HEK293 cells were maintained in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum, 100 units/ml penicillin, and 100 μg/ml streptomycin. As described previously, human intestinal organoid cultures and passages were conducted (40 ). Briefly, organoids were cultured with a 25% L-WRNH–conditioned medium embedded in Matrigel Matrix (Corning). The entire medium was changed every 2 to 3 days, and passages were performed every 5 to 6 days. Following passages, cell dissociation was performed using TrypLE express (Corning) at a general ratio of 1:8 embedded in Matrigel Matrix on a Nunclon Delta 4-well dish (Thermo Scientific). All cells were cultured at 37 °C in a 5% CO2 atmosphere. The experiments using primary human organoids complied with the Declaration of Helsinki and were approved by the human ethical committee of The University of Tokyo and Osaka University. All tissues were sampled with informed consent.
Tryple express
Manufactured by Corning
TrypLE Express is a ready-to-use solution for detaching adherent cells from cell culture surfaces. It is a formulation of enzymes and other components designed to efficiently dissociate cells from substrates while maintaining cell viability and functionality.
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Lab products found in correlation
4 protocols using tryple express
1
Culturing Human Intestinal Organoids
2
Evaluating Epigenetic Drug Toxicity in 2D Tumor Cells and Organoids
3
Glioblastoma Spheroid Formation for Invasion
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To form GBM6 spheroids for invasion studies, transduced GBM6 cells were dissociated using TrypLE Express and added to 96‐well spheroid low‐attachment plates (5000 cells per well) (Corning). Plates were centrifuged and then incubated on a rotating platform (60 RPM) at 37 °C for 48 h.
4
Evaluating Epigenetic Drug Toxicity in 2D Tumor Cells and Organoids
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For 2D tumour cell culture, tumour cells were seeded into 96 well plates at 2,000 cells per well and cultured for 72 h. For tumour organoids, tumour organoids with diameter between 70 and 150 μm were seeded at 50 organoids per well into 96-well microplates with ultra-low attachment surface (Corning). Cells or tumour organoids were treated with indicated compound at the final concentration of 1μM. After 3-day culture, tumour organoids were dissociated into single cells using TrypLE Express (Corning) at 37°C with a shaking velocity of 500 rpm on a thermomixer (Eppendorf) for 15 min. The 96-well microplates were added with 10 μl/well of Premix WST-1 reagent (TaKaRa) and incubated for 1 h. The microplates were read at absorbance of 450 nm on the Epoch microreader (BioTek). The toxicity of the epigenetic drugs was quantified using the Premix WST-1 reagent according to the manufacturer’s protocol.
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