title: Isolation, culture and genetic manipulation of mouse pancreatic ductal cells.
abstract: The most common subtype of pancreatic cancer is pancreatic ductal adenocarcinoma (PDAC). PDAC resembles duct cells morphologically and, to some extent, at a molecular level. Recently, genetic-lineage labeling has become popular in the field of tumor biology in order to study cell-fate decisions or to trace cancer cells in the mouse. However, certain biological questions require a nongenetic labeling approach to purify a distinct cell population in the pancreas. Here we describe a protocol for isolating mouse pancreatic ductal epithelial cells and ductlike cells directly in vivo using ductal-specific Dolichos biflorus agglutinin (DBA) lectin labeling followed by magnetic bead separation. Isolated cells can be cultured (in two or three dimensions), manipulated by lentiviral transduction to modulate gene expression and directly used for molecular studies. This approach is fast (~4 h), affordable, results in cells with high viability, can be performed on the bench and is applicable to virtually all genetic and nongenetic disease models of the pancreas.
authors.affiliation: Division of Gastroenterology, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: Maximilian
authors.initials: M
authors.lastname: Reichert
authors.forename: Shigetsugu
authors.initials: S
authors.lastname: Takano
authors.forename: Steffen
authors.initials: S
authors.lastname: Heeg
authors.forename: Basil
authors.initials: B
authors.lastname: Bakir
authors.forename: Gregory P
authors.initials: GP
authors.lastname: Botta
authors.forename: Anil K
authors.initials: AK
authors.lastname: Rustgi
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D015870:Gene Expression; D016086:Lentivirus / Q000235:genetics; D051379:Mice; D010183:Pancreatic Ducts / Q000166:cytology*; D037121:Plant Lectins / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014161:Transduction, Genetic / Q000379:methods*
chemical_list: D037121:Plant Lectins; C036554:dolichos biflorus agglutinin