title: Transfection of Primary Cultures of Rat Hepatocytes.
abstract: Five different transfection reagents-calcium phosphate, TransFast™ Transfection Reagent, Superfect™ Transfection Reagent, Effectene™ Transfection Reagent, and Tfx™-20-were compared for their ability to effectively transfect primary cultures of male rat hepatocytes. Hepatocytes were isolated by the collagenase perfusion method and then cultured on Matrigel-coated plates for 24 h before transfection. The cells were transfected with either pGL3-Control or pGL3-Basic plasmids. The efficiency of transfection of each reagent was monitored using the dual luciferase reporter gene assay system. Superfect Transfection Reagent, Effectene Transfection Reagent and Tfx-20 were the most effective for the transfection of primary hepatocytes and gave comparable transfection efficiencies. Calcium phosphate was found to be the least effective transfection reagent and gave the most variable transfection results. Tfx-20 gave the least variable transfection results when different hepatocyte preparations were compared.
authors.affiliation: Department of Biochemistry and Molecular Biology, University College London, London, UK.
authors.forename: Mina
authors.initials: M
authors.lastname: Edwards
authors.affiliation: Institute of Molecular and Cell Biology, Proteos, Singapore.
authors.forename: Siew Cheng
authors.initials: SC
authors.lastname: Wong
authors.affiliation: Department of Clinical Chemistry, Faculty of Associated Medical Science, Chiang Mai University, Chiang Mai, Thailand.
authors.forename: Roongsiri
authors.initials: R
authors.lastname: Chotpadiwetkul
authors.affiliation: Department of Biochemistry, Hellenic Pasteur Institute, Athens, Greece.
authors.forename: Despina
authors.initials: D
authors.lastname: Smirlis
authors.affiliation: School of Biological Sciences, Queen Mary, University of London, London, UK.
authors.forename: Ian R
authors.initials: IR
authors.lastname: Phillips
authors.affiliation: Department of Biochemistry and Molecular Biology, University College London, London, UK.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Shephard
keywords: Hepatocytes; reporter gene assay; transfection