AIEgen-Conjugated Magnetic Nanoparticles as Magnetic–Fluorescent

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Cite This: ACS Appl. Nano Mater. 2019, 2, 3292−3299

AIEgen-Conjugated Magnetic Nanoparticles as Magnetic− Fluorescent Bioimaging Probes Kuheli Mandal,† Debabrata Jana,‡ Binay K. Ghorai,*,‡ and Nikhil R. Jana*,† †

Centre for Advanced Materials and School of Materials Science, Indian Association for the Cultivation of Science, Kolkata 700 032, India ‡ Department of Chemistry, Indian Institute of Engineering Science and Technology, Shibpur, Howrah 711 103, India

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S Supporting Information *

ABSTRACT: Colloidal nanoparticles with a magnetic−fluorescent property and varied surface chemistry have enormous biomedical application potential. However, the synthesis of a high quality of such a nanoparticle with 100 nm with limited surface chemistry for functionalization, restricting wider biomedical application potential.27−30 We designed and developed a nanoprobe for cellular/ subcellular targeting, imaging, and therapy.22,32 We used quantum dots,22 magnetic nanoparticles,22 plasmonic nanoparticles,22 and AIEgens33,34 for development of these nanoprobes. We have designed a nanoprobe for controlling the endocytotic uptake mechanism and for targeting different Received: April 6, 2019 Accepted: May 7, 2019 Published: May 7, 2019 3292

DOI: 10.1021/acsanm.9b00636 ACS Appl. Nano Mater. 2019, 2, 3292−3299

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ACS Applied Nano Materials

with MN-control I by the processes described above. Colloidal solution of the nanoparticle was used to measure the optical property (using a BioTek Synergy MX microplate reader) and hydrodynamic size and zeta potential (using Malvern, Nano ZS). A superconducting quantum interference device (SQUID) magnetometer was used for magnetic measurement study of the solid composite materials. Fourier transform infrared spectroscopy on KBr pellets was performed using a Shimadzu FT-IR 8400S instrument. TEM study was performed using an FEI Tecnai G2F20 microscope with a field-emission gun operating at 200 kV. FESEM was performed with a JEOL JSM-7610F microscope. Estimation of Primary Amine and TPE in Nanoparticle. The number of primary amines per nanoparticle was determined before TPE conjugation via a fluorescamine test. For this, borate buffer solution (pH 9) of polymer coated nanoparticles and acetone solution of fluorescamine (1 mg/mL) were prepared separately. Next, equal volumes (100 μL) of fluorescamine solution and nanoparticle solution were mixed, and then, fluorescence was measured at an excitation wavelength of 400 nm. For determination of the number of TPEs per particle, a standard calibration curve was prepared from the absorbance value of the TPE molecule at 354 nm at different concentrations (Supporting Information, Figure S1). Next, the concentration of TPE present in the nanoparticle was determined by measuring the absorbance value at 354 nm and fitting it with the above-mentioned calibration graph. The concentration of γ-Fe2O3 nanoparticles was determined by a reported method.32 Cell Labeling Experiment. Human cervical cancer cell line (HeLa) was cultured as described earlier.34 For cell labeling study, cells were seeded in 24 well plates with 500 μL of culture media. After overnight growth, cells were treated with 100−200 μL of sample solution with a final concentration of 0.16 mg/mL. After 1 h of incubation, washed cells were imaged using an Olympus IX81 microscope attached with a digital camera. For colocalization study, nanoparticle labeled cells were further incubated with fresh media for 24 h and then treated with LysoTracker Red for 30 min. Then, washed cells were used for imaging study. For cytotoxicity study, cells were incubated with different doses of sample for 24 h. Then, cells were washed with PBS to remove unbound particles, and a fresh 500 μL of culture media was added. Next, each well plate with attached cells was treated with 50 μL of freshly prepared MTT solution (5 mg/mL) for 4 h. After that, supernatant was removed carefully, leaving formazan in plate. Next, formazan was dissolved in freshly prepared sodium dodecyl sulfate (SDS) solution (8 g of SDS dissolved in 40 mL of 50% water−DMF solution), and absorbance was measured at 570 nm. Cell viability was estimated assuming 100% viability for control without any sample treatment.

subcellular compartments such as the mitochondria, lipid droplet, and perinuclear regions.22 We also developed a multifunctional nanoprobe via a hybrid-nanoparticle approach9 and fluorophore conjugation to a nanoparticle.13 Here we report an AIEgen-based magnetic−fluorescent nanoparticle with 25−50 nm hydrodynamic size and variable surface chemistry. The approach involves “switching on” of AIEgen fluorescence via its covalent linking to a polymer coated colloidal magnetic nanoparticle surface. The resultant nanoparticle becomes magnetic−fluorescent with three distinct advantages over reported AIEgen-based multifunctional nanoparticles.27−31 First, the hydrodynamic size is small (average size < 50 nm) and suitable for cellular and subcellular targeting application. Second, nanoparticle surface charge can be varied to be positive, negative, or zwitterionic, and this can be used to control the cellular interaction. Third, the nanoparticle surface is terminated with primary amines that can be used for conjugation chemistry and preparation of various functional nanoparticles and nanobioconjugates.

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EXPERIMENTAL SECTION

Materials. Poly(ethylene glycol) methacrylate, 3-sulfopropyl methacrylate, N,N-methylene-bis-acrylamide, N-(3-aminopropyl)methacrylamide hydrochloride, N,N-methylene-bis-acrylamide (MBA), ammonium persulfate, 4-methyl morpholine-N-oxide (MNO), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), bis[2-(methacryloyloxy)ethyl]phosphate, and octadecene were purchased from Sigma-Aldrich and used as received. Details of the synthesis of tetraphenylethene (TPE) are described elsewhere.33 Preparation of Polyacrylate Coated Magnetic Nanoparticle (MN-Control I/II/III). The hydrophobic γ-Fe2O3 nanoparticle was synthesized via a high temperature colloidal method.32 In brief, a mixture of 0.37 g of iron stearate, 0.16 g of octadecylamine, 0.16 g of MNO, and 8 mL of octadecene was heated at 60 °C under inert atmosphere for 15 min. After that, the temperature of the reaction mixture was increased to 300 °C, and heating was continued at this temperature for 10 min. The nanoparticle was then purified from excess surfactants via acetone/ethanol-based precipitation and chloroform/cyclohexane-based redispersion. The hydrophobic γFe2O3 nanoparticle was then converted into a hydrophilic nanoparticle by using polyacrylate coating as described earlier.13,32 The molar ratios of the acrylate monomers were appropriately adjusted as described previously.13,32 In brief, 30 mg of N-(3-aminopropyl)methacrylamide hydrochloride was used for preparation of MNcontrol I, 14 mg of N-(3-aminopropyl)-methacrylamide hydrochloride and 12 mg of 3-sulfopropyl methacrylate were used for preparation of MN-control II, and 9 mg of N-(3-aminopropyl)methacrylamide hydrochloride and 12 mg of 3-sulfopropyl methacrylate were used for preparation of MN-control III. In all cases, 6 μL of bis[2-(methacryloyloxy)ethyl]phosphate was used as a cross-linker in addition to other acryl monomers. The resultant polyacrylate coated γ-Fe2O3 nanoparticle was then washed with chloroform and ethanol repeatedly and dispersed in 3−5 mL of distilled water. Finally, polyacrylate coated γ-Fe2O3 nanoparticle solution was dialyzed using nitrocellulose membrane (MWCO 12 000 Da) to remove unreacted reagents. Preparation of TPE-Conjugated Magnetic Nanoparticle (MN-TPE-I/II/III). Aqueous solution (500 μL) of MN-control I/II/ III was diluted to 1 mL by adding methanol. In another vial, 2.0 mg of TPE was dissolved in 1 mL of methanol, mixed with nanoparticle solution, and stirred for 15 min. After that, 100 μL of aqueous solution of NaCNBH3 (10 mg/mL) was added and stirred for overnight. Next, the resultant TPE-conjugated nanoparticle (MNTPE-I/II/III) was purified by dialysis followed by chloroform extraction to remove unbound TPE. In order to vary the number of TPEs per nanoparticle, three different amounts of TPE (0.5, 1, 2.0 mg) were dissolved in 1 mL of methanol and used for conjugation

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RESULTS Conversion of Magnetic Nanoparticle to Magnetic− Fluorescent Colloid via AIEgen Conjugation. The synthetic approach of AIEgen conjugation to a magnetic nanoparticle is shown in Scheme 1. First, a surfactant capped hydrophobic γ-Fe2O3 nanoparticle of 6−10 nm size is synthesized via a high temperature colloid−chemical approach, and then, it is converted into a primary amine terminated hydrophilic nanoparticle via polyacrylate coating.13,32 The hydrophobic γ-Fe2O3 nanoparticle is insoluble in aqueous media because of the presence of hydrophobic surfactants on their surface. So, we have transformed it to a polyacrylate coated nanoparticle where the hydrophilic shell provides colloidal stability. Next, primary amines are used for covalent conjugation with an AIEgen molecule.33,34 We have used tetraphenylethene (TPE) as the AIEgen. The TPE molecule has one aldehyde group in its structure so that it can react with a primary amine and form a covalent bond. We have used three different types of polyacrylate shells and depending on the shell 3293

DOI: 10.1021/acsanm.9b00636 ACS Appl. Nano Mater. 2019, 2, 3292−3299

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ACS Applied Nano Materials Scheme 1. Synthetic Strategy for AIEgen-Conjugated Magnetic−Fluorescent Nanoparticlea

It involves transformation of a 6−10 nm hydrophobic γ-Fe2O3 nanoparticle to a primary amine terminated hydrophilic nanoparticle of 25−50 nm hydrodynamic size via polyacrylate coating followed by conjugation with AIEgen (TPE). Depending on the acrylate monomer used, nanoparticles are cationic (MN-control I/MN-TPEI), zwitterionic (MN-control II/MN-TPE-II), or anionic (MNcontrol III/MN-TPE-III) in nature. a

Figure 1. (a) Aggregation-induced emission property of TPE as observed by precipitation of the same concentration of TPE with increasing water volume % in the water−ethanol mixture. (b) Evidence of “switching on” of TPE fluorescence after binding with a colloidal magnetic nanoparticle. A varied amount of TPE is conjugated with MN-TPE-I, and the average number of TPEs per particle is 60 (1), 120 (2), and 230 (3). TPE concentration is kept the same (by adjusting the same absorbance of TPE at 325 nm for three samples), in order to show that the “switching on” property appears with an increased number of TPEs per particle.

type, the TPE-conjugated nanoparticles are named as MNTPE-I, MN-TPE-II, and MN-TPE-III. The MN-TPE-I is derived from MN-control I with the shell having primary amine (NH2) and polyethylene glycol. Similarly, MN-TPE-II and MN-TPE-III are derived from MN-control II and MNcontrol III, respectively, both of which have NH2, PEG, and sulfopropyl (SO3−). The only difference between MN-control II and MN-control III (and hence MN-TPE-II/MN-TPE-III) is the different % of functional groups. The presence of primary amines on the surface of MN-control I/II/III has been estimated via a fluorescamine test. The tentative numbers of primary amines per particles are 600, 450, and 300 for MNcontrol I, MN-control II, and MN-control III, respectively. In addition, the tentative number of TPEs present per MN-TPEI/II/III has been determined from the absorbance of TPE at 354 nm. The values are 230, 230, and 120 for MN-TPE-I, MNTPE-II, and MN-TPE-III, respectively. These values indicate that a fraction of primary amines are reacted, and the remaining numbers of primary amines are 370, 220, and 180 for MN-TPE-I, MN-TPE-II, and MN-TPE-III, respectively. Physical properties of all of the nanoparticles are summarized in Table 1 as well as Figures 1−4, S2, S6, and S7. Figure 1a shows the AIE property of TPE. As the water is

added to the ethanol solution of TPE, molecularly soluble TPE starts aggregating, which results in the “switch on” of green fluorescence. A similar type of fluorescence “switch on” property is also observed after conjugation of TPE with the polymer coated magnetic nanoparticle (Figures 1b and S2). In particular, it is shown that as the number of attached TPEs per particle increases from 60 to 230, the “switching on” property of TPE is enhanced. In contrast, molecularly soluble TPE solution of a similar concentration is completely nonfluorescent (see Figure 1a). This result clearly suggests that a dominant TPE−TPE interaction occurs at the nanoparticle surface that leads to the fluorescence “switch on” property. As the fluorescence “switching on” property of TPE increases with the increased number of TPEs per nanoparticle, we have tried to conjugate the maximum number of TPEs per particle to encourage the maximum TPE−TPE interaction.

Table 1. Properties of AIEgen-Derived Magnetic−Fluorescent Nanoparticles nanoparticle type

functional groups (number per particle)a

average hydrodynamic size, fluorescence quantum yield (%)

surface charge at pH 4.5, 7.4, 9

cell labeling performance, cytotoxicity

MN-TPE-I MN-control I MN-TPE-II MN-control II MN-TPE-III MN-control III

−NH2 (370), TPE (230) −NH2 (600) −NH2 (220), −SO3−, TPE (230) −NH2 (450), −SO3− −NH2 (180), −SO3−, TPE (120) −NH2 (300), −SO3−

25 ± 10 nm, 6−8% 25 ± 10 nm, -----35 ± 10 nm, 6−8% 25 ± 10 nm, -----50 ± 20 nm, 3−4% 50 ± 20 nm, ------

+23, +4, −6 +18, +4, −4 +9, −1, −13 +8, 0, 0 −2, −13, −15 −1, −3, −10

good, low good, high good, low good, low poor, low poor, low

a

Each nanoparticle has PEG functional groups in addition to the mentioned functional groups. 3294

DOI: 10.1021/acsanm.9b00636 ACS Appl. Nano Mater. 2019, 2, 3292−3299

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Figure 2. (a) Absorption/emission spectra and dynamic-light-scattering-based hydrodynamic size of MN-TPE-I (a), MN-TPE-II (b), and MNTPE-III (c).

are 25−50 nm in average hydrodynamic size, including inorganic core and polymer shell along with all chemical functional groups (Figure 2). The surface charge of the nanoparticles is determined at different solution pHs by zeta potential measurement (Table 1). It is observed that MNTPE-I shows a positive surface charge at pH 4.5 because of protonated NH2(NH3+), and as the pH increases to 7−9, the charge approaches zero due to deprotonation of NH2 groups. We called this nanoparticle cationic, as it shows positive charge in the pH range of 4.5−7.4. In contrast, MN-TPE-II shows a relatively lower positive surface charge at pH 4.5, as NH3+ ions are partially balanced by anionic sulfopropyl (SO3−) groups. As the pH increases to 7.4, the charge approaches zero, but at pH 9.0, the surface charge becomes anionic. We called this nanoparticle zwitterionic, as it changes the surface charge from positive to negative, as the pH changes from 4.5 to 9.0. In contrast, the surface charge of MN-TPE-III shows a negative value in all pH because of dominant SO3− groups, and we termed it as an anionic nanoparticle. Magnetic property measurement system (MPMS) has been used to study the magnetic property of MN-TPE-I and MNcontrol I (Figure 3). The result shows characteristic hysteresis, similar to superparamagnetic materials, which almost disappears at room temperature. The saturation magnetization values for MN-control I are 3.8 and 2.9 emu g−1 at 10 and 300 K, respectively. Similarly, for MN-TPE-I, the saturation

Figure 2 shows UV−visible absorption spectra of MN-TPEI, MN-TPE-II, and MN-TPE-III with the characteristic absorption band of TPE at 325 nm and an emission band between 450 and 550 nm. However, the absorption band and emission band maxima of TPE in MN-TPE-I/II/III blue shifts from 354 to 325 nm and from 530 to 500 nm, respectively, as compared to ethanol-based TPE aggregates. This is due to the transformation of the −HCO group of TPE to an −H2C− NH− group that minimizes π-conjugation. The fluorescence quantum yield has been measured using quinine sulfate as the reference, and the values are in the range of 6−8% for MNTPE-I/MN-TPE-II and 3−4% for MN-TPE-III (Table 1). The lower quantum yield value for MN-TPE-III is due to a lower number of TPEs per particle with lower TPE−TPE interaction. Polyacrylate coating on the nanoparticle surface has been verified by Fourier transform infrared (FTIR) spectra. FTIR spectroscopy of MN-control I and MN-TPE-I shows the appearance of hydroxyl group signals around 3000−3500 cm−1 because of the polyacrylate component. Figure S3 TEM image shows the inorganic γ-Fe2O3 core of 6−10 nm, and the polymer shell is observed as a light gray color around a dark core after the uranyl acetate staining (Figure S4). The SEM image shows an overall size of 15−22 nm that includes polyacrylate coating and the core nanoparticle (Figure S5). Hydrodynamic sizes of nanoparticles are measured before and after TPE conjugation. The results show that all nanoparticles 3295

DOI: 10.1021/acsanm.9b00636 ACS Appl. Nano Mater. 2019, 2, 3292−3299

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ACS Applied Nano Materials

Figure 3. Field dependent magnetization curves of MN-control I (a) and MN-TPE-I (b) at 10 and 300 K and the respective temperature dependent zero-field-cooled (ZFC) and field-cooled (FC) magnetization curves.

magnetization values are 3.4 and 2.5 emu g−1 at 10 and 300 K, respectively. The zero-field cooling (ZFC) and field cooling (FC) magnetization curves of MN-control I and MN-TPE-I show well-defined blocking temperatures of 20 and 10 K, respectively, which also indicate the superparamagnetic nature of the nanoparticles. Colloidal stability of MN-TPE-I, MN-TPE-II and MN-TPEIII has been investigated in buffer solution of varied pH. Digital images of all colloidal dispersions are shown in Figure 4. The result shows that nanoparticles have good colloidal stability in all buffer solutions for more than 1 month. However, colloidal nanoparticles can be precipitated by adding a large amount of salt, and precipitated particles can then be attracted by laboratory-based bar magnets (Figure 4). Magnetically separated particles can be redispersed in fresh aqueous media or by removing of salt via dialysis. Fluorescence stability study of MN-TPE-I, MN-TPE-II, and MN-TPE-III has been carried out in the presence of buffer solution of different pH (Figure S6). Results show that the fluorescence intensity of MN-TPE-I remains intact at pH 7.4 and 9.0 but decreases to some extent at pH 4.5. This may be due to protonation of the primary/secondary amine group that reduces restricted intramolecular motion (RIM) to a certain extent. Fluorescence stability of MN-TPE-I, MN-TPE-II, and MNTPE-III is further investigated under continuous UV exposure of colloidal solution of nanoparticles and drop-casted film of nanoparticle, which show that fluorescence remains intact under continuous light exposure of 30 min (Figure S7). These results suggest that fluorescence of nanoparticles would be stable under continuous excitation and useful for long-term imaging application. AIEgen-Conjugated Magnetic Nanoparticle as Magnetic−Fluorescent Cell Imaging Probe. In order to explore the bioimaging application potential, MN-TPE-I,

Figure 4. Colloidal solutions of MN-TPE-I, MN-TPE-II, and MNTPE-III in daylight or under handheld UV light (UV) in different buffer solutions. Right panel shows the magnetic separation of nanoparticles by using a laboratory-based bar magnet after their precipitation.

MN-TPE-II, and MN-TPE-III have been used to label cells. Typically, HeLa cells are incubated with nanoparticles for 1 h, and washed cells are incubated with fresh media for 24 h. Next, washed cells are imaged under fluorescence mode with UV excitation. Results are summarized in Figures 5 and S8. It is observed that MN-TPE-I and MN-TPE-II can label cells within 1 h, and their fluorescence intensity within cells remains intact even up to 24 h, and they localize in cell cytoplasm (Figure 5). In contrast, labeling efficiency of MN-TPE-III is 3296

DOI: 10.1021/acsanm.9b00636 ACS Appl. Nano Mater. 2019, 2, 3292−3299

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ACS Applied Nano Materials

with a cytotoxic effect,22 we have intentionally designed MNTPE-I with a lower cationic charge and PEGylated functional groups. This allows MN-TPE-I with high cell uptake with lower cytotoxicity.

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DISCUSSION An AIE active hydrophobic TPE molecule is nonemissive in organic solvents such as methanol/ethanol, in which it remains in molecularly soluble form. In aqueous environment, it started to aggregate, where intramolecular rotation of phenyl rotors becomes restricted and “switches on” its fluorescence with increasing water volume % in a water−ethanol mixture. Here we have used the strategy of covalent conjugation of TPE to a polymer coated magnetic nanoparticle surface to “switch on” its fluorescence. Transformation of a nonfluorescent magnetic nanoparticle into a fluorescent magnetic nanoparticle by this type of covalent conjugation with a conventional organic fluorophore has significant limitations due to the fluorescence quenching issue via intermolecular energy transfer. In contrast, the AIE active TPE molecule does not have such a quenching issue.13,23 The presented approach involves covalent conjugation of TPE to the primary amine terminated magnetic nanoparticle. Considering the fact that nanoparticles are 25−50 nm in hydrodynamic size and each nanoparticle has 120−230 TPEs, it can be expected that TPEs are close to each other at the nanoparticle surface, offering TPE−TPE interaction that leads to restriction in intramolecular motion (RIM) and the fluorescence “switch on” phenomenon. However, this TPE− TPE interaction is too weak to offer any red/blue shifting of their UV−vis absorption spectrum. The presented synthetic approach has three additional advantages coming from polyacrylate coating. First, a nanoparticle can be prepared with good colloidal stability as compared to other coating approaches. For example, we have demonstrated good colloidal and fluorescence stability of three designed nanoparticles under physiological conditions. Second, a nanoparticle with a wide variation of surface chemistry can be generated using different acryl monomers and by varying their molar ratio. For example, we have prepared three different nanoparticles with cationic, anionic, and zwitterionic surface charges. Third, nanoparticles can be further functionalized using the remaining primary amines on their surface. In particular, the poor cell labeling property of MN-TPE-III is ideal to minimize nonspecific labeling of nanoparticles and can be used for selective labeling after conjugation with bioaffinity molecules.

Figure 5. Application of MN-TPE-I (a), MN-TPE-II (b), and MNTPE-III (c) as a fluorescent cell imaging probe. Inset shows magnetic separation of MN-TPE-I labeled cells. HeLa cells are incubated with sample for 1 h, washed cells are incubated with fresh culture media up to 24 h and imaged under bright field or fluorescence mode, and merged images are shown here. Results show that MN-TPE-I and MN-TPE-II can label cells very efficiently as compared to MN-TPEIII and stay/localize inside cytoplasm in the next 24 h. For magnetic separation, cells are detached from the culture plate (via trypsin− EDTA), and then, water dispersed cells are observed under handheld UV light in the absence/presence of magnet.

relatively poor, and its fluorescence intensity inside cells decreases with longer incubation time. It is observed that the localization of MN-TPE-I/MN-TPE-II inside a cell takes 8−24 h, depending on the nature of the surface charge. In order to investigate the multifunctional application potential of these nanoparticles, MN-TPE-I labeled cells are dispersed in cell culture media and kept under a laboratory-based bar magnet. It can be found that labeled cells are easily separated by magnet (Figure 5a). In order to understand the nature of localization, a colocalization study with LysoTracker Red has been performed. It has been found that particles significantly colocalize with LysoTracker Red (Figure S8). This result suggests that nanoparticles enter into the cell via endocytosis processes that traffic them to the endosome/lysozome, and a different extent of labeling is linked to the surface property of the nanoparticle. MN-TPE-I with a cationic surface charge has a strong interaction with the anionic cell membrane, but MNTPE-II with zwitterionic surface charge has modular interaction with the cell membrane, and MN-TPE-III with anionic charge can only weakly interact with the membrane via lipophilic TPE groups. Thus, MN-TPE-III has a weaker labeling property than the other two nanoparticles. Cell viability study of MN-TPE-I/II/III nanoparticles has been performed using HeLa cells via MTT assay. The result shows low cytotoxicity of all three nanoparticles at a wider concentration range (Figure S9). Although cationic nanoparticles are known to have a good labeling property along

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CONCLUSION We have shown that a magnetic−fluorescent colloidal nanoparticle with an average hydrodynamic size of 25−50 nm can be prepared via covalent linking of an AIEgen to a polymer coated magnetic nanoparticle surface. This processes involves the “switching on” of AIEgen fluorescence, which is otherwise nonfluorescent in a molecularly soluble state. The resultant colloidal nanoparticle has a cationic/anionic/ zwitterionic surface charge with primary amine termination. It is demonstrated that the nanoparticle can be used as an cell imaging probe, and labeled cells can be magnetically separated. The primary amines present at the nanoparticle surface can be used for the development of nanoparticles and nanobioconjugates with various functionalities. 3297

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ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acsanm.9b00636.

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Details of quantification of primary amines in the nanoparticle surface, TEM image of polymer coated magnetic nanoparticle, colocalization study of nanoparticle with LysoTracker and cytotoxicity data (PDF)

AUTHOR INFORMATION

Corresponding Authors

*E-mail: [email protected]. (N.R.J.) *E-mail: [email protected]. (B.K.G.) ORCID

Kuheli Mandal: 0000-0001-9705-5419 Nikhil R. Jana: 0000-0002-4595-6917 Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS N.R.J. acknowledges the DST Nano Mission (Grant number SR/NM/NB/1009/2016) and CSIR (Grant number 02(0249) 15/EMR-II), Government of India, for financial assistance. K.M. acknowledges CSIR, India, for providing a research fellowship.

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DOI: 10.1021/acsanm.9b00636 ACS Appl. Nano Mater. 2019, 2, 3292−3299