Figure 3.

The binding ability of bCAT1/SLC7A1 with BLV particles based on flow cytometry analysis. CHO-K1 cells were transfected with or without bCAT1/pEGFP-N1. After 48 h cultivation, the cells were incubated with BLV particles purified from the culture supernatant of FLK-BLV cells for 1 h (A) or 2 h (B) at 4°C and then labeled using anti-gp51 mAb (BLV-1) followed by incubation with APC-conjugated anti-mouse IgG. After staining, cells were analyzed using a BD Accuri C6 Plus flow cytometer. Each profile was separated into 4 quadrants, which signified single-positive staining (red for BLV or green for CAT1-EGFP), double-negative staining, or double-positive staining. C) The binding ability of CAT1/SLC7A1 and BLV particles significantly decreased by adding BLV serum. CC81-GREMG cells were added either with or without BLV serum (red) or control serum (blue) (1:10 and 1:50 dilution) and BLV particles that purified from the culture supernatant of FLK-BLV cells to bind for 2 h at 4°C. The cells were washed and then labeled using anti-gp51 antibody followed by incubation with APC-conjugated anti-mouse IgG. After staining, cells were analyzed using a BD Accuri C6 Plus flow cytometer. Staining cells expressed as histograms in profile. The dotted line indicated the CC81-GREMG cells only as negative control. The gray solid curve indicated cells were added BLV particles only as positive control. The percentage of BLV particles binding cells expressed as a graph (lower column). Each column and error bar represents the means + sd of results from triplicate experiments. **P = 0.01, ***P = 0.001 (Student’s t test).