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A High Throughput Apoptosis Assay using 3D Cultured Cells

A high throughput apoptosis assay using 3D cultured cells was developed with a micropillar/microwell chip platform. Live cell apoptosis assays based on fluorescence detection have been useful in high content screening. To check the autofluorescence of drugs, controls (no caspase-3/7 reagent in the a...

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Autores principales: Lee, Sang-Yun, Doh, Il, Lee, Dong Woo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6767243/
https://www.ncbi.nlm.nih.gov/pubmed/31527418
http://dx.doi.org/10.3390/molecules24183362
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author Lee, Sang-Yun
Doh, Il
Lee, Dong Woo
author_facet Lee, Sang-Yun
Doh, Il
Lee, Dong Woo
author_sort Lee, Sang-Yun
collection PubMed
description A high throughput apoptosis assay using 3D cultured cells was developed with a micropillar/microwell chip platform. Live cell apoptosis assays based on fluorescence detection have been useful in high content screening. To check the autofluorescence of drugs, controls (no caspase-3/7 reagent in the assay) for the drugs are necessary which require twice the test space. Thus, a high throughput capability and highly miniaturized format for reducing reagent usage are necessary in live cell apoptosis assays. Especially, the expensive caspase-3/7 reagent should be reduced in a high throughput screening system. To solve this issue, we developed a miniaturized apoptosis assay using micropillar/microwell chips for which we tested seventy drugs (six replicates) per chip and reduced the assay volume to 1 µL. This reduced assay volume can decrease the assay costs compared to the 10–40 µL assay volumes used in 384 well plates. In our experiments, among the seventy drugs, four drugs (Cediranib, Cabozatinib, Panobinostat, and Carfilzomib) induced cell death by apoptosis. Those results were confirmed with western blot assays and proved that the chip platform could be used to identify high potency apoptosis-inducing drugs in 3D cultured cells with alginate.
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spelling pubmed-67672432019-10-02 A High Throughput Apoptosis Assay using 3D Cultured Cells Lee, Sang-Yun Doh, Il Lee, Dong Woo Molecules Technical Note A high throughput apoptosis assay using 3D cultured cells was developed with a micropillar/microwell chip platform. Live cell apoptosis assays based on fluorescence detection have been useful in high content screening. To check the autofluorescence of drugs, controls (no caspase-3/7 reagent in the assay) for the drugs are necessary which require twice the test space. Thus, a high throughput capability and highly miniaturized format for reducing reagent usage are necessary in live cell apoptosis assays. Especially, the expensive caspase-3/7 reagent should be reduced in a high throughput screening system. To solve this issue, we developed a miniaturized apoptosis assay using micropillar/microwell chips for which we tested seventy drugs (six replicates) per chip and reduced the assay volume to 1 µL. This reduced assay volume can decrease the assay costs compared to the 10–40 µL assay volumes used in 384 well plates. In our experiments, among the seventy drugs, four drugs (Cediranib, Cabozatinib, Panobinostat, and Carfilzomib) induced cell death by apoptosis. Those results were confirmed with western blot assays and proved that the chip platform could be used to identify high potency apoptosis-inducing drugs in 3D cultured cells with alginate. MDPI 2019-09-16 /pmc/articles/PMC6767243/ /pubmed/31527418 http://dx.doi.org/10.3390/molecules24183362 Text en © 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Technical Note
Lee, Sang-Yun
Doh, Il
Lee, Dong Woo
A High Throughput Apoptosis Assay using 3D Cultured Cells
title A High Throughput Apoptosis Assay using 3D Cultured Cells
title_full A High Throughput Apoptosis Assay using 3D Cultured Cells
title_fullStr A High Throughput Apoptosis Assay using 3D Cultured Cells
title_full_unstemmed A High Throughput Apoptosis Assay using 3D Cultured Cells
title_short A High Throughput Apoptosis Assay using 3D Cultured Cells
title_sort high throughput apoptosis assay using 3d cultured cells
topic Technical Note
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6767243/
https://www.ncbi.nlm.nih.gov/pubmed/31527418
http://dx.doi.org/10.3390/molecules24183362
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