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. 2018 Sep 21;8:14224. doi: 10.1038/s41598-018-32664-z

Figure 4.

Figure 4

Heterologous expression of tick GALTs in HL-60 and E. coli cells is associated to α-Gal synthesis. (A) HL-60 cells were transfected with b4galt7, a4galt-1 and a4galt-2 in a fusion protein expression system that uses Red Fluorescent Protein (RFP) as a reporter of heterologous gene expression. α-Gal production was then measured by immunofluorescence. Empty plasmid was used as control. Host cell nucleus was stained with DAPI (blue). The α-Gal-specific monoclonal antibody M86 (primary antibody) and the goat anti-mouse IgM-FITC antibody (secondary antibody) were used to detect the production of α-Gal (green). RFP was also detected in human cells (red). Merged images show that the presence of α-Gal was observed exclusively in cells with heterologous gene expression (arrows). Images are at magnification X 63. (B) E. coli BL21 cells were transformed with plasmids containing b4galt7, a4galt-1 and a4galt-2. α-Gal production was then measured by immunofluorescence. Empty plasmid was used as control. Host cell nucleus was stained with DAPI (blue). The α-Gal-specific monoclonal antibody M86 (primary antibody) and the goat anti-mouse IgM-FITC antibody (secondary antibody) were used to detect the production of α-Gal (green). Merged images show the presence of α-Gal in E. coli BL21 (arrows). Images are at magnification higher than X 63. Bars represent 10 µm.