CellTrace Violet Cell Proliferation Kit Clearly Labels Living Tumor Cells

Protocol Overview

Tumor cells in suspension were collected and then washed. Cells were resuspended in either warm PBS alone or warm PBS containing 5 uM Cell Trace Violet prepared according to manufacturer's instructions. Cells were incubate for 20 minutes at 37 degrees Celsius. Cells were then washed, labelled with the appropriate antibody on ice for 20 minutes, then resuspended in warm media and were incubated for 3-24 hours at 37 degrees Celsius with or without macrophages. After incubation, cells were washed and resuspended in PBS containing Fc block (diluted 1:250) and LIVE/DEAD (diluted 1:5,000), using 100 uL for one million cells). Cells were incubated on ice in the dark for 20 minutes. Cells were washed and resuspended in FACS buffer containing the appropriate cell surface markers (diluted 1:500). Cells were incubated on ice in the dark for 20 minutes.