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Development of a miniaturized 3D organoid culture platform for ultra-high-throughput screening
Yuhong Du1,* , Xingnan Li2 , Qiankun Niu1 , Xiulei Mo1 , Min Qui1 , Tingxuan Ma1 , Calvin J. Kuo2 , Haian Fu1
1Department of Pharmacology and Chemical Biology and Emory Chemical Biology Discovery Center, Emory University School of Medicine, Atlanta, GA, USA
2Department of Medicine, Division of Hematology, Stanford University School of Medicine, Stanford, CA, USA
*Correspondence to:Yuhong Du ,
J Mol Cell Biol, Volume 12, Issue 8, August 2020, Pages 630-643
Keyword: human colon organoids, KRASG12D, 3D culture, 384-well plate, high-throughput screening (HTS), 1536-well plate, ultra-HTS (uHTS)

The recent advent of robust methods to grow human tissues as 3D organoids allows us to recapitulate the 3D architecture of tumors in an in vitro setting and offers a new orthogonal approach for drug discovery. However, organoid culturing with extracellular matrix to support 3D architecture has been challenging for high-throughput screening (HTS)-based drug discovery due to technical difficulties. Using genetically engineered human colon organoids as a model system, here we report our effort to miniaturize such 3D organoid culture with extracellular matrix support in high-density plates to enable HTS. We first established organoid culturing in a 384-well plate format and validated its application in a cell viability HTS assay by screening a 2036-compound library. We further miniaturized the 3D organoid culturing in a 1536-well ultra-HTS format and demonstrated its robust performance for large-scale primary compound screening. Our miniaturized organoid culturing method may be adapted to other types of organoids. By leveraging the power of 3D organoid culture in a high-density plate format, we provide a physiologically relevant screening platform to model tumors to accelerate organoid-based research and drug discovery.