Member since: 2016
Organization: University Of Miami
Great result for cell cycle analysis!
Application Area: Cell cycle by flow cytometry
"A very easy and efficient way of cell cycle staining."
Live cell studies of cellular DNA content and cell cycle distribution are useful to detect variations of growth patterns due to a variety of physical, chemical, or biological means, to monitor apoptosis, and to study tumor behavior and suppressor gene mechanisms. In a given population, cells are distributed among three major phases of cell cycle: G0/G1 phase (one st of paired chromosomes per cell), S phase (DNA synthesis with variable amount of DNA), and G2/M phase (two sets of paired chromosomes per cell, prior to cell division). DNA content can be measured using fluorescent, DNA-selective stains that exhibit emission signals proportional to DNA mass. Flow cytometric analysis of these stained populations is then used to produce a frequency histogram that reveals the various cell cycle phases. This analysis is typically performed on permeabilized or fixed cells using a cell-impermeant nucleic acid stain, but is also possible using live cells and a cell-permeant nucleic acid stain. While the choices for fixed cell staining are varied, there are only a few examples of useful cell-permeant nucleic acid stains.
The Vybrant® DyeCycle™ Green and Orange stains are DNA-selective, cell membrane-permeant, and nonfluorescent stains for DNA content analysis in living cells. The Vybrant® DyeCycle™ Green and Orange stains are fluorescent upon binding to double-stranded DNA. These stains take advantage of the commonly available 488 nm excitation source, placing cell cycle studies on live cells within reach of all flow cytometrists. Vybrant® DyeCycle™ Green stain is excited at 488 nm with emission ~520 nm. Vybrant® DyeCycle™ Orange stain is excited using both 488 nm and 532 nm laser lines with emission ~570 nm.
The staining protocol is simple and includes incubating suspended cells in the presence of Vybrant® DyeCycle™ stain and directly measuring the fluorescence without the need for any additional treatment or centrifugation steps. This live cell stain allows the simultaneous co-staining of the cell population for other parameters, and allows for the possibility of cell sorting based on DNA content.