Liquid nitrogen ultra-low temperature freezing method of microbial species preservation method.

Biotechnology Channel News: Liquid nitrogen ultra-low temperature preservation technology is to preserve the bacteria species in -196 degrees C of liquid nitrogen, or -150 degrees C of nitrogen in the long-term preservation method, its principle is to use microorganisms in -130 degrees C metabolism tend to stop and effectively preserve microorganisms.
procedures are as follows: 1, the preparation of ampoule tube or freeze storage tube with round bottom boron silicon glass products ampoule tube, or the plastic freeze storage tube at the helix port.
note that the glass tube must not have cracks.
cleaning the freezer or ampoule tube, autocultension at 121 degrees C for 15-20 minutes, spare.
2, the preparation of protective agents the type of protective agents to be selected according to the category of microorganisms.
preparation of protective agents, attention should be paid to its concentration, generally using 10-20% glycelycer.
3, microbial preservation of different physiological states of microorganisms have an impact on survival rate, generally using static or mature culture.
should be operated under sterile conditions when packing. The preparation of
strains can be prepared by the following methods: scraping the spores or bacteria on the slope of the culture, mixing with the protective agent and adding them to the freeze storage tube; The culture is cultured in flat dishes, the formation of bacteria backward, with sterile punchers from the plate to cut some uniform size of small pieces (diameter of about 5-10 mm), fungi preferably take the bacteria at the edge of the bacteria, mixed with the protective agent and added to the freeze storage tube;
4, prefreeze when the general freezing speed is controlled at a rate of 1 degree C per minute, so that the sample freezes to -35 degrees C.
there are three commonly used temperature control methods: program temperature control cooling method, the application of electronic computer programs to control cooling devices, can stabilize continuous cooling, can control the cooling rate very well.
cooling method: cool the bacteria in different temperature levels of refrigerator or liquid nitrogen tank section cooling, or hanging in the air mist of ice gradually cool down.
generally using two-step temperature control, the ampoule tube or plastic tube, first put -20 degrees C to -40 degrees C refrigerator for 1-2 hours, and then remove and put into the liquid nitrogen tank for rapid freezing.
the freezing rate is reduced by about 1-1.5 degrees Celsius per minute.
can be placed directly into the gas phase or liquid phase nitrogen for low temperature-resistant microorganisms.
5, preservation The ampoule tube or plastic freezer tube is placed in the liquid nitrogen tank to be stored.
temperature in the general gas phase is -150 degrees C and the temperature in the liquid phase is -196 degrees C.
6, resuscitation method Remove the ampoule tube or plastic freezer tube from the liquid nitrogen tank and immediately place it in a water bath at 38 degrees C-40 degrees C to recover quickly and shake appropriately.
until all the internal ice dissolves, it usually takes about 50 to 100 seconds.
the ampoule tube or plastic freezer tube and move the contents to a suitable culture base for culture.
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