Zui recently used fetal bovine serum in cultured cells. Because of the precipitation, I don’t know whether it has deteriorated. I didn’t get the exact answer when I consulted the relevant literature. Therefore, I dare not use the original serum again, resulting in waste. I remind everyone to do cell culture. When you pay attention to the dispensing of serum.
Serum is one of the important elements of Zui in cell culture. Pay attention to and deal with in laboratory operations:
1. Serum preservation: The recommended serum should be stored at -5 ° C to -2 ° ° C. However, if stored at 4 ° C, do not exceed one month. If you are unable to use one bottle at a time, it is recommended that you aliquot the serum into a suitable sterile container and put it back in the freezer.
2. Method of thawing serum: It is recommended that you take the serum out of the freezer, put it in a refrigerator at 2-8 °C to melt it, and then let it melt at room temperature. However, it must be noted that the melting process must be regularly shaken evenly.
3. After the serum is thawed, flocculent deposits are found. What should I do?
There are many reasons for the presence of precipitates in serum, but the common cause of Zui is due to the degeneration of lipoproteins in serum, and fibrin (one of the proteins that form blood coagulation) is also present in serum after serum thawing. It is also one of the main causes of sedimentation. However, these flocculent precipitates do not affect the quality of the serum itself.
If you want to remove these flocculent deposits, the serum can be dispensed into a sterile centrifuge tube and centrifuged slightly at 400 g. The supernatant can then be added to the medium for filtration. We do not recommend that you remove these flocculent deposits by filtration as it may block your filter membrane.
4. Why do you want to heat inactivate serum?
Heating can inactivate the complement system. Activated complement is involved in lytic cell events, stimulates smooth muscle contraction, releases histamine from cells and platelets, and activates lymphocytes and macrophages. In immunological studies, the use of heat-inactivated serum is recommended when culturing ES cells, insect cells and smooth muscle cells.
5. Is it necessary to do heat inactivation?
Experiments have shown that properly treated heat inactivated serum is not required for most cells. The serum thus treated has only a slight promotion to the growth of the cells, or has no effect at all, and even the high temperature treatment usually affects the quality of the serum, resulting in a decrease in the cell growth rate. The heat-treated serum, the formation of precipitates will increase significantly, these precipitates observed under an inverted microscope, such as "small black spots", often let researchers mistakenly believe that serum is contaminated, and put serum in 37 In the °C environment, this precipitate will be increased, which makes the researchers mistakenly believe that it is the splitting and amplification of microorganisms.
Therefore we recommend that you do not need to do the heat treatment if it is not necessary. In this way, not only save your precious time, but also ensure the quality of serum!
6. Why does the fetal bovine serum stored in the refrigerator precipitate?
Fetal bovine serum is not pre-aged. When stored at 2-8 ° C, various proteins and lipoproteins (such as cold agglutinin, fibrinogen, villogen, etc.) in the serum may aggregate to form a precipitate or visible turbidity. This should not affect the quality of the serum. It is recommended to store fetal bovine serum at -20 °C to avoid repeated freezing and thawing.
7. How to avoid the formation of sediment?
When thawing serum, follow the recommended step-by-step thawing method (-20 ° C to 4 ° C to room temperature). If the temperature is too large when the serum is thawed (eg -20 ° C to 37 ° C), the experiment shows that precipitation is very likely to occur.
When thawing serum, please shake it at any time to make the temperature and composition uniform, reducing the occurrence of sedimentation.
Do not leave the serum at 37 °C for too long. If left at 37 ° C for too long, the serum will become cloudy, and many of the more unstable components in the serum will be damaged, which will affect the quality of the serum.
The heat inactivation of serum is very likely to cause an increase in sediment, which may not be necessary if not necessary.
If you must do the heat inactivation of serum, please observe the principle of 56 ° C, 30 minutes, and shake it at any time. If the temperature is too high, the time is too long or the shaking is uneven, it will cause the precipitation to increase.
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