Fig. 2. Functional ST6GAL-1 is variably expressed in breast cancer cells.

A mRNA levels of ST6GAL1 were determined from ER + and TNBC human and mouse breast cancer cells, as indicated by quantitative real-time PCR (qPCR) and normalized to GAPDH (2^-delta Ct). N = 3, data are means ± s.e. B Total cell lysates from separate cell cultures (A) were used for Western blot analysis with antibodies against ST6GAL1. β-tubulin was used as a specific marker for cytosol and to show equal loading and transfer. C As indicated, an equal amount of proteins from cell lysates (B) was used for the ST6GAL1 enzyme assay, as previously mentioned [65]. ST6GAL1 activity is presented as fmol/min/mg protein; enzyme assays were performed in triplicates. Data are mean ± s.e. D Human and mouse representative ER+ and TNBC cell lines, as indicated, were immunostained with FITC- (green) labeled SNA lectin and DAPI (blue) for the nuclei before fixing. During fluorescent microscopy, exposure time and weighting for both DAPI and FITC fluorescence were kept consistent between samples. Representative images (N = 4) were shown on a scale bar of 50 µm. E Images were processed in ImageJ; background subtraction and MFI per cell calculations were carried out using the same parameters for each condition, and mean fluorescence intensity (MFI) per cell was shown with mean ± s.d. for four fields of view.