NUS Graduate School for Integrative Sciences and Engineering 1 article published in JoVE Biology Isolation of Primary Rat Hepatocytes with Multiparameter Perfusion Control Inn Chuan Ng*1, Li Zhang*2, Narelle Nichola Yi Ying Shen3, Yun Ting Soong4, Chan Way Ng5, Phoebe Kang Sheing Koh1, Yan Zhou3, Hanry Yu1,3,4,5,6 1Department of Physiology & The Institute for Digital Medicine (WisDM), National University of Singapore, 2College of Agriculture and Biology, Zhongkai University of Agriculture and Engineering, 3Mechanobiology Institute, National University of Singapore, 4Institute of Bioengineering and Nanotechnology, A*STAR, 5NUS Graduate School for Integrative Sciences and Engineering, 6CAMP, Singapore-MIT Alliance for Research and Technology This protocol details the use of a special intravenous catheter, standardized sterile disposable tubing, temperature control complemented by real-time monitoring, and an alarm system for two-step collagenase perfusion procedure to improve the consistency in the viability, yield, and functionality of isolated primary rat hepatocytes.