Skip to main content
SpringerLink
Log in

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

  • Published:
Cytotechnology Aims and scope Submit manuscript

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.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Abbreviations

DM:

differentiation medium

GM:

growth medium

PI:

propidium iodide

YP:

YO-PRO-1

References

  • F.G. Blankenberg J.F. Tait H.W. Strauss (2000) ArticleTitleApoptotic cell death: its implications for imaging in the next millennium Eur. J. Nucl. Med. 27 359–367 Occurrence Handle10.1007/s002590050046 Occurrence Handle10774891 Occurrence Handle1:STN:280:DC%2BD3c3jsV2htg%3D%3D

    Article  PubMed  CAS  Google Scholar 

  • D.J. Boffa J. Waka D. Thomas S. Suh K. Curran V.K. Sharma M. Besada T. Muthukumar et al. (2005) ArticleTitleMeasurement of apoptosis of intact human islets by confocal optical sectioning and stereologic analysis of YO-PRO-1-stained islets Transplantation 79 842–845 Occurrence Handle10.1097/01.TP.0000155175.24802.73 Occurrence Handle15818328 Occurrence Handle1:CAS:528:DC%2BD2MXjtVOjtb0%3D

    Article  PubMed  CAS  Google Scholar 

  • R.G. Breuls C.V. Bouten C.W. Oomens D.L. Bader F.P. Baaijens (2003a) ArticleTitleCompression induced cell damage in engineered muscle tissue: an in vitro model to study pressure ulcer aetiology Ann. Biomed. Eng. 31 1357–1364 Occurrence Handle10.1114/1.1624602 Occurrence Handle1:STN:280:DC%2BD2c%2FlslOisQ%3D%3D

    Article  CAS  Google Scholar 

  • R.G. Breuls A. Mol R. Petterson C.W. Oomens F.P. Baaijens C.V. Bouten (2003b) ArticleTitleMonitoring local cell viability in engineered tissues: a fast quantitative and non-destructive approach Tissue Eng. 9 269–281 Occurrence Handle10.1089/107632703764664738

    Article  Google Scholar 

  • K.T. Chung (1983) ArticleTitleThe significance of azo-reduction in the mutagenesis & carcinogenesis of azo dyes Mutat. Res. 114 269–281 Occurrence Handle6339890 Occurrence Handle1:CAS:528:DyaL3sXitFSru78%3D

    PubMed  CAS  Google Scholar 

  • T. Idziorek J. Estaquier F. Bels ParticleDe J.C. Ameisen (1995) ArticleTitleYOPRO-1 permits cytofluorometric analysis of programmed cell death (apoptosis) without interfering with cell viability J. Immunol. Methods 185 249–258 Occurrence Handle10.1016/0022-1759(95)00172-7 Occurrence Handle7561136 Occurrence Handle1:CAS:528:DyaK2MXotFCkt7Y%3D

    Article  PubMed  CAS  Google Scholar 

  • K.R. Jerome Z. Chen R. Lang M.R. Torres J. Hofmeister S. Smith R. Fox C.J. Froelich L. Corey (2001) ArticleTitleHSV and glycoprotein J inhibit caspase activation and apoptosis induced by granzyme B or Fas J. Immunol. 167 3928–3935 Occurrence Handle11564811 Occurrence Handle1:CAS:528:DC%2BD3MXntFajtL8%3D

    PubMed  CAS  Google Scholar 

  • X. Liu T. Vleet ParticleVan R.G. Schnellmann (2004) ArticleTitleThe role of calpain in oncotic cell death Annu. Rev. Pharmacol. Toxicol. 44 349–370 Occurrence Handle10.1146/annurev.pharmtox.44.101802.121804 Occurrence Handle14744250 Occurrence Handle1:CAS:528:DC%2BD2cXhvVCktLs%3D

    Article  PubMed  CAS  Google Scholar 

  • A. McArdle A. Maglara P. Appleton A.J. Watson I. Grierson M.J. Jackson (1999) ArticleTitleApoptosis in multinucleated skeletal muscle myotubes Lab. Invest. 79 1069–1076 Occurrence Handle10496525 Occurrence Handle1:STN:280:DyaK1MvisVyntg%3D%3D

    PubMed  CAS  Google Scholar 

  • T.F. McGuire D.L. Trump C.S. Johnson (2001) ArticleTitleVitamin D(3)-induced apoptosis of murine squamous cell carcinoma cells. Selective induction of caspase-dependent MEK cleavage and up-regulation of MEKK-1 J. Biol. Chem. 276 26365–26373 Occurrence Handle11331275 Occurrence Handle1:CAS:528:DC%2BD3MXlsVKntr0%3D

    PubMed  CAS  Google Scholar 

  • T. Mosmann (1983) ArticleTitleRapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays J. Immunol. Methods 65 55–63 Occurrence Handle10.1016/0022-1759(83)90303-4 Occurrence Handle6606682 Occurrence Handle1:STN:280:BiuD1cbgsFE%3D

    Article  PubMed  CAS  Google Scholar 

  • J.C. Park Y.S. Hwang H. Suh (2000) ArticleTitleViability evaluation of engineered tissues Yonsei Med. J. 41 836–844 Occurrence Handle11204834 Occurrence Handle1:CAS:528:DC%2BD3MXhtVaqsr8%3D

    PubMed  CAS  Google Scholar 

  • M. Rieseberg C. Kasper K.F. Reardon T. Scheper (2001) ArticleTitleFlow cytometry in biotechnology Appl. Microbiol. Biotechnol. 56 350–360 Occurrence Handle10.1007/s002530100673 Occurrence Handle11549001 Occurrence Handle1:CAS:528:DC%2BD3MXmsFKhur4%3D

    Article  PubMed  CAS  Google Scholar 

  • C. Stadelmann H. Lassmann (2000) ArticleTitleDetection of apoptosis in tissue sections Cell. Tissue Res. 301 19–31 Occurrence Handle10.1007/s004410000203 Occurrence Handle10928278 Occurrence Handle1:CAS:528:DC%2BD3cXlsFGnsrs%3D

    Article  PubMed  CAS  Google Scholar 

  • T. Suzuki K. Fujikura T. Higashiyama K. Takata (1997) ArticleTitleDNA staining for fluorescence and laser confocal microscopy J. Histochem. Cytochem. 45 49–53 Occurrence Handle9010468 Occurrence Handle1:CAS:528:DyaK2sXmslKmsA%3D%3D

    PubMed  CAS  Google Scholar 

  • H. Vandenburgh M. Del Tatto J. Shansky J. Lemaire A. Chang F. Payumo P. Lee A. Goodyear L. Raven (1996) ArticleTitleTissue-engineered skeletal muscle organoids for reversible gene therapy Hum. Gene Ther. 7 2195–2200 Occurrence Handle8934233 Occurrence Handle1:STN:280:ByiD1cfivFE%3D

    PubMed  CAS  Google Scholar 

  • Y.N. Wang C.V. Bouten D.A. Lee D.L. Bader (2004) ArticleTitleCompression induced damage in a muscle cell model “in vitro’ Proc. Inst. Mech. Eng. [H] 219 1–12

    Google Scholar 

  • R. Wronski N. Golob E. Grygar M. Windisch (2002) ArticleTitleTwo-color, fluorescence-based microplate assay for apoptosis detection BioTechniques 32 666–668 Occurrence Handle11911668 Occurrence Handle1:CAS:528:DC%2BD38XitFCntr0%3D

    PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Biomedical Engineering, Eindhoven University of Technology, WH 4.105, P.O. Box 513, 5600 MB, Eindhoven, The Netherlands

    Debby Gawlitta, Cees W. J. Oomens, Frank P. T. Baaijens & Carlijn V. C. Bouten

Authors
  1. Debby Gawlitta
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Cees W. J. Oomens
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Frank P. T. Baaijens
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Carlijn V. C. Bouten
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Debby Gawlitta.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Gawlitta, D., Oomens, C.W.J., Baaijens, F.P.T. et al. Evaluation of a Continuous Quantification Method of Apoptosis and Necrosis in Tissue Cultures. Cytotechnology 46, 139–150 (2004). https://doi.org/10.1007/s10616-005-2551-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10616-005-2551-7

Keywords

Search

Navigation