Cancer Management and Research (Feb 2019)
A modified method for isolating human quiescent pancreatic stellate cells
Abstract
Fu-Tao Meng,1,2,* Mei Huang,1,2,* Fang-Fang Fan,3,* Feng Shao,1,2 Chao Wang,1,2 Qiang Huang1,2 1Department of General Surgery, Anhui Provincial Hospital Affiliated to Anhui Medical University, Hefei, Anhui Province, People’s Republic of China; 2Anhui Province Key Laboratory of Hepatopancreatobiliary Surgery, Anhui Provincial Hospital, Hefei, Anhui Province, People’s Republic of China; 3Perelman School of Medicine, University of Pennsylvania, Pennsylvania, PA, USA *These authors contributed equally to this work Background: This study explored a simple, high-yield method for isolating quiescent human pancreatic stellate cells (PSCs) to provide sufficient and reliable raw materials for PSC-related studies. Materials and methods: Single-cell suspensions were prepared from normal human pancreatic tissue specimens using the gentleMACS™ tissue processor, which enhanced the yield and viability of the suspensions. Percoll density gradient centrifugation was then performed to isolate quiescent normal PSCs (NPSCs). Cell viability was determined by trypan blue staining, and the states of the NPSCs were determined by autofluorescence and oil red O staining. The purity of human activated PSCs (APSCs) was determined by immunofluorescence assays. Results: The yield of NPSCs was ~(2.75±0.65)×106 cells/g. The maximum cell viability was 92%, whereas the maximum cell purity was 95%. Conclusion: The method employed in this study to isolate PSCs is a simple, high-yield and stable method that is worth popularizing. Keywords: pancreatic stellate cells, gentleMACS™ Dissociator, Percoll, primary cell culture
