Analysis of Cellular DNA Content by Flow Cytometry
- Curr Protoc Cytom. 2017 Oct 2:82:7.5.1-7.5.20. doi: 10.1002/cpcy.28.
- 1. Department of Pathology and Brander Cancer Research Institute, New York Medical College, Valhalla, New York.
- 2. Hematology/Oncology, Case Western Reserve University Cleveland, Ohio.
Cellular DNA content can be measured by flow cytometry with the aim of : (1) revealing cell distribution within the major phases of the cell cycle, (2) estimating frequency of apoptotic cells with fractional DNA content, and/or (3) disclosing DNA ploidy of the measured cell population. In this unit, simple and universally applicable methods for staining fixed cells are presented, as are methods that utilize detergents and/or proteolytic treatment to permeabilize cells and make DNA accessible to fluorochrome. Additionally, supravital cell staining with Hoechst 33342, which is primarily used for sorting live cells based on DNA-content differences for their subsequent culturing, is described. Also presented are methods for staining cell nuclei isolated from paraffin-embedded tissues. Available algorithms are listed for deconvolution of DNA-content-frequency histograms to estimate percentage of cells in major phases of the cell cycle and frequency of apoptotic cells with fractional DNA content. © 2017 by John Wiley & Sons, Inc.