Jem 2010hr

The morphologies of BPNS, BATNS were characterized by transmission electron microscope (TEM, JEM-2010HR, JEOL, Japan) operating at 120 kV. Atomic force microscopy (AFM) was performed on Bruker Diension Icon microscope. The size and zeta potential of BPNS, BATNS were measured by Malvern Zetasizer Nano ZS90. The structures of BPNS, BATNS were determined by Raman scattering, X-ray photoelectron spectroscopy (XPS, Axis HSi, Kratos Ltd., UK), UV–vis–NIR spectrophotometry (Lambda 950, PerkinElmer, UK), X-ray diffraction (XRD), Inductively coupled plasma atomic emission spectrometer (ICP-AES) and Micro infrared spectrophotometry. The surface morphology and elemental distributions were analyzed by scanning electron microscope (SEM) and energy-dispersive X-ray spectroscopy (EDS).

The preparation of the PAMAM-Es plasmid mixture was performed according to the Dendritech Company guidelines. Six PAMAM dendrimers were dissolved in water at a concentration of 1 mg/mL and stored at 4°C. The human recombined endostatin plasmid was stored at −20°C. When used, the PAMAM solution and the endostatin plasmid were both rewarmed in room temperature on the bench for 20 min and mixed together at a ratio of 3.25 μg : 1 μg (PAMAM : endostation). After 20 min of coincubation, the mixture was identified by JEM-2010HR transmission electron microscopy (JEOL, Tokyo, Japan).
As a traditional gene vector, Lipofectamine 2000 was taken to compare with PAMAM in both in vitro and in vivo studies. The preparation of the Lipofectamine-endostatin plasmid mixture (Lipo-Es) was in accordance with the routine of our laboratory. Briefly, Lipofectamine 2000 and endostatin plasmid were rewarmed in room temperature on the bench top for 20 min and mixed together at a ratio of 3.25 μg : 1 μg and cultured for another 20 min before injection.

The TEM images were taken by using a JEM-2010HR transmission electron microscope (JEOL, Tokyo, Japan) with a tungsten filament at an accelerating voltage of 200 kV. High-resolution transmission electron microscopy (HR-TEM) and energy-dispersive X-ray analysis spectroscopy (EDAX) were performed on an FEI Tecnai G2 F30 transmission electron microscope (at 300 kV, FEI, USA). Magnetic measurements were carried out on a magnetic property measurement system (MPMS XL-7, Quantum Design, San Diego, USA). The UV-vis absorption of different nanoparticle samples was measured with a UV-vis-NIR spectrophotometer (UV-3150, Shimadzu, Japan). The hydrodynamic size and surface potential of the nanoparticles were determined in aqueous phase by using a Malvern Zetasizer Nano-ZS (Malvern Instruments, Worcestershire, UK). The r₂ relaxivity was determined by a linear fitting of 1/T₂ as a function of the Fe concentration of the particles. The instrumental parameters were set as follows: point resolution of 156 × 156 mm², section thickness of 2 mm, TR of 5000 ms and number of signal acquisitions of 3.

The Supo HHS-6 electro-thermostatic water bath and the Evela N-1200A rotary evaporator were used. The freeze-drying method was operated by Biosafer Biosafer-10A and Christ Alpha 1–2 LD plus vacuum freeze-dryers. The size distribution and zeta potential were determined using a Malvern Zetasizer nano ZS analyzer. The UV used a Shimadzu UV-2600 UV-vis spectrophotometer, and the ATR-FTIR used a Perkin Elmer Spectrum two infrared analyzer. The JEOL JEM-2010HR was operated for TEM observation.
The cell cycles were performed by a Beckman CytoFLEX S flow cytometry analyzer. Cell proliferation was determined with a Multiskan FC ThermoFisher microplate reader. The quantitative RT-PCR was conducted with 7500 apparatus, Applied Biosystems (Waltham, MA, USA).

Freeze-fracture transmission electron microscopy (FF-TEM) was employed to characterize the morphology of UB-NLC [40 (link),41 (link),42 (link),43 (link),44 (link)]. For freeze-fracture (FF) measurements, droplets of the sample were dropped onto a specimen holder and frozen via a rapid plunge into liquid nitrogen. Then, samples were fractured and replicated using a freeze-etching apparatus (Balzers BAF 400D, Balzers, Liechtenstein). Platinum-carbon was deposited to shadow the replicas. Finally, the prepared samples were loaded onto copper grids and then observed by a transmission electron microscopy (JEOL JEM-2010HR, Tokyo, Japan).

The cyclic voltammetry (CV) and galvanostatic charge/discharge (GCD) measurements were carried out using a CHI 760D potentiostat (Shanghai Chenhua Instrument Co. Ltd., Shanghai, China) in both the three-electrode system and asymmetrical two-electrode configurations. In a three-electrode system, a saturated Ag/AgCl and a platinum (Pt) wire were used as reference electrode and counter electrode, respectively. For electrochemical impedance spectroscopy (EIS), the amplitude signal was 5 mV, and the frequency was scanned from the 10 kHz to 5 mHz. All the electrochemical measurements were performed in a 5 M LiCl solution at room temperature. The areal capacitance (C_A) of the supercapacitor was calculated from the CV curves on the basis of the following equation: $C_A=\frac{S}{2\Delta VAv}$
where S stands for the integral area of CV curves, v presents the scan rate, ΔV is the potential window, and A refers to the surface area of electrodes.
An X-ray diffractometer (D8 ADVANCE) (Bruker, Billerica, MA, USA) was used to study the phases of the samples. The samples underwent morphological analysis by using field emission scanning electron microscope (JSM-6330F) (JEOL Ltd., Akishima, Japan) and transmission electron microscopy (JEM2010-HR) (JEOL Ltd., Akishima, Japan).