Figure 7.

Characterization of the murine NB cell line NXS2-HGW. (A-C) Representative immunohistochemical images of GD₂ expression (left) and respective negative controls (right). NXS2-HGW cells cultivated onto chamber slides (A-B) and primary tumors obtained from A/J mice inoculated subcutaneously with NXS2-HGW cells (C) were analyzed for GD₂-expression (magnification of 100x (A), 200x (B) and 400x (C)). (D) PD-L1 expression analysis by the parental NXS2 cells (left histogram) in comparison with NXS2-derived NXS2-HGW cells (right histogram) by flow cytometry. Cells were stained with PE-labeled anti-mouse PD-L1 Ab (filled black curve) relative to isotype control (open gray curve). Results are shown as representative histograms from at least 5 independent experiments. (E) RT-PCR analysis of MYCN- (product size: 248 bp), TH- (187 bp) and PD-L1 (180 bp) mRNA in NXS2-HGW cells. GAPDH (223 bp) served as internal control. NTC - no template control. (F) GD₂ expression by the parental NXS2 cells in comparison with NXS2-derived NXS2-HGW cells (passage 49 for NXS2 and passage 1, 12 and 29 for NXS2-HGW) by flow cytometry. Cells were stained with chimeric ch14.18/CHO and PE-labeled anti-human IgG served as primary and secondary antibody, respectively (filled black curve). Rituximab served as isotype control (open gray curve). Results are shown as representative histograms from independent experiments.