Chapter 3. Flow Cytometry

  1. Mary Hannon-Fletcher Lecturer2 and
  2. Perry Maxwell Principal Clinical Scientist3
  1. Ian Dimmick Flow Cytometry Core Facility Manager

Published Online: 11 MAY 2009

DOI: 10.1002/9780470745069.ch3

Advanced Techniques in Diagnostic Cellular Pathology

Advanced Techniques in Diagnostic Cellular Pathology

How to Cite

Dimmick, I. (2009) Flow Cytometry, in Advanced Techniques in Diagnostic Cellular Pathology (eds M. Hannon-Fletcher and P. Maxwell), John Wiley & Sons, Ltd, Chichester, UK. doi: 10.1002/9780470745069.ch3

Editor Information

  1. 2

    The University of Ulster, Coleraine, UK

  2. 3

    Belfast Health & Social Care Trust, Centre for Cancer Research & Cell Biology, Belfast, UK

Author Information

  1. Institute of Human Genetics, Bioscience Centre, International Centre for Life, Newcastle Upon Tyne, UK

Publication History

  1. Published Online: 11 MAY 2009
  2. Published Print: 17 APR 2009

ISBN Information

Print ISBN: 9780470515976

Online ISBN: 9780470745069

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Keywords:

  • flow cytometry - semi-automated procedure for interrogation of single cells in continuous fluid stream;
  • cell surface and cytoplasmic measurements – cellular antigen expression;
  • flow cytometer principles;
  • photomultiplier tube (PMT);
  • excitation and emission spectra;
  • defining what is negative - important step in flow cytometry with isotype control;
  • flow cytometry - acting as invaluable aid in diagnosis of haematological malignancies;
  • acute myeloid leukaemia (AML);
  • future for flow cytometry and Amnis ImageStream system;
  • fluorescent in situ hybridization (FISH) spot counting

Summary

Flow Cytometry continues to be a rapidly expanding field of science. The demands put upon this branch of science are generated primarily by the requirement of more data created faster, under more complex conditions, such as multicolour immunophenotyping involving now quite routinely six and more colours. To keep up with these requirements a large number of new fluorochromes have been created as more instruments are being developed and sold with more lasers. Unique fluorphores are being developed to utilise the use of these new lasers and expand the ability to do multicolour Flow Cytometry. In this chapter, we look at how good Flow Cytometry still relies upon the basics of sample preparation, antibody-fluorochrome choice and the understanding of spectral overlap within experiments that involve two or more fluorochromes. The basic principles of the hardware that make up all flow cytometers are very similar. The end user however, has to be aware of the small differences from each instrument to optimise and work within each individual instruments performance to allow the generation of relevant and good data; this requires not only an understanding of the instrumentation but also the cells and cellular components under analysis. Finally, the future of Flow Cytometry lies in instruments where the operator gets both graphical data distribution of cells and cell probes and an option to visualise the actual cell staining on a cell-by-cell basis by using state of the art camera technology.