Cell name | Bovine endometrial epithelial cells BEND |
CELLBIO Item No. | CBR-131460 |
Animal species | Cattle |
Number of cells | 1X10^6 |
Culture bottle specification | 25T |
Passage | 2-3 generations |
Stable pass times | 10-15 generation |
Passage | 1:2 or 1:3 passage |
Recommended training program | 90% Hyclone DMEM high sugar |
Medium; 10% Gibco fetal calf | |
Serum, 1%-1.5% Cellbio Double | |
anti- | |
Culture temperature | 37 ° C |
Carbon dioxide concentration | 5% |
Cell purity | ≥ 99% |
Mycoplasma detection | (-) |
Toxin detection in culture system | ≤ 0.5EU/ml |
Cell handling considerations
1. Receive cells, observe the cells under a microscope. Due to transportation
The problem is that the adherent cells in the cell culture bottle may fall off from the bottle wall, and the cell suspension may occur under the microscope. When this state occurs, please do not open the cell culture bottle, and immediately place the culture bottle in the cell. Leave the incubator in the incubator for 3-5 hours, let the cells stabilize first, and then observe under the microscope. At this time, most of the cells will reattach to the bottle wall. If the cells still do not adhere to the wall, please use the trypan blue staining method to identify the cell viability. If the trypan blue staining confirms that the cell viability is normal, please treat it by suspending cells.
2. After receiving the cells, observe the cells under the microscope and treat the cells in an appropriate manner. If there are more cells in suspension, collect the cells by centrifugation and inoculate them into a new culture flask. Discard the stock solution, use freshly prepared medium, and use imported fetal bovine serum. Just after receiving the cells, if the cells are not much, the serum concentration can be added to 15% for culture. If the cell is about 80%, the serum concentration is still 10%.
3. If there is no abnormality when receiving the cells, please observe the cell density under the microscope. If it is adherent cells, if the concentration is not more than 80%, the medium in the culture flask will be aspirated, leaving 5-10ML medium to continue the culture. : When more than 80% confluence, please subculture according to cell culture conditions. For suspension cells, aspirate the culture solution, centrifuge at 1000 rpm for 3 minutes, aspirate the supernatant, and drain the cells in the fresh medium to return to the flask.
After 4 hours, the cell morphology has been restored and covered with the bottle wall, which can be passaged. (Adherent cells) Pour the medium in the flask and add 3-5 ml (according to the cell growth surface) to wash with PBS or Hanks' solution and discard. plus
0.5-1ml 0.25% trypsin digestion with EDTA, the digestion time is based on the specific cells, usually 1-3 minutes, no more than 5 minutes. It can be digested in a 37 ° C incubator. Gently shake the bottle wall, see the cells fall off, add 3-5ml
The medium is terminated for digestion. The cells on the wall of the bottle were gently pipetted with a pipette to completely fall off, and then the solution was aspirated into a centrifuge tube and centrifuged at 1000 rpm for 5 min.
Discard the supernatant, depending on the number of cells, determine the number of bottles, generally one pass two, such as the amount of cells can be passed three, some cells are not easy to pass too thin, some cells that grow faster can pass a few bottles to specific cells And experience prevails. (suspended cells) Use a pipette to gently blow the bottle wall and directly inject the solution into the centrifuge tube for centrifugation.
5, adherent cells, suspension cells. Strict aseptic operation. When changing the solution, renew the cell culture flask and change to the fresh culture medium.
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