Cell cycle experiment outsourcing test
Time：2018-10-02 Click：1150
Cell cycle experimental outsourcing detection is one of the common experiments of the cell experiment platform, and is also one of the synthesis of the swantopure project, which is operated by experienced experiments.
The reagents used in the experiment of the company are all imported reagents, preparatory preparations.
The principle of cell cycle detection:
PI method is a classic cycle detection method, PI is propididium, a double-stranded DNA Fluorescent dyes, iodide and double stranded DNA can produce fluorescence, and the content of fluorescence intensity and double stranded DNA is proportional. After DNA in the cells were stained with propidium iodide, the cells can be measured by flow cytometry, and then cell cycle and apoptosis analysis can be performed according to the distribution of DNA content.
Generally, the G0 / G1 of normal cells has a DNA content (2n) of diploid cells (2n), and the G2 / M period has a DNA content (4n) of tetraploid cells, while the S phase DNA
The The content is between 2 N and 4N. After the cells were fixed to the cells, the PI can bind to the DNA of the cell, and the fluorescence intensity directly reflects the intracellular DNA content.
Therefore, the cell cycle is divided into G1 / G0 period, S-phase and G2 / M phase, and can be obtained by detection of the cell cycle in the stream intracellular DNA. Cell cycle testing experimental process:
1. Collect cells: A: Collect cells 5 ~ 20 u0026 Times; 105 in centrifugal tubes, if the cells are relatively small ( Such as lymphocytes) 400g centrifugation, if the cells are large (such as tumor cells) 300g centrifugation, 5 ~ 10min, discard the culture solution;
2. Wash: Add 1 ml cold PBS (pre-cooling in advance 4 ° C) Wash 1 time, leave the supernatant;
3. Fixed pre-treatment: Use the liquid left by the tubular floor, the bottom of the tube, and the precipitate is repeated into a cell suspension to avoid the cells; [123 ]
4. Cellular fixation: Add about 1 mL-20 ° C pre-cold anhydrous ethanol, gently blow mixing, fixed 1 h at -20 ° C or overnight; 7.pi Dyeing: Add 400 u0026 Mu; L of the PI solution and mix well, 4EtOAc (EtOAc) EtOAc.
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