azurin (AZ) and cytochrome c (cyt c).

Colloidal gold nanoparticles protected with alkanethiolate ligands called monolayer protected gold clusters (MPCs) are synthesized and subsequently ...

Bioengineering

Gold Nanostar Synthesis with a Silver Seed Mediated Growth Method

The physical, chemical and optical properties of nano-scale colloids depend on their material composition, size and shape 1-5. There is a great interest in using nano-colloids for photo-thermal ablation, drug delivery and many other biomedical applications 6. Gold is particularly used because of its low toxicity 7-9. A property of metal nano-colloids is that they can have a strong surface plasmon resonance 10. The peak of the surface plasmon resonance mode depends on the structure and composition of the metal nano-colloids. Since the surface plasmon resonance mode is stimulated with light there is a need to have the peak absorbance in the near infrared where biological tissue transmissivity is maximal 11, 12.
We present a method to synthesize star shaped colloidal gold, also known as star shaped nanoparticles 13-15 or nanostars 16. This method is based on a solution containing silver seeds that are used as the nucleating agent for anisotropic growth of gold colloids 17-22. Scanning electron microscopy (SEM) analysis of the resulting gold colloid showed that 70 % of the nanostructures were nanostars. The other 30 % of the particles were amorphous clusters of decahedra and rhomboids. The absorbance peak of the nanostars was detected to be in the near infrared (840 nm). Thus, our method produces gold nanostars suitable for biomedical applications, particularly for photo-thermal ablation.

We synthesized star shaped gold nanostars using a silver seed mediated growth method. The diameter of the nanostars ranges from 200 to 300 nm and the number of tips vary from 7 to 10. The nanoparticles have a broad surface plasmon resonance mode centered in the near infrared.

The physical, chemical and optical properties of nano-scale colloids depend on their material composition, size and shape 1-5. There is a great interest ...

Synthesis, Characterization, and Functionalization of Hybrid Au/CdS and Au/ZnS Core/Shell Nanoparticles

Plasmonic nanoparticles are an attractive material for light harvesting applications due to their easily modified surface, high surface area and large extinction coefficients which can be tuned across the visible spectrum. Research into the plasmonic enhancement of optical transitions has become popular, due to the possibility of altering and in some cases improving photo-absorption or emission properties of nearby chromophores such as molecular dyes or quantum dots. The electric field of the plasmon can couple with the excitation dipole of a chromophore, perturbing the electronic states involved in the transition and leading to increased absorption and emission rates. These enhancements can also be negated at close distances by energy transfer mechanism, making the spatial arrangement of the two species critical. Ultimately, enhancement of light harvesting efficiency in plasmonic solar cells could lead to thinner and, therefore, lower cost devices. The development of hybrid core/shell particles could offer a solution to this issue. The addition of a dielectric spacer between a gold nanoparticles and a chromophore is the proposed method to control the exciton plasmon coupling strength and thereby balance losses with the plasmonic gains. A detailed procedure for the coating of gold nanoparticles with CdS and ZnS semiconductor shells is presented. The nanoparticles show high uniformity with size control in both the core gold particles and shell species allowing for a more accurate investigation into the plasmonic enhancement of external chromophores.

The synthesis of uniform gold nanoparticles coated with semiconductor shells of CdS or ZnS is performed. The semiconductor coating is conducted by first depositing a silver sulfide shell and exchanging the silver cations for zinc or cadmium cations.

Plasmonic nanoparticles are an attractive material for light harvesting applications due to their easily modified surface, high surface area and large ...

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

Reducing dilute aqueous HAuCl4 with sodium thiocyanate (NaSCN) under alkaline conditions produces 2 to 3 nm diameter nanoparticles. Stable grape-like oligomeric clusters of these yellow nanoparticles of narrow size distribution are synthesized under ambient conditions via two methods. The delay-time method controls the number of subunits in the oligoclusters by varying the time between the addition of HAuCl4 to alkaline solution and the subsequent addition of reducing agent, NaSCN. The yellow oligoclusters produced range in size from ~3 to ~25 nm. This size range can be further extended by an add-on method utilizing hydroxylated gold chloride (Na+[Au(OH4-x)Clx]-) to auto-catalytically increase the number of subunits in the as-synthesized oligocluster nanoparticles, providing a total range of 3 nm to 70 nm. The crude oligocluster preparations display narrow size distributions and do not require further fractionation for most purposes. The oligoclusters formed can be concentrated &#62;300 fold without aggregation and the crude reaction mixtures remain stable for weeks without further processing. Because these oligomeric clusters can be concentrated before derivatization they allow expensive derivatizing agents to be used economically. In addition, we present two models by which predictions of particle size can be made with great accuracy.

We describe a simple method for producing highly stable oligomeric clusters of gold nanoparticles via the reduction of chloroauric acid (HAuCl4) with sodium thiocyanate (NaSCN). The oligoclusters have a narrow size distribution and can be produced with a wide range of sizes and surface coats.

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

Hydroquinone Based Synthesis of Gold Nanorods

Gold nanorods are an important kind of nanoparticles characterized by peculiar plasmonic properties. Despite their widespread use in nanotechnology, the synthetic methods for the preparation of gold nanorods are still not fully optimized. In this paper we describe a new, highly efficient, two-step protocol based on the use of hydroquinone as a mild reducing agent. Our approach allows the preparation of nanorods with a good control of size and aspect ratio (AR) simply by varying the amount of hexadecyl trimethylammonium bromide (CTAB) and silver ions (Ag+) present in the "growth solution". By using this method, it is possible to markedly reduce the amount of CTAB, an expensive and cytotoxic reagent, necessary to obtain the elongated shape. Gold nanorods with an aspect ratio of about 3 can be obtained in the presence of just 50 mM of CTAB (versus 100 mM used in the standard protocol based on the use of ascorbic acid), while shorter gold nanorods are obtained using a concentration as low as 10 mM.

This paper describes a protocol for the synthesis of gold nanorods, based on the use of hydroquinone as reducing agent, plus the different mechanisms for controlling their size and aspect ratio.

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

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

Due to drug resistance and toxicity in healthy cells, use of doxorubicin (DOX) has been limited in clinical cancer therapy. This protocol describes the designing of poly(ethylenimine) grafted with polyethylene glycol (PEI-g-PEG) copolymer functionalized gold nanoparticles (AuNPs) with loaded aptamer (AS1411) and DOX through amide reactions. AS1411 is specifically bonded with targeted nucleolin receptors on cancer cells so that DOX targets cancer cells instead of healthy cells. First, PEG is carboxylated, then grafted to branched PEI to obtain a PEI-g-PEG copolymer, which is confirmed by 1H NMR analysis. Next, PEI-g-PEG copolymer coated gold nanoparticles (PEI-g-PEG@AuNPs) are synthesized, and DOX and AS1411 are covalently bonded to AuNPs gradually via amide reactions. The diameter of the prepared AS1411-g-DOX-g-PEI-g-PEG@AuNPs is ~39.9 nm, with a zeta potential of -29.3 mV, indicating that the nanoparticles are stable in water and cell medium. Cell cytotoxicity assays show that the newly designed DOX loaded AuNPs are able to kill cancer cells (A549). This synthesis demonstrates the delicate arrangement of PEI-g-PEG copolymers, aptamers, and DOX on AuNPs that are achieved by sequential amide reactions. Such aptamer-PEI-g-PEG functionalized AuNPs provide a promising platform for targeted drug delivery in cancer therapy.

In this protocol, doxorubicin-loaded AS1411-g-PEI-g-PEG modified gold nanoparticles are synthesized via three-step amide reactions. Then, doxorubicin is loaded and delivered to target cancer cells for cancer therapy.

Due to drug resistance and toxicity in healthy cells, use of doxorubicin (DOX) has been limited in clinical cancer therapy. This protocol describes the ...

Gold Nanoparticle Synthesis

Gold nanoparticles (Au nanoparticles) that are ~12 nm in diameter were synthesized by rapidly injecting a solution of 150 mg (0.15 mmol) of tetrachloroauric acid in 3.0 g (3.7 mmol, 3.6 mL) of oleylamine (technical grade) and 3.0 mL of toluene into a boiling solution of 5.1 g (6.4 mmol, 8.7 mL) of oleylamine in 147 mL of toluene. While boiling and mixing the reaction solution for 2 hours, the color of the reaction mixture changed from clear, to light yellow, to light pink, and then slowly to dark red. The heat was then turned off, and the solution was allowed to gradually cool down to room temperature for 1 hour. The gold nanoparticles were then collected and separated from the solution using a centrifuge and washed three times; by vortexing and dispersing the gold nanoparticles in 10 mL portions of toluene, and then precipitating the gold nanoparticles by adding 40 mL portions of methanol and spinning them in a centrifuge. The solution was then decanted to remove any remaining byproducts and unreacted starting materials. Drying the gold nanoparticles in a vacuum environment produced a solid black pellet; which could be stored for long periods of time (up to one year) for later use, and then redissolved in organic solvents such as toluene.

A protocol for synthesizing ~12 nm diameter gold nanoparticles (Au nanoparticles) in an organic solvent is presented. The gold nanoparticles are capped with oleylamine ligands to prevent agglomeration. The gold nanoparticles are soluble in organic solvents such as toluene.

Gold nanoparticles (Au nanoparticles) that are ~12 nm in diameter were synthesized by rapidly injecting a solution of 150 mg (0.15 mmol) of ...

Synthesis of Functionalized 10-nm Polymer-coated Gold Particles for Endothelium Targeting and Drug Delivery

Gold nanoparticles (AuNPs) have been used extensively in medical research due to their size, biocompatibility, and modifiable surface. Specific targeting and drug delivery are some of the applications of these AuNPs, but endothelial extracellular matrices&#39; defensive properties hamper particle uptake. To address this issue, we describe a synthesis method for ultrasmall gold nanoparticles to improve vascular delivery, with customizable functional groups and polymer lengths for further adjustments. The protocol yields 2.5 nm AuNPs that are capped with tetrakis(hydroxymethyl)phosphonium chloride (THPC). The replacement of THPC with hetero-functional polyethylene glycol (PEG) on the surface of the AuNP increases the hydrodynamic radius to 10.5 nm while providing various functional groups on the surface. The last part of the protocol includes an optional addition of a fluorophore to allow the AuNPs to be visualized under fluorescence to track nanoparticle uptake. Dialysis and lyophilization were used to purify and isolate the AuNPs. These fluorescent nanoparticles can be visualized in both in vitro and in vivo experiments due to the biocompatible PEG coating and fluorescent probes. Additionally, the size range of these nanoparticles render them an ideal candidate for probing the glycocalyx without disrupting normal vasculature function, which may lead to improved delivery and therapeutics.

We describe a method of synthesizing biocompatible 10-nm gold nanoparticles, functionalized by coating poly-ethylene glycol onto the surface. These particles can be used in vitro and in vivo for delivering therapeutics to nanoscale cellular and extracellular spaces that are difficult to access with conventional nanoparticle sizes.

Gold nanoparticles (AuNPs) have been used extensively in medical research due to their size, biocompatibility, and modifiable surface. Specific targeting ...

Aqueous Synthesis of Plasmonic Gold-Tin Alloy Nanoparticles

This protocol describes the synthesis of Au nanoparticle seeds and the subsequent formation of Au-Sn bimetallic nanoparticles. These nanoparticles have potential applications in catalysis, optoelectronics, imaging, and drug delivery. Previously, methods for producing alloy nanoparticles have been time-consuming, require complex reaction conditions, and can have inconsistent results. The outlined protocol first describes the synthesis of approximately 13 nm Au nanoparticle seeds using the Turkevich method. The protocol next describes the reduction of Sn and its incorporation into the Au seeds to generate Au-Sn alloy nanoparticles. The optical and structural characterization of these nanoparticles is described. Optically, prominent localized surface plasmon resonances (LSPRs) are apparent using UV-visible spectroscopy. Structurally, powder X-ray diffraction (XRD) reflects all particles to be less than 20 nm and shows patterns for Au, Sn, and multiple Au-Sn intermetallic phases. Spherical morphology and size distribution are obtained from transmission electron microscopy (TEM) imaging. TEM reveals that after Sn incorporation, the nanoparticles grow to approximately 15 nm in diameter.

Author Spotlight: Designing Sustainable Nanomaterials for Advancing Synthesis and Element Mixing

Here, the synthesis of gold (Au) seeds is described using the Turkevich method. These seeds are then used to synthesize gold-tin alloy (Au-Sn) nanoparticles with tunable plasmonic properties.

This protocol describes the synthesis of Au nanoparticle seeds and the subsequent formation of Au-Sn bimetallic nanoparticles. These nanoparticles have ...

Synthesis of Near-Infrared Emitting Gold Nanoclusters for Biological Applications

Over the past decade, fluorescent gold nanoclusters (AuNCs) have witnessed growing popularity in biological applications and enormous efforts have been devoted to their development. In this protocol, a recently developed, facile method for preparation of water soluble, biocompatible, and colloidally stable near-infrared emitting AuNCs have been described in detail. This room-temperature, bottom-up chemical synthesis provides easily functionalizable AuNCs capped with thioctic acid and thiol-modified polyethylene glycol in aqueous solution. The synthetic approach requires neither organic solvents or additional ligand exchange nor extensive knowledge of synthetic chemistry to reproduce. The resulting AuNCs offer free surface carboxylic acids, which can be functionalized with various biological molecules bearing a free amine group without adversely affecting the photoluminescent properties of the AuNCs. A quick, reliable procedure for flow cytometric quantification and confocal microscopic imaging of AuNC uptake by HeLa cells also been described. Due to the large Stokes shift, proper setting of filters in flow cytometry and confocal microscopy is necessary for efficient detection of near-infrared photoluminescence of AuNCs.

A reliable and easily reproducible method for preparation of functionalizable, near-infrared emitting photoluminescent gold nanoclusters and their direct detection inside HeLa cells by flow cytometry and confocal laser scanning microscopy is described.

Over the past decade, fluorescent gold nanoclusters (AuNCs) have witnessed growing popularity in biological applications and enormous efforts have been ...