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. 2005 Nov 30;46(2-3):139–150. doi: 10.1007/s10616-005-2551-7

Evaluation of a Continuous Quantification Method of Apoptosis and Necrosis in Tissue Cultures

Debby Gawlitta 1,✉, Cees W J Oomens 1, Frank P T Baaijens 1, Carlijn V C Bouten 1
PMCID: PMC3449716  PMID: 19003268

Abstract

In tissue-engineering and other life sciences, there is a growing need for real-time, non-destructive information on apoptosis and necrosis in 2D and 3D tissue cultures. Previously, propidium iodide was applied as a fluorescent marker for monitoring necrosis. In the current study this technique was extended with a fluorescent apoptosis marker, YO-PRO-1, to discriminate between both stages of cell death. The main goal was to evaluate the performance of YO-PRO-1 and propidium iodide during monitoring periods of up to 3 days. Apoptosis was induced in C2C12 cultures and the numbers of YP-positive and PI-positive nuclei were counted in time. The performance of the dual staining was evaluated with a metabolic measure and a probe intensity study. Cell metabolism was unaffected during the first 24 h of testing. In conclusion, the YP/PI dual staining method was found to be a powerful tool in obtaining real-time spatial information on viability in cell and tissue culture without culture disruption.

Keywords: Apoptosis, Monitoring, Necrosis, Propidium iodide, Quantification, YO-PRO-1

Full Text

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Glossary

DM

differentiation medium

GM

growth medium

PI

propidium iodide

YP

YO-PRO-1

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