Ix53 fluorescent microscope

The morphology of nanoparticles was measured using a transmission electronic microscope (TEM, FEI Tecnai G2) at an accelerating voltage of 200 kV. Zeta potential was recorded on Malvern Zetasizer Nano Series. Scanning electron microscopy (SEM) images were taken on a FEI Nova NanoSEM 230 under 10 kV acceleration voltage with a secondary electron detector. The samples were coated using Emitech K575× evaporative premium coater prior to imaging. X‐ray diffraction (XRD) measurements were performed on powder and film samples using PANalytical X‐ray diffraction system with a CoKα and CuKα source respectively. The angles were converted to CuKα source for comparison using HighScore Plus software. Synchrotron terahertz infrared spectroscopy (THz‐FIR) was conducted on Bruker IFS 125/HR Fourier Transform (FT) spectrometer. The fluorescence intensity of cells and nanoparticles was examined by using BD LSRFortessa SORP X‐20 analyzing flow cytometer and Olympus IX53 fluorescent microscope. ¹H NMR spectra were acquired by dissolving the ZIF‐8 NPs in diluted HCl with D₂O (1/99 v/v) at 25 °C on a Bruker Avance III 400 MHz NMR (Rabi). MRI measurements were carried on the 9.4 T Bruker BioSpec Avance III 94/20 system (Bruker, Ettlingen, Germany). The temperature change during the irradiation was recorded by an IR thermal camera. Flow cytometry data were analyzed and represented by Flowjo software.

Exosomes and PKH26 were incubated for 30 min and centrifuged at 4°C for 60 min. After incubation with LoVo cells, PKH26-labeled exosomes were stained with 4ʹ,6-diamidino-2-phenylindole 2 and viewed under a IX53 fluorescent microscope (Olympus, Tokyo, Japan).
To explore the transfer of miR-431-5p, FITC-miR-431-5p was packaged into exosomes using electroporation, and isolated from 300 μl hUCMSCs cell culture-conditioned medium and packaged with FITC-miR-431-5p. Briefly, 30 μl FITC-miR-431-5p was added to 200 ng exosomes in 400 μl electroporation buffer (21% Optiprep, 25 mm KCl, 100 mm potassium phosphate, pH 7.2) and transferred to a 4-mm electroporation tube (Eppendorf, Hauppauge, NY, USA). Samples were electroporated in an E2510 electroporator (Eppendorf, Hauppauge, NY, USA) with three pulses at 0–2000 V and stored at 4°C for 5 min. The exosomes containing FITC-miR-431-5p were labeled with Dil (red), and finally incubated with LoVo cells for 48 h. The co-localization of FITC and Dil was observed in recipient cells using a fluorescence microscope (ECLIPSE E800, Nikon, Japan).