Detection and quantification of macropinosomes in pancreatic tumors

Abstract

Macropinocytosis is a mechanism of fluid-phase endocytosis that functions in the nonspecific internalization of extracellular fluid. This uptake pathway has specialized roles in different cell types and organisms, and its importance has recently been established in several diseases, including cancer. In cancer, macropinocytosis is stimulated by oncogenes, such as Ras, and macropinocytic cargo is targeted to lysosomes for degradation, providing a catabolic route for tumor cells to obtain amino acids from the tumor microenvironment. Here, we describe a protocol to employ fluorescently labeled dextran molecules in order to visualize and quantify the extent of macropinocytosis in pancreatic tumors. Multiple samples can be processed in parallel by this method, and the protocol can be easily customized for pancreatic tumor tissue isolated from subcutaneous, orthotopic and genetically engineered mouse models (GEMM), or human patients.