Xenogen ivis imaging system 200 series

In vitro transfection efficiency of polyplexes in HCASMCs was assessed by a luciferase gene reporter assay. HCASMCs were cultured in 96-well plates (12 000 cells per well for ~70–80% confluent) in SMC medium for 24 h. Cells were then treated with LPS (1 μg mL⁻¹) and IFN-γ (100 units per mL) for 6 h to induce ROS production, followed by treatment with PPDDBP and LF2K polyplexes at N/P 10 ratio for 24 h at 37 °C. Luciferin (150 μg mL⁻¹) was added to the samples, and bioluminescence from cells was measured using a Xenogen IVIS Imaging system 200 series (Perkin Elmer, Waltham, MA, USA). Cells were then washed twice with DPBS and lysed with 200 μL of lysis buffer (Promega, Madison, WI, USA). The protein content was using a microBCA protein assay reagent kit (Pierce) and reading absorbance at 562 nm using the M1000 Pro platen reader. The luciferase expression was reported in relative light units (RLU) normalized to mg protein (RLU per mg).

All animals treated with GFP⁺ Luc⁺ cells, across all groups, were evaluated at 7 day intervals with in vivo fluorescence optical imaging to evaluate persistence, distribution, and relative signal of donor cells using a Xenogen IVIS Imaging System 200 Series (PerkinElmer, MA, USA) [15 (link)].