Cell cycle analysis by quantitation of DNA content was one of the earliest applications of flow cytometry. The DNA of mammalian, yeast, plant or bacterial cells. Cell cycle analysis by DNA content measurement is a method that most frequently employs flow cytometry to distinguish cells in different phases of the cell cycle. Cell cycle analysis was one of the earliest applications developed on the LSC platform and continues to be one of the most important ones. As in FC, the total.
cycle analysis Cell
The basic version can perform a single cell cycle analysis without background, aggregate or debris correction. To analyze aneuploid populations or background and debris fitting, you can purchase the complete Multicycle package.
The DNA fitting engine in Multicycle has been used for over a decade by researchers all over the world and is described in several publications: DNA content histogram and cell-cycle analysis. Methods in Cell Biology. Practical considerations for DNA content and cell cycle analysis. Principles and Applications Bauer, K.
Williams and Wilkins, Baltimore, pp. Cell Growth and Division, 2nd Edition. Oxford University Press, Oxford, Automated peak detection and cell cycle analysis of flow cytometric histograms.
Multiple levels of automation, including manual, semi automated auto detect number of cycles and full automation autofit modes. Jurkat cells were incubated with media only control , nocodazole 0.
Control and drug-treated cells were fixed then stained with propidium iodide. The fluorescence intensity for each cell was measured.
The Cellometer instrument acquires a bright field image for each sample tested. The bright field image allows researchers to verify cell morphology, evaluate the degree of homogeneity of the sample, and identify the presence of cellular debris.
Because all of the cells have been fixed, all of the cells are stained with propidium iodide and appear in the fluorescent image. The fluorescent counted image can be used to confirm that cells are counted correctly.
Individual counted cells are outlined in green. Uncounted cells are outlined in yellow. Cellometer software uses proprietary algorithms to accurately count individual cells within clumps. A cell cycle histogram is automatically generated for each sample using the optimized Nexcelom cell cycle data layout in FCS Express Flow Software.
Gating can be manually optimized directly on the histogram with automatic update to the associated data table. Histograms and data tables for control and experimental samples are shown in the experimental data section below. Gating was set for the control sample and applied to histograms for the two experimental conditions presented. Population histogram for the control sample generated with FCS Express software. Figures 2 and 3.
Population histogram for the 0. Sub-G 1 population is indicated in red. Population histograms for control, 0. Data for each phase of the cell cycle is show in the table on right. As the concentration of the nocodazole increased from 0. Similar correlation data for etoposide is presented in the Nexcelom publication referenced below.
There are a number of fluorescent-based dyes that are capable of binding to double stranded DNA. Since the amount of bound fluorescent dye is directly proportional to the amount of DNA present within a cell, these dyes can be used to detect the cell cycle within a population of cells.
Below, is a representative example of cell cycle detection using DAPI. In this experiment, control and drug treated cells were stained with DAPI and analyzed using a Cellometer. Cells that were treated with 0. Type I interferons induce autophagy in certain human cancer cell lines. Cells are preparing for DNA replication. At this stage the cells contain about the same amount of DNA.
The diploid number of a cell is the number of chromosomes in the cell. This number is commonly abbreviated as 2n, where n stands for the number of chromosomes.
Since there are normally 23 pairs of chromosomes in human cells:
Cell Cycle Analysis by Flow: DNA Stains and Beyond
Cell cycle analysis appears to be deceptively easy in concept, but details are absolutely critical. It is not possible to hide the data if there is poor. Described are four widely used procedures to analyze the cell cycle by flow cytometry. The first two are based on univariate analysis of cellular DNA content. G1 (G=Gap) phase. Cells are preparing for DNA replication. At this stage the cells contain about the same amount of DNA. The diploid number.