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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...
Autores principales: | , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2019
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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. |
format | Online Article Text |
id | pubmed-6767243 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
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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