Lx71 epifluorescence microscope

Rhodamine B-labeled particles were reacted with hydroxyl group in HER-PEG-PCL and PEG-PCL copolymers as previously described [33 (link)]. The labeled NPs were loaded with FAM-labeled AMO-21 (FAM-AMO-21) and the fluorescent signals were used to track the location of AMO-21 and NPs in NUGC4 and SGC7901 cells. Cells were loaded in 6-well plates at a density of 2×10⁵ cells/well and incubated overnight. An additional incubation for 2 h was performed using a fixed concentration of FAM-AMO-21-loaded Rhodamine B-labeled HER-PEG-PCL and PEG-PCL NPs. The cells were washed three times with PBS, and fixed with 4% paraformaldehyde (PFA) for 30 min at room temperature. Following another round of washing the media were replaced with fresh culture media. The fluorescence was determined using an Olympus LX71epifluorescence microscope (Olympus, Tokyo, Japan). For quantification, the color change in the PEG-PCL and HER-PEG-PCL groups was detected by the naked eye and analyzed using image analysis software (ImageJ). The integrated optical density (IOD) was used to quantify the fluorescence. The IOD of Rhodamine B and FAM were evaluated by performing tests under identical experimental conditions, in triplicate