β-Lactoglobulin Influences Human Immunity and Promotes Cell Proliferation

Figure 3

LG is transported into the cell cytoplasm after membranous LG-LG receptor binding process. (a) Immunofluorescence analysis by confocal microscopy (magnification = 1000x): FITC-LG (1 mg) was bound to cell membranes (green colored fluorescence) at 4°C after a 30 min incubation. (b) Flow cytometry: the cells were incubated with various doses of FITC-LG (0, 0.0625 mg, 0.125 mg, 0.25 mg, and 0.5 mg) at 37°C for 30 min and then their fluorescence intensities were assessed by flow cytometry. The results showed that, as the treatment dose increased, the fluorescence intensity detected in cells obviously increased (the peak shifts to the right or a higher intensity). (c) Immunofluorescence analysis by confocal microscopy: hybridoma cells were incubated using the following conditions: (1) negative control: no green fluorescence was observed in the cell membrane or the cytoplasm; (2) cells incubated with FITC-LG at 4°C for 30 min; green fluorescence was only observed on the cell membranes; (3) cells incubated with FITC-LG at 37°C for 5 min followed by an incubation at 4°C for 25 min: green fluorescence was observed in both the cell membrane and the cytoplasm; (4) cells incubated with FITC-LG at 37°C for 30 min: green fluorescence was observed mostly in the cytoplasm. (d) Flow cytometry further confirmed these results. The data indicated that FITC-LG could be transported into the cytoplasm after binding the cell membrane.

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