Cancer Chemotherapy and Pharmacology

, Volume 27, Issue 6, pp 445–450

Detection of multidrug resistance and quantification of responses of human tumour cells to cytotoxic agents using flow cytometric spectral shift analysis of Hoechst 33342-DNA fluorescence

  • Paul J. Smith
  • Sally A. Morgan
  • James V. Watson
Original Articles Multidrug Resistance, Human Tumour Cells, Hoechst 33342


We describe the application of a flow cytometric technique for assessing the radiation or drug sensitivity characteristics of human tumour cells. The technique makes use of the phenomenon that a red shift occurs in the fluorescence emission spectrum of a DNA-specific dye (Hoechst 33342) as an increasing number of dye molecules bind to nuclear DNA. Intact, viable cells undergo a time-dependent spectral shift that can be distinguished from the rapid shift observed in cells with damaged membranes by the use of multiparametric flow cytometry. The responses of various human cell lines were compared, namely, those of normal and ataxia-telangiectasia (A-T) lymphoblastoid lines, a small-cell lung carcinoma line and its (in vitro) derived multidrug-resistant variants. A close correlation was found between dye toxicity and the degree of DNA binding of Hoechst 33342 independent of cellular DNA content, with lymphoblastoid and multidrug-resistant small-cell lung cancer cells showing enhanced and restricted dye-binding rates, respectively. VP16-and radiation-induced cell kill was found to result in a quantifiable increase in the fraction of cells undergoing a rapid spectral shift and was capable of detecting the increased radiation sensitivity of A-T-derived cells. Spectral shift analysis provides a rapid method for assessing the responses of tumour cells to cytotoxic agents and for determining the general ability of cells to protect cellular DNA from a model DNA-binding agent (Hoechst 33342) that participates in the multidrug resistance phenotype.


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Copyright information

© Springer-Verlag 1991

Authors and Affiliations

  • Paul J. Smith
    • 1
  • Sally A. Morgan
    • 1
  • James V. Watson
    • 1
  1. 1.Medical Research Council, Clinical Oncology and Radiotherapeutics UnitMRC CentreCambridgeUK

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