Q1: What color should Serum be?
A1: Color of serum is affected by many factors. The colors of serum from different batches are likely to have a great difference with each other. The color may be yellow, red, amber or interposed between the three. And, the color does not have direct relation with the quality of serum products.
Q2: How to store the serum?
A2: Serum should be stored at the temperature range from -15 ℃ to -25 ℃, as well as at -70 ℃. Because in most cases, a bottle of 500ml of serum will not be used all at once, it is recommended to dispense serum into disposable volume, which can be used up in once experiment (usually 40ml). We do not recommend keeping it at 4 ℃ for more than a month.
Q3: How to defrost serum?
A3: Bottle (500ml) serum thawing steps (gradual thawing method): remove it from -20 ℃ or -70℃ refrigerator to 4 ℃ refrigerator to thaw for one day. Subpackage at room temperature after the serum thawing out. In the process of thawing, shake the bottle regularly (be careful not to generate air bubbles), to make the temperature and composition uniformity and reduce the occurrence of precipitation. Do not thaw the serum directly from the -20 ℃ to 37 ℃, because the temperature change is too large and easily lead to protein coagulation and precipitation.
Q4: After thawing serum, there are floccules found. How to deal with this situation?
A4: There are several reasons for floccules in serum; one of the most common reasons is denaturation of lipoprotein in serum. Apart from that, fibrin (one kind of clotting protein) is also one of the main reason causing floccules in serum after thawing. These floccules do not affect the quality and use of serum. If the pipette can avoid the floccules, you can totally ignore them and directly add serum to the culture medium. If you want to remove the floccules, it is recommended to dispense the serum into sterile centrifuge tubes and briefly centrifuge them with 400g centrifugal force. Then use the supernatant after centrifugation. We do not recommend filter, because floccules will clog filters.
Q5: Why will FBS stored in the refrigerator precipitate?
Q5: When FBS stored at 2-8 ℃ without pre-aging, various serum proteins (such as cold agglutinin, fibrinogen, fibronectin, etc.) and lipoproteins may aggregate to form the visible precipitate or turbidity. These deposits will not affect the serum’s using effect.
Q6: How to avoid the serum precipitation?
A6: When using the serum, we recommend paying attention to the following:
a) When thawing serum, please follow the gradual thawing method above. We do not recommend you directly removing serum from -20 ℃ to 37 ℃ or removing -70 ℃ into a water bath to thaw. Dramatic changes in temperature can lead to sediment and black spots.
b) When thawing serum, please gently shake the bottle, so that the composition and temperature will be uniform. In this way, you can reduce the precipitation caused by temperature variations.
c) Do not keep serum at 37 ℃ for too long. If so, the serum will become turbid, while many of the labile components in serum will be metamorphic, and influence the quality of serum.
d) Heat-inactivation is very likely to cause increases of sediment in serum. If not necessary, do not need to do this step.
e) If you have to do heat-inactivation, please observe the principle that keeping at fifty six degree centigrade for less than thirty minutes, and shake the bottle regularly. The factors-too high temperature, heating too long or shake unevenly, will all result in an increase in sediment.
Q7: Why do people heat-inactivate serum?
A7: The heating can inactivate the complement system. Activated complement will participate in cell lysis, stimulate smooth muscle contraction, and stimulate many cells and blood platelets to release histamine, activate the lymphocytes and macrophages. In the experiment of immunology research, and culturing embryonic stem cells, insect cells, smooth muscle cells, it is recommended to use heat-inactivated serum.
Q8: Is it necessary to heat-inactivate serum?
A8: Experiments show that it is not necessary for the majority of cells to use to heat-inactivated serum. Heat-inactivated serum can only slightly promote the growth of cells, or just has no effect. However, sometimes heat-inactivated serum may even have negative affect on the quality of serum because of the high temperature treatment, and resulting in cell growth rate decreased. After heat treatment of serum, precipitates have a significant increase. These deposits can be observed by microscope. They look like "black spot". Some researchers will mistakenly believe that the serum is contaminated, while the serum, long time keeping in 37 ℃ environment, will show an increase of the precipitates, which looks like the clonal expansion of microorganisms.
Q9: How to correctly inactivate serum?
A9: When heat-inactivating serum, please place serum in 56 ℃ water bath for 30 minutes. To minimize the impact of heat-inactivation on serum’s quality, you should control the serum volume not too large. It is better to prepare the same container of water with equal volume as serum (the same temperature as serum). Place The vessel containing serum and water at the same time at 56 ℃ water bath. thermometer place in the container filled with water, and begin counting time when the thermometer reached about 56℃. During the heating process, gently rotate the serum from time to time.
Q10: Specks were observed in serum sometimes. Are they bacteria?
A10: Some laboratory personnel observe black specks in serum-containing culture system and think they are bacteria. In fact, these black substances may be salt or serum protein crystallization. The motion of the matters in the culture system is Brownian motion, which is the result of liquid movement instead of the independent organism movement. And never had any biological laboratory once isolated genetic material from the black substance. With the increase of time to keep the system at 37 ℃, the formation of precipitation will increase. Correct thawing and inactivation (without heat inactivation if not necessary) can reduce the generation of black spots.
Q11: Serum partially melted during transportation, whether it can still be usable.
A11: During the transportation, the serum will undergo several freeze-thaw processes, and will not affect serum quality in a way. Partially thawed serum can be normally used. But we should reduce the amount of freezing and thawing process as far as possible.