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Clinical & Experimental Metastasis

, Volume 24, Issue 5, pp 317–327 | Cite as

Specific isolation of disseminated cancer cells: a new method permitting sensitive detection of target molecules of diagnostic and therapeutic value

  • Siri Tveito
  • Gunhild M. Maelandsmo
  • Hanne K. Hoifodt
  • Heidi Rasmussen
  • Oystein Fodstad
Original Paper

Abstract

Molecular studies of rare cells, such as circulating cancer cells, require efficient pre-enrichment steps to obtain a pure population of target cells for further characterization. We have developed a two-step approach, starting with immunomagnetic enrichment, followed by specific isolation of individual, easily identifiable bead-rosetted target cells using a new semi-automated CellPick system. With this procedure, 1–50 live target cells can now be isolated. As a model system, we spiked a small number of tumor cells into millions of normal mononuclear cells (MNCs). Efficient isolation of pure target cells was obtained by use of the CellPick system, and the nature of isolated, bead-rosetted cells was verified by use of FISH. Single breast cancer cells were picked directly into an RNA preserving lysis buffer, reverse transcribed, and PCR amplified with two cDNA specific primer sets. With the isolated cells we consistently obtained both ubiquitously expressed and tumor cell specific PCR products. We also performed a successful mutation analysis of single cells using PCR and cycling temperature capillary electrophoresis (CTCE). This may have significant clinical implications in cancer and in other diseases, e.g. in characterizing micrometastatic cancer cells in blood and lymph nodes to help identifying patients who most likely will respond to therapies like tyrosine kinase inhibitors and compounds targeting specific mutations. By use of the CellPick system it is possible to specifically isolate bead-rosetted or otherwise labelled target cells from a heterogeneous cell population for further molecular characterization.

Keywords

Micrometastasis Circulating cells Single cells Pure cell population RT-PCR Mutation-detection 

Notes

Acknowledgements

We would like to thank Stine Kresse for excellent tutorials in the FISH procedure and Karen Marie Heintz for performing all the CTCE analyses. This work was supported by the Norwegian Cancer Society, the Research Council of Norway and the Anna Lovise Lundeby Memorial Fund.

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Copyright information

© Springer Science + Business Media B.V. 2007

Authors and Affiliations

  • Siri Tveito
    • 1
  • Gunhild M. Maelandsmo
    • 1
  • Hanne K. Hoifodt
    • 1
  • Heidi Rasmussen
    • 1
  • Oystein Fodstad
    • 1
    • 2
    • 3
  1. 1.Department of Tumor BiologyRikshospitalet-Radiumhospitalet Medical CenterOsloNorway
  2. 2.Faculty of MedicineUniversity of OsloOsloNorway
  3. 3.University of South Alabama-Mitchell Cancer InstituteMobileUSA

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