Objective：To develop an in vitro peroxisome proliferator-activated receptor（PPAR）γ ligand screening system for identify－ing novel compounds with PPARγ agonist activity. Methods：Human PPARγ plasmid（pIRES2-PPARγ），firefly luciferase reporter plasmid（pPPRE×3-TK-Luciferase） and an internal reference plasmid（pRL-TK）were cotransfected into HEK293 cells by using LipofectamineTM 2000，and the optimal ratio of three plasmids was determined. Luciferase expression intensity was determined to assess the PPARγ agonist activity after the potential ligand being added to cotransfected HEK293 cells. Specificity of the model was exam－ined by treatment of PPARγ agonist and PPARγ specific antagonist and several nuclear receptor agonists and PPARα agonist；re－peatability was also determined by Z’ value and time characteristics of PPAR agonist action was observed. Results：PPARγ plasmid，reporter gene vector and internal control plasmid ratio of 2∶6∶1 was proved to be suitable for highest relative luciferase activity. PPARγ agonist rosiglitazone can significantly increase the relative luciferase activity and co-treatment with PPARγ specific antago－nist GW9662 resulted in significantly reduced expressions of luciferase and increased relative luciferase activity in the HEK293 cells in a time dependent manner. Dexamethasone，all-trans retinoic acid，17β-estradiol and fenobrate did not alter the relative luciferase activity. Value of Z’ was 0.70 after repeating experiments for 9 times. Conclusions：An in vitro PPARγ agonist screening system is developed with acceptable specificity and repeatability. The model can be used for PPARγ partial agonist screening from synthetic or natural compounds.