title: Distinguishing modes of cell death using the ImageStream multispectral imaging flow cytometer.
abstract: BACKGROUND
  Here we demonstrate the ability of the ImageStream 100 Multispectral Imaging Cytometer to discriminate between live, necrotic, and early and late apoptotic cells, using unique combinations of photometric and morphometric features.
  
  
  METHODS
  Live, necrotic, and early and late apoptotic cells were prepared and analyzed by immunofluorescence microscopy, conventional flow cytometry, and imaging flow cytometry, both as single populations and as a heterogeneous mixture of cells.
  
  
  RESULTS
  Live (annexin V(-), 7-AAD(-)) and early apoptotic (annexin V(+), 7-AAD(-)) cells were readily identifiable using either conventional or ImageStream based flow cytometric methods. However, inspection of multispectral images of cells demonstrated that the annexin V(+), 7-AAD(+) population contained both necrotic and late-stage apoptotic cells. Although these cells could not be distinguished using conventional flow cytometric techniques, they were separable using unique combinations of photometric and morphometric measures available using ImageStream technologies.
  
  
  CONCLUSIONS
  Using multispectral imagery, morphologically distinct cell populations can be distinguished using features not available with conventional flow cytometers. In particular, the ability to couple morphometric with photometric measures makes it possible to distinguish live cells from cells in the early phases of apoptosis, as well as late apoptotic cells from necrotic cells.
authors.affiliation: Amnis Corporation, Seattle, Washington 98121, USA. t.george@Amnis.com
authors.forename: Thaddeus C
authors.initials: TC
authors.lastname: George
authors.forename: David A
authors.initials: DA
authors.lastname: Basiji
authors.forename: Brian E
authors.initials: BE
authors.lastname: Hall
authors.forename: David H
authors.initials: DH
authors.lastname: Lynch
authors.forename: William E
authors.initials: WE
authors.lastname: Ortyn
authors.forename: David J
authors.initials: DJ
authors.lastname: Perry
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Seo
authors.forename: Cathleen A
authors.initials: CA
authors.lastname: Zimmerman
authors.forename: Philip J
authors.initials: PJ
authors.lastname: Morrissey
mesh_terms: D016923:Cell Death / Q000502:physiology; D002469:Cell Separation / Q000379:methods; D005434:Flow Cytometry / Q000295:instrumentation* / Q000379:methods; D005455:Fluorescent Antibody Technique; D006801:Humans; D019044:Image Cytometry / Q000379:methods*; D019169:Jurkat Cells; D008856:Microscopy, Fluorescence / Q000379:methods