1 Equipment and materials
1.1 Low temperature refrigerator: -70 ° C
1.2 Autoclave.
1.3 Porous glass beads (or plain porcelain beads): 2~3mm in diameter.
1.4 Commercial cell cryotubes (optional).
2 Operation steps
2.1 The porcelain beads with a diameter of 2-3 mm are divided into small screw cap bottles (up to 30 capsules per bottle) to make a cryotube and autoclave.
2.2 Mark the cryotube.
2.3 Open the cryostat under aseptic conditions
2.4 The cells are cultured to the stable or mature stage under the optimum nutrient conditions of the deposited strains. After the purity check, the fresh culture of the strain is added to the medium to prepare a suspension, and the suspension is aseptically operated. Transfer to the freezing tube.
2.5 Seal the cryotube and invert the suspension of the emulsified culture medium four to five times. Do not vortex.
2.6 After the strain is adsorbed on the porcelain beads, the excess suspension is aspirated.
2.7 Commercial bacterial cryotubes can also be used, in which case they should be operated in accordance with the product instructions.
3 preservation
The cryotubes were stored at -70 °C.
4 preservation period
Can be stored for 10 years.
5 recovery training
5.1 Commonly used medium See Appendix G. Open the cryotube under aseptic conditions, take out a porcelain bead with a sterile or inoculation ring, seal the cryotube and return it to the low temperature condition as soon as possible.
5.2 Strains oh enamel beads can be directly inoculated in solid medium or an appropriate amount of liquid medium.
6 Precautions
Considerations include:
a In order to keep the strains stable for a long time, pay attention to aseptic operation during the use of the cryotubes.
b Perform operations such as strain preservation and resuscitation in the biosafety cabinet.
c Pay attention to biosafety when handling applied cryotubes.
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