Evaluation of a continuous quantification method of apoptosis and necrosis in tissue cultures.
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.
Gawlitta
Debby
D
Department of Biomedical Engineering, Eindhoven University of Technology, WH 4.105, P.O. Box 513, 5600 MB, Eindhoven, The Netherlands, d.gawlitta@tue.nl.
Oomens
Cees W J
CW
Baaijens
Frank P T
FP
Bouten
Carlijn V C
CV