U hglgps fluorescence microscope

The cellular
uptake of PSFG coassembly polymeric micelles was evaluated by confocal
laser scanning microscopy (CLSM). The cells were seeded in 6-well
plates at a density of 2 × 10⁵ cells per well in 2
mL complete F12K media containing 10% fetal bovine serum, supplemented
with 100 U/mL penicillin and 100 U/mL streptomycin, and incubated
at 37 °C in 5% CO₂ atmosphere for 24 h. PSFG micelles
in a medium with the same concentration of FCy (10 μg/mL) were
added. After different time points, the plate was observed with an in vivo fluorescence imaging system or an Olympus U-HGLGPS
fluorescence microscope with or without replacing the original medium.
For a 6-well plate observed with a fluorescence microscope, 1 mM GSH,
which could imitate the reduction environment, was added to each well
for further observation. Then, the cells were washed with PBS, and
4% formaldehyde was added for 25 min of incubation. 4,6-Diamino-2-phenyl
indole (DAPI, 5 μg/mL) was further used for nucleus staining
(10 min). After washing with PBS, each well was observed with a fluorescence
microscope for cellular uptake evaluation. To better compare the cell-uptake
results, flow cytometry analysis was performed using a flow cytometer
(Beckman), which collected 10,000 gated events for each sample.

¹H-NMR spectra, using either DMSO-_d6 or CDCl₃ containing 0.03% (v/v) TMS as solvent, were measured on a Bruker AC 300 NMR spectrometer. UV-Vis spectra were measured on a Shimadzu UV-2401 PC Ultraviolet while fluorescence spectra were recorded on Shimadzu RF-5301PC. FT-IR spectra, applying a KBr method, were recorded on a Bruker EQUINOX 55 spectrometer. The molecular weights of the prepared samples (5mg/mL) were determined with gel permeation chromatography (GPC, KD-804 column and RID-10A refractive index detector), using DMF as the mobile phase at 60 ^oC. Raman spectra were recorded on a Bruker spectrometer with a 532nm laser. Dynamic light scattering (DLS) were measured via a Malvern Zetasizer Nano ZS90. JEOL-2010 microscope was used to obtain transmission electron microscopy (TEM) images. The fluorescence microscope imaging experiments were conducted on an Olympus U-HGLGPS fluorescence microscope. A Bio-rad iMark microplate reader was applied to record the MTT absorbance. The dissolved oxygen is estimated by a dissolved oxygen meter AZ 8403 at room temperature.