A High Throughput Apoptosis Assay using 3D Cultured Cells.

Area of Science:

• Biotechnology
• Cell Biology
• Drug Discovery

Background:

• Live cell apoptosis assays are crucial for high-content screening.
• Current methods require large reagent volumes and test space, increasing costs.
• Miniaturization is needed to reduce expenses, especially for expensive reagents like caspase-3/7.

Purpose of the Study:

• To develop a high-throughput and miniaturized apoptosis assay using a micropillar/microwell chip platform.
• To reduce reagent usage and assay costs for screening drugs in 3D cell cultures.
• To validate the chip platform's efficacy in identifying apoptosis-inducing drugs.

Main Methods:

• Development of a micropillar/microwell chip for 3D cell culture.
• Implementation of a miniaturized live cell apoptosis assay with a 1 µL assay volume.
• Screening of seventy drugs with six replicates per chip.
• Confirmation of apoptosis-inducing drugs using western blot assays.

Main Results:

• The chip platform successfully tested seventy drugs in a miniaturized format.
• Four drugs (Cediranib, Cabozatinib, Panobinostat, and Carfilzomib) were identified as potent apoptosis inducers.
• The assay volume was reduced to 1 µL, significantly lowering costs compared to 384-well plates.
• Results were validated through western blot analysis.

Conclusions:

• The developed micropillar/microwell chip platform offers a high-throughput, cost-effective solution for apoptosis assays.
• This technology enables efficient identification of apoptosis-inducing drugs in 3D cultured cells.
• The miniaturized assay format significantly reduces reagent consumption and associated costs.