Figure 2

Transduction of mouse fibroblasts with IDO-expressing lentiviral vector. a Fluorescence microscopy analysis of IDO-lentiviral vector-transduced fibroblasts. Cultured mouse fibroblasts were incubated with IDO-lentiviral vector either in the presence or absence of polybrene. Cells that were transduced in the presence of polybrene were then sorted by FACS. Panels a and e, b and f, c and g, as well as d and h represent bright-field and fluorescence images of non-transduced fibroblasts, IDO-lentiviral vector-transduced fibroblasts, fibroblasts transduced with IDO-expressing lentiviral vector in the presence of polybrene, and fibroblasts transduced with IDO-expressing lentiviral vector in the presence of polybrene after sorting, respectively. b These panels show the Western blot analysis (a) as well as kynurenine levels (b) in different indicated cells. The data shown are the mean and standard deviation of kynurenine measurements obtained from three different experiments. c Flow cytometry analysis of the IDO-expressing cell population. Fibroblasts were left either non-transduced or transduced with IDO-expressing lentiviral vector in the absence or presence of polybrene and flow cytometry was then conducted by gating the cells based on their mCherry fluorescence. The selected cell population was prepared by cell sorting and the number of mCherry-positive cells was greater than 95%. Panel a shows non-transduced fibroblasts, b transduced IDO-expressing fibroblasts in the absence of polybrene, c transduced IDO-expressing fibroblasts in the presence of polybrene, and d FACS-sorted IDO-expressing fibroblasts (in the presence of polybrene). Panel e shows the percentage of mCherry-positive fluorescent cells in the total cell population studied as detected by flow cytometry. The data shown are the mean and standard deviation of mCherry-positive cells obtained from three separate experiments.