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. 2018 Nov 14;19(11):3592. doi: 10.3390/ijms19113592

Figure 2.

Figure 2

Expression of CXCR4 and/or ACKR3 in transfected CHO cells. CHO cells were transfected with pcDNA3.1/Zeo-CXCR4 and/or pcDNA3-ACKR3 using Effectene and selected with antibiotics. (A) Histograms showing CXCR4 or ACKR3 expression in the final selected colonies. Cells were stained with PE or APC-conjugated antibodies and expression was assessed using flow cytometry (Red = Isotype, blue = antibody) on CHO-CXCR4 (left) and CHO-ACKR3 (middle); and red = isotype for CXCR4, purple = isotype for ACKR3, green = CXCR4, blue = ACKR3 on CHO-CXCR4-ACKR3 (right) cells. (B) Mean fluorescence levels of the transfected CHO cells were determined and compared to several breast cancer cell lines to assess CXCR4 (left) and ACKR3 (right) receptor levels. Data represents the mean ± SEM of three independent experiments and statistical significance was calculated using a one way ANOVA (* p < 0.05). (C) Formation of heterodimers was assessed using fluorescence resonance energy transfer (FRET) and quantified through the FRET ratio from APC (the acceptor fluorochrome). Data represents the mean ± SEM of three independent experiments and statistical significance was calculated using a one way ANOVA (* p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001).