In spite of advances in surgical and medical care, pancreatic cancer remains a leading cause of cancer-related death in the United States. An understanding of cancer-cell interactions with host cells is critical to our ability to develop effective antitumor therapeutics for pancreatic cancer. We report here a color-coded model system for imaging cancer cell interactions with host immune cells within the native pancreas. A human pancreatic cancer cell line engineered to express green fluorescent protein (GFP) in the nucleus and red fluorescent protein (RFP) (DsRed2) in the cytoplasm was orthotopically implanted into the pancreas of a nude mouse. After 10-14 days, red or green fluorescent splenocytes from immune-competent transgenic-mouse donors expressing RFP and GFP, respectively, were delivered systemically to the pancreatic cancer-bearing nude mice. Animals were imaged after splenocyte delivery using high-resolution intravital imaging systems. At 1 day after iv injection red- or green-fluorescent spleen cells were found distributed in lung, liver, spleen and pancreas. By 4 days after cell delivery, however, the immune cells could be clearly imaged surrounding the tumor cells within the pancreas as well as collecting within lymphatic tissues such as lymph nodes and spleen. With the high-resolution intravital imaging afforded by the Olympus IV100 and OV100 systems, the interactions of the dual-colored cancer cells and the red- or green-fluorescent spleen cells could be clearly imaged in this orthotopic pancreatic cancer model. This color-coded in vivo imaging technology offers a novel approach to imaging the interactions of cancer and immune cells in the tumor microenvironment (TME).