not-yet-known not-yet-known not-yet-known unknown Synthesis and characterization of FVIOs The 3,4-Dihydroxyhydrocinnamic acid (DHCA)-coated FVIOs (DHCA-FVIOs) with an average diameter of 52 nm were first synthesized using a wet chemical method reported previously[48]. Moreover, 20 nm DHCA-SPIOs were prepared as controls in this study[49]. Scanning electron microscopy (SEM) revealed that the DHCA-FVIOs had a uniform ring shape with an average outer diameter, inner diameter and thickness of 52 nm, 18 nm and 15 nm, respectively (Figure S1, Supporting Information). For precisely targeted magnetothermal stimulation, DHCA-FVIOs and DHCA-SPIOs were further covalently bound to anti-His antibodies using a 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide/N-hydroxysuccinimide (EDC/NHS) coupling reaction to enable TRPV1 targeting via an extracellular 6× His epitope tag on the transfected cells. The scanning electron micrograph of FVIOs is presented in fig. S2a. The transmission electron microscopy (TEM) images of the negatively stained FVIOs and SPIOs modified with anti-His antibodies were displayed in Fig. 1 (a and b). The antibody coatings are observed as a marked layer (~ 5 nm thick) on the surface of the nanoparticles. The covalent binding of the anti-His antibody to the nanoparticles was confirmed using infrared spectroscopy (Figure S2b, Supporting Information). Moreover, the powder X-ray diffraction patterns (Figure 1c) confirmed the cubic inverse spinel phase of Fe3O4 in both FVIOs and SPIOs. As shown in fig. S3a, after bioconjugation, the hydrodynamic diameters of the anti-His-FVIOs and anti-His-SPIOs increased to 105.71 nm and 37.84 nm, respectively. In addition, both samples maintained their original hydrodynamic diameters after incubation in PBS for more than 80 days, suggesting their good colloidal stability (Figure S3b, Supporting Information). The magnetic properties of these two samples were also examined using a vibrating sample magnetometer. Compared with SPIOs, FVIOs exhibited a vortex magnetic structure with a much greater saturation magnetization and larger hysteresis loop area at room temperature (Figure 1d). Induction heating measurements were then performed with the FVIOs using an induction heating system (Supermag M5), in which the temperature changes in the samples were recorded by an optical fiber probe. And the applied AMF had a frequency (f) of 275 kHz and an amplitude (H) of 20 mT. Figure 1e presents the temperature profiles of the anti-His-FVIOs and anti-His-SPIOs aqueous suspensions with Fe concentrations of 0.05, 0.15, and 0.30 mg/mL. The calculated SAR of the FVIOs was approximately 3462 W/g, which is 10.78 times greater than that of the SPIOs (321 W/g) (Figure 1f). The superior heat-generating ability of the FVIOs can be attributed to the significantly increased hysteresis loss during the vortex-to-onion magnetization reversal process in response to AMF exposure[50]. Cytotoxicity evaluations were further performed with anti-His-FVIOs and anti-His-SPIOs with Fe concentrations ranging from 0 to 1000 μg/mL using HEK293T cells. After 24 h of incubation, the FVIOs and SPIOs did not alter the viability of HEK293T cells or cortical neurons at Fe concentrations ranging from 0 to 200 μg/mL (Figure S4a, b, Supporting Information ). When the Fe concentration increased from 200 to 1000 μg/mL , both the FVIOs and SPIOs exhibited severe cytotoxicity with significant decreases in cell viability (much lower than 80% viability). These results imply that the biocompatible anti-His-FVIOs with superior heat induction properties can be successfully prepared as efficient magnetothermal transducers for neurostimulation applications.