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中洪博元,细胞保种
z-bo
2020-07-20
725
Zhonghong Boyuan has been cultivating cells for many years, and you must have the most caring moment than when the new cells first arrived, and you know little about them and don't know how to deal with them. As a professional organization that has carried out cell culture for many years, it often encounters the problem of maintaining seeds after the research team purchases new cells, so today I will share with you the experience of handling new cells.
First of all, there are generally two types of transportation for cell procurement, one is to send frozen cells directly, and the other is to send live cells after resuscitation. The former is to take out the cell cryopreservation tube from liquid nitrogen and transport it on dry ice; the latter is to resuscitate the cells into a T-25 culture flask and transport it at room temperature. Both have their own advantages and disadvantages. The advantage of the first method is that it can be transported over long distances. Because the cells are relatively stable in dry ice, it can generally ensure that the cell viability is not affected within a few weeks. However, the disadvantage is that the transportation cost is high and the cell recovery is very difficult. A complicated process. If the cells are in poor condition after resuscitation, it is difficult to determine whether it is the source of the problem or the resuscitation operation. The advantage of the second method is that after the cells are received, they can be directly observed to roughly understand the cell state, and The transportation requirements are relatively low, but the disadvantage is that it is only suitable for short-distance transportation, because even if serum is not added to the medium during transportation, the cells will continue to proliferate. When the cells are full, the state of the cells will continue to deteriorate with time. . In summary, if you purchase from a large foreign cell bank, the first method is generally adopted, which can adapt to long-distance transportation, and the quality of cryopreserved cells in large cell banks is generally good, and the recovery and survival rate is high under good operation; if When purchasing from a domestic cell bank, the second method is generally used to facilitate the better grasp of the cell status when receiving the cells and reduce the transportation cost (i.e. purchase cost). Most of our cells are purchased from domestic cell institutes (such as Shanghai Institute of Cell Research, Chinese Academy of Sciences, etc.). Firstly, the quality is guaranteed, and secondly, it can reduce the time and cost of procurement. Of course, some major issues are to purchase cells from foreign cell banks. The results made in this way are more recognized by the international academic community.
is a bit off topic, so let's talk about the processing of new cells for the two modes of transportation.
In the case of sending cryopreserved cells directly, the first thing to check is whether there is enough dry ice remaining after the cells are received. If there is insufficient dry ice or there is no dry ice, the cell status may be affected. It is recommended to recover as soon as possible; if If there is sufficient dry ice, it is recommended to transfer to -80℃ refrigerator overnight, and then recover according to the normal procedure on the second day, that is, quickly melt in a 37℃ water bath. After low speed centrifugation, remove the cryopreservation solution containing DMSO and replace with fresh cells suitable for cells. The medium is gently blown and transferred to a petri dish to make up the medium; generally, adherent cells can be observed within 24 hours to determine the cell status, and the cell status can be observed after the suspension cells are recovered. For cells that have been cryopreserved for a long time, it may take 1 or 2 passages before the cells can adjust to their normal state. Therefore, if the cells are not in good condition after resuscitation, don't be discouraged, but culture them carefully.
For the case of sending live cells after resuscitation, check whether the cell bottle is damaged after receiving the cells. Generally, the express delivery of cells in China is mixed, and the cell bottle is likely to be collided and damaged during transportation. Squeeze, causing damage to the bottle. If the cell flask is found to be damaged, it needs to be disposed of as soon as possible. If the cell bottle is intact, disinfect the bottle with alcohol, remove the sealing film, and then let it stand at room temperature for 1-2 hours. After the cell is stable, observe the cell status and record it in time. If the cells are contaminated when received, they should be photographed in time. If the cells sent by the public cell bank are contaminated, they will be sent again after they confirm. If the cells are not abnormal, passage them as soon as possible and replace with fresh medium suitable for the cells. Since new cells are generally contacted for the first time, it is especially necessary to pay attention to the trypsin digestion time during cell passage. Too short or too long digestion time will affect the cell state and even lead to cell death. Zhonghong Boyuan recommends that everyone use low-concentration trypsin digestion, and shorten the interval between placing in the incubator and observation during digestion, in order to find a suitable digestion time as much as possible.