Self-regulated co-assembly of soft and hard nanoparticles

Controlled self-assembly of colloidal particles into predetermined organization facilitates the bottom-up manufacture of artificial materials with designated hierarchies and synergistically integrated functionalities. However, it remains a major challenge to assemble individual nanoparticles with minimal building instructions in a programmable fashion due to the lack of directional interactions. Here, we develop a general paradigm for controlled co-assembly of soft block copolymer micelles and simple unvarnished hard nanoparticles through variable noncovalent interactions, including hydrogen bonding and coordination interactions. Upon association, the hairy micelle corona binds with the hard nanoparticles with a specific valence depending exactly on their relative size and feeding ratio. This permits the integration of block copolymer micelles with a diverse array of hard nanoparticles with tunable chemistry into multidimensional colloidal molecules and polymers. Secondary co-assembly of the resulting colloidal molecules further leads to the formation of more complex hierarchical colloidal superstructures. Notably, such colloidal assembly is processible on surface either through initiating the alternating co-assembly from a micelle immobilized on a substrate or directly grafting a colloidal oligomer onto the micellar anchor.

). The preformed BA2 trimers retained intact and the further added S2090V380 micelles failed to associate with them, indicating that the surface of the central silica NP is saturated due to the coverage of the P2VP coronas. Silica-NPs-capped AB3 tetramers assembled by S2090V380 micelles and 50-nm silica NPs (corresponding to Supplementary Fig. 5). (b) Silica-NPs-capped AB2 trimers assembled by S2090V380 micelles and 50-nm silica NPs (corresponding to Supplementary Fig. 6). (c) AB dimers assembled by S2000V1910 micelles and 50-nm silica NPs (corresponding to Supplementary Fig. 28). (d) Micelles-capped BA2 trimers assembled by S2090V380 micelles and 50-nm silica NPs (corresponding to Supplementary Fig. 9). (e) Micelles-capped BA3 tetramers assembled by S2090V380 micelles and 75-nm silica NPs (corresponding to Supplementary Fig. 16). (f) Micelles-capped BA4 pentamers assembled by S2090V380 micelles and 90-nm silica NPs (corresponding to Supplementary Fig. 22). Additional insights to the formation of silica-NP-centered and SV-micelle-centered colloidal molecules. The structure (valence) of the colloidal molecules is generally regulated by the relative size and feeding ratio of SV micelles (denoted as A) and hard nanoparticles (NPs, denoted as B) (Fig. 2). Taking silica-NP-centered colloidal molecules as an example, the saturated valence (Vmax) of a silica NP is determined by the surface area of a silica NP (Ssilica) and the area that a SV micelle can cover on the silica NP (Smicelle) ( Supplementary  Fig. 26a). For example, the saturated valence of a 50-nm silica NP is 2 when S2090V380 micelles are employed as evidenced by the co-existence of BA2 molecules and excessive S2090V380 micelles ( Supplementary Fig. 10). However, when S2000V1910 micelles are used the saturated valence of a 50-nm silica NP turns to be 1 since longer P2VP coronas can cover a larger surface area (Supplementary Figs. S29 27 and 28). Similarly, the increase in silica NP size (75-nm/90-nm silica NPs, with larger surface area) can increase its saturated valence (Supplementary Figs. 16 and 22). Ideally, the saturated (maximum) valence of a silica NP Vmax can be calculated by

Supplementary
The actual valence of a silica NP (Vactual, the number of surrounding, actually associated SV micelles) is an integrative consequence of the saturated valence Vmax and the feeding ratio of micelles to silica NPs Nmicelle/Nsilica. As a result, we can conclude that where Nmicelle and Nsilica are the numbers of micelles and silica NPs added in the co-assembly system, respectively; Vdynamic is the dynamic valence of a silica NP in the assembled dynamic structures (such as 1D colloidal chains and 2D/3D superstructures). When the feeding ratio Nmicelle/Nsilica is greater than or equal to the saturated valence Vmax, the number of micelles in the system can meet the silica NP demand of achieving a saturated valence and micelles-capped silica-NP-centered colloidal molecules will be generated. However, when Nmicelle/Nsilica is less than Vmax, the number of micelles is insufficient and various dynamic structures will be formed. For example, for S2090V380 micelles and 50-nm silica NPs at A:B ≈ 1:1, micelles and silica NPs can connect with each other to form 1D colloidal chains ( Supplementary Fig. 8) in which the valence of a silica NP remains to be 2 in order to achieve H-bonding maximization in spite of the insufficient number of micelles. Similar situation occurs for 2D/3D superstructures assembled by 75-nm/90-nm silica NPs (Supplementary Figs. 14 and 20). For silica-NP-centered clusters, the surface of the silica NP was saturatedly covered by the P2VP coronas and the repulsion from the solvent-swollen P2VP coronas may cause the distance between the surrounding micelles. For SV-micelle-centered clusters, similarly, the electrostatic repulsion between the silica surface may render a specific distance between the surrounding NPs. (AB3)(BA2)3 colloidal conjugates were assembled by AB3 colloidal molecules (assembled by S2090V380 micelles and 50-nm silica NPs) and BA2 colloidal molecules (assembled by S2090V380 micelles and 50-nm silica NPs).  Supplementary Fig. 37). (b) CA2 trimers assembled by S1520V370 micelles and 47-nm Au NPs (corresponding to Supplementary  Fig. 38). (c) CA3 tetramers assembled by S460V190 micelles and 47-nm Au NPs (corresponding to Supplementary Fig. 39). (d) DA2 trimers assembled by S2090V380 micelles and 53-nm ZIF-8 NPs (corresponding to Supplementary Fig. 42). (e) DA4 pentamers assembled by S1220V440 micelles and 70-nm ZIF-8 NPs (corresponding to Supplementary Fig. 43). (f) DA6 heptamers assembled by S1030V150 micelles and 70-nm ZIF-8 NPs (corresponding to Supplementary Fig. 44).  Fig. 49) into an ethanol solution of silica-NPs-capped short or long colloidal oligomers (formed by S2090V380 micelles and 67-nm silica NPs, Supplementary Fig. 51), respectively. The white arrows denote the micellar anchors preformatively immobilized on the silicon wafer surface.

Preparation of silica NPs
Silica NPs with various diameters were synthesized by a modified Stöber method 1 . In a typical experiment, 1 mL of aqueous ammonia (28%) and 20 mL of ethanol were mixed thoroughly for 30 min, followed by the rapid injection of 0.06 mL of tetraethyl orthosilicate (TEOS) under vigorous stirring. After stirred for 30 min, the solution was allowed to age for 24 h at room temperature. Subsequently, the resulting solution was dialyzed against ethanol to remove the remaining ammonia before analysis.

Preparation of Au NPs
Au NPs with various diameters were synthesized through a seed-mediated growth method assisted with mild oxidation as previously reported 2 . To prepare the seed solution, 0.6 mL of an ice-cold, freshly prepared aqueous solution of NaBH4 (0.01 M) was rapidly injected into a mixture containing 0.

Preparation of core/shell structured silica-coated Au NPs
Silica-coating of Au NPs was carried out as previously reported with slight modifications 3 . In a typical experiment, 20 µL of an aqueous solution of NaOH (0.1 M) was added to 4 mL of the as-prepared 20-fold concentrated Au NPs solution, followed by the addition of 150 µL of a methanol solution of TEOS (20%, v/v). The resultant mixture was gently stirred for 1 h and left undisturbed for 24 h at room temperature. Subsequently, the mixture was washed with methanol by multiple centrifugation-redispersion cycles. The obtained silica-coated Au NPs were collected and redispersed in ethanol.

Preparation of PDA NPs
PDA NPs with various diameters were synthesized according to a previous literature 4 . In a typical experiment, 6 mL of ethanol, 14 mL of deionized water, and 0.3 mL of aqueous ammonia (28%) were mixed thoroughly for 30 min, followed by the rapid injection of 1.5 mL of an aqueous solution of dopamine hydrochloride (50 mg/mL) under vigorous stirring. After stirring for 24 h at room temperature, the resulting PDA NPs were collected by centrifugation and redispersion in ethanol.

Preparation of ZIF-8 NPs
ZIF-8 NPs were prepared as previously reported with slight modifications 5 . In a typical experiment, 5 mL of an aqueous solution of Zn(CH3COO)2·2H2O (60 mg/mL) was added to 5 mL of an aqueous solution containing 2-methylimidazole (2.72 M) and CTAB (0.54 mM) with gentle stirring for a few seconds. After 15 s, the mixture was left undisturbed at room temperature for 2 h. Subsequently, the resulting ZIF-8 NPs were washed with deionized water and finally redispersed in deionized water. ) diblock copolymers were synthesized through sequential anionic polymerization in tetrahydrofuran (THF) in an inert argon atmosphere glovebox as reported in our previous work 6 . In a typical procedure, a solution of n-butyllithium (n-BuLi) in hexane (0.28 mL, 2.5 M) was added to 30 mL of dried THF under stirring and the mixture was allowed to age at room temperature overnight to remove the reactive impurities. Subsequently, 1.4 mL of styrene (12.18 mmol) was added to above solution at -78 °C, followed by the rapid injection of a solution of sec-butyllithium (sec-BuLi) in hexane (3.8 μL, 1.3 M) under vigorous stirring to initiate the polymerization of styrene. After 6 h, an aliquot (1/10) of the mixture was removed out and terminated by p-tert-butylphenol (PTBP). Subsequently, an excess amount (4.7 μL, 26.68 μmol) of diphenylethylene (DPE) was added into the mixture. A red color gradually developed within a few minutes. Then the solution was cooled to -78 °C and a solution of 2-vinylpyridine (0.14 mL, 1.3 mmol) and LiCl (1.88 mg, 44.46 μmol) in 1 mL of dried THF was added. After 1.5 h, PTBP was added to the resultant solution to terminate the living chain ends. Finally, the PS-b-P2VP diblock copolymer was isolated by precipitation into hexane and further into deionized water. The molecular weight of diblock copolymer was determined by combining the number-average molecular weight (Mn) of the first polystyrene block from GPC measurements with the block ratio obtained by integration of the corresponding 1 H NMR spectrum.

Preparation of PS-b-P2VP micelles
In a typical experiment, 100 mg of PS2090-b-P2VP380 (S2090V380) diblock copolymer solid was firstly dissolved in 10 mL of THF to yield a 10 mg/mL stock solution. Subsequently, to 0.9 mL of the above solution was added dropwise 33 mL of methanol via a syringe pump with an injection rate of 1.5 mL/h under mild stirring. The resulting solution was dialyzed against ethanol to remove THF, followed by filtration with a syringe filter (Titan, polytetrafluoroethylene membrane with a 0.22 μm pore size) to remove impurities. Finally, the solution was diluted to 30 mL by the addition of ethanol before use.

Characterization
Transmission electron microscopy (TEM) micrographs were obtained on a Thermo Scientific Talos L120C G2 microscope operated at 120 kV. High-resolution TEM micrographs, bright-field scanning transmission electron microscopy (BF-STEM) micrographs, high-angle annular dark-field STEM S58 (HAADF-STEM) micrographs and corresponding energy dispersive X-ray spectroscopy (EDS) elemental mapping micrographs were obtained on a Thermo Scientific Talos F200X microscope operated at 200 kV. Samples were prepared by placing one drop (ca. 10 µL) of solution on a carbon film-coated copper grid, which was placed on a piece of filter paper to remove excess solvent. No staining of the samples was necessary. Images were analyzed using the ImageJ software package (version 1.52a) developed at the US National Institute of Health. Scanning electron microscopy (SEM) micrographs were obtained on a TESCAN MAIA3 microscope operated at 5.0 kV. Samples were prepared by placing one drop (ca. 10 µL) of the solution on a clean silicon wafer. Before use, the silicon wafers were sonicated in acetone and deionized water for 30 min, respectively, and dried under a gentle stream of nitrogen. Then, the silicon wafers were subjected to oxygen-plasma treatment for 10 min. Before SEM characterization, the samples were sputter-coated with a thin layer of platinum for 15 s. Atomic force microscopy (AFM) height micrographs were obtained at ambient conditions using a Bruker Bio-FastScan atomic force microscope. Samples were prepared by placing one drop (ca. 10 µL) of the solution on a clean silicon wafer. Images were analyzed by NanoScope Analysis (version 1.80, an open source software program developed for AFM images). Gel permeation chromatography (GPC) measurements were performed on a Tosoh HLC-8320GPC gel permeation chromatograph. THF or DMF was used as the eluent at a flow rate of 1.0 mL/min. Sample was dissolved in the eluent overnight and filtered with a filter (Titan, polytetrafluoroethylene membrane with a 0.22 μm pore size) before analysis. Calibration was conducted using polystyrene standards. 1 H nuclear magnetic resonance (NMR) spectrum was recorded with a Bruker AVANCE III HD 500 NMR spectrometer. The samples were prepared by dissolving the polymers in CDCl3. Static water contact angle (CA) measurements were conducted on a KRÜSS DSA100 Drop Shape Analyzer equipped with a high-speed camera at room temperature. 2 μL of water was dropped via the sessile drop method and the angle was measured by the circle fitting method. For each sample, at least five locations were tested in order to obtain the average CA value.