Internalization of antibody-bound wild-type CD33 in transduced cell lines. (A) 3 2D cell sublines, (B) OCI-A ML3 cell sublines, and (C) KG-1a cell sublines transduced with wild-type CD33 at different MOIs were labeled with unconjugated hP67.6 on ice water before the cells were incubated in 37°C in antibody-free medium to allow internalization of antibody-bound CD33 up to 4 hours as indicated. Subsequently, remaining cell surface–associated hP67.6 was detected with biotin-conjugated mouse anti–human IgG4 monoclonal antibody and a streptavidin-PE conjugate. Each left panel shows the percentage of internalized hP67.6 relative to cells kept at 0°C, whereas the right panels depict total amount of internalized hP67.6, expressed as change in arbitrary fluorescence units. Results are shown as mean ± SEM from 2 to 5 independent experiments.
