FLOW CYTOMETRY

WHAT IS A FLOW CYTOMETRY?

Flow cytometry uses a technology that analyzes and quantitates the physical and chemical characteristics of particles in a fluid as it passes through at least one laser. Flow cytometers can measure cell size, cell granularity, the amounts of cell components, amounts of specific surface receptors, the amount of intracellular proteins, or transient signaling events in living cells. The fluorescence is measured to determine the properties of a single particle, quantities are usually relative, up to three to six properties or components are quantified in a single sample, cell by cell, for about 10 thousand cells, in less than a minute.

THREE MAIN SYSTEM OF A FLOW CYTOMETER

  • Fluidics System ― transports particles in a stream to the laser beam for interrogation, the section of the fluid stream that contains the particles is called the sample core. Particles that are 0.2 to 150 micrometers in size can be analyzed by the flow cytometer.
  • Optics System ― consists of lasers to illuminate the particles in the sample stream and optical filters to direct the resulting light signals to the appropriate detectors. Optical filters and beam splitters are directed to the light signals. Relevant detectors emit electronic signals proportional to the signals that hit them.
  • Electronics System ― converts the detected light signals into electronic signals that can be processed by the computer. It has the capability to initiate sorting decisions to charge and deflect particles.

The data are collected and stored in the computer, they can be analyzed to provide information, they are presented in a form of single parameter histogram or as a plot of correlated parameters, are referred to as cytograms. Cytograms may be displayed in the dot plot, contour plot or density plot.

APPLICATION OF FLOW CYTOMETRY

Flow cytometry has a relevant application to health science, it is used to diagnose conditions, for basic research and clinical trials, and vital laboratory diagnostics.

  • Cell counting
  • Cell sorting
  • Detection of biomarkers
  • Protein engineering ― to identify cell surface-displayed protein variants with desired properties
  • Molecular biology
  • Pathology
  • Immunology
  • Plant Biology
  • Marine Biology ― to characterize the abundance and community structure of photosynthetic planktons
  • Transplantation
  • Tumor immunology and chemotherapy
  • Prenatal diagnosis
  • Sex preselection ― genetics and sperm sorting
  • Detection of DNA damage, caspase cleavage, and apoptosis

Measurable Parameters

  • Apoptosis
  • Cell adherence, pigments, surface antigens, viability
  • Characterizing multidrug resistance
  • Chromosome analysis and sorting
  • Enzymatic activity
  • Protein expression and localization, modifications, phospho-proteins
  • Total DNA and RNA content
  • Intracellular antigens
  • Nuclear antigens
  • Oxidative burst
  • Membrane fluidity
  • pH, intracellular ionized calcium, magnesium, membrane potential