Cryo Tubes

Content
1. What are cryo tubes?
    1.1 Cryo tubes sizes
2. Features of cryo tubes
    2.1 Safety of cryo tubes
    2.2 Lids of cryo tubes
    2.3 Advantages of cryo tubes
3. Use of cryo tubes
4. How to buy cryo tubes?

To ensure reproducibility and continuity of results in research and biomedical processes, today's scientists are faced with the task of preserving living cells and organisms with optimal viability and ensuring that subcellular components such as nucleic acids and proteins are unchanged. Cryo tubes, also known as cryogenic storage vials, are a special type of test tube. They can be used for cryogenic storage of samples, preserving samples such as whole blood, serum, cells, and so on. They are generally applied in the biological industry, pharmaceutical industry, and food industry as a carrier for low-temperature transportation and storage of biological materials.

Not all cryo tubes can be used for liquid nitrogen, cryogenic vials for liquid nitrogen are designed to meet the demands of liquid nitrogen temperatures. They are special cryotubes with high sealing and low-temperature resistance.

There is no specific classification of cryo tube, it is generally divided according to capacity as follows:

Cryo tubes are laboratory tubes used to store samples at low temperatures. Improper use of cryotube may cause explosions, resulting in leakage of hazardous materials and, in severe cases, even injuries. To reduce the risk of cryo tube explosion, please pay attention to the following guidelines when using it.

a. Cryo tube is usually made of high-density polyethylene or polypropylene. It can be divided into two categories according to different usage scenarios. One is cryotubes for cell culture and the other is cryotubes for liquid nitrogen. These two types of cryotubes need to be carefully distinguished before purchase and use, as their applicable temperatures are particularly different.

b. All cryotube vials should have a silicone gasket, rubber gasket, or O-ring gasket to ensure a tight seal. Please note that overtightening the cap can deform the seal.

c. Cryo tubes should be stored in the gas phase above the liquid nitrogen and not immersed in the liquid to avoid penetration of the liquid nitrogen into the tubes. Overcrowding of storage containers should be avoided. If samples must be stored in the liquid phase, it is recommended to use a professional protective sleeve from a reliable supplier that provides an additional external seal.

d. During freezing and storage of cells, the freezing temperature of the tubes must be uniform. Uneven freezing will result in the development of ice plugs in the tube. Ice plugs can inhibit fluid temperature transfer on both sides, creating dangerously high pressures and causing damage to the tube.

e. Do not freeze more samples than the maximum working volume of the cryo tube limit.

f. All cryo tubes have the possibility of explosion. Especially, cryotubes used to contain samples stored under liquid nitrogen may explode without warning. The tube explosion is thought to have been caused by liquid nitrogen entering the tube through tiny cracks and then expanding rapidly as the tube thawed. Therefore, safety equipment should be used throughout the operation of cryo vial ice-out. You should wear a face shield, goggles, heavy gloves, and a buttoned lab coat over pants or a long skirt when having this operation. When thawing the cryo tubes, you should place them in a 37 °C water bath immediately after taking them out. Then, you should gently shake the tube to thaw it within 1 minute. Be careful that the water does not exceed the edge of the freezer tube cover, otherwise, contamination may occur. Due to the high indoor temperature in summer, extra care should be taken when accessing refrigerated pipes.

The lid of the cryo tube is divided into two types -- internal screw cap and external screw cap.

Cryo tubes with internal screw caps are used to freeze biological samples in liquid nitrogen. The silicone gasket at the mouth of the tube enhances the sealing of the tube, which can effectively stop the infiltration of liquid nitrogen.

Cryo tubes with external screw caps are used to freeze samples in the refrigerator. The threaded cap with external screw cap reduces the probability of contamination when handling samples.

a. The cap is designed to be easy to rotate and open.

b. Both tube cap and tube body are produced from the same batch and polypropylene (PP) material. PP is durable and chemically resistant. The same coefficient of expansion ensures a seal of the cryo tube at any temperature.

c. Large marking area for easy writing.

d. The tube is transparent, making it easy to view the sample.

e. Boilable and autoclavable, RNase/DNase free, no heat source.

f. Round bottom design of the cryotube facilitates pouring of liquids and reduces residue.

The function of cryotubes is to preserve samples at low temperatures. It has better low-temperature resistance than EP tubes. Generally, the material becomes brittle as the temperature decreases, and the brittleness temperature of the frozen tube is lower.

The current method of preserving cells for a long time is to freeze them in liquid nitrogen (-196 °C) for up to one year or even several years. Most cells can still grow and multiply when thawed at the time of use. For proper storage, cells should be resuscitated and cultured after one year of freezing and then frozen again. Protective agents should be added when freezing cells, otherwise, the water inside and outside the cells will form ice crystals and cause mechanical damage to the cells. The freezing point can be lowered by adding a protective agent. Under the condition of slow freezing, the water inside the cell leaks out before freezing to avoid ice crystal damage. In addition, culture fluid is added to the lyophilization solution to provide nutrition, osmotic pressure, and pH. Cell recovery refers to thawing frozen cells. Thawing cells should be fast so that they quickly pass through the most vulnerable -5℃ to 0 ℃.

The use of cryo tube is not a simple process like first, opening the liquid nitrogen tank; second, putting in the tubes; third, closing the tank. There are certain safety risks when using cryo tubes, so the scientific and correct use of them can avoid the loss of samples and protect the safety of test personnel.

a. Wash the cells with a pre-warmed PBS solution. Cover the cells with a solution containing trypsin and EDTA at the time of aspiration, a thin layer of liquid is sufficient. The concentration of trypsin and EDTA needs to be determined according to the cell line.

b. Incubate cells at 37°C for 3-5 minutes.

c. After the cells are detached from the bottom, terminate the incubation. Add the medium containing serum and gently suspend the cells with a pipette.

d. Centrifuge the cell suspension (500 x g, 5 min) and resuspend it with a medium containing serum.

e. Cell Counting.

f. Centrifuge the cell suspension (500 x g, 5 min). Remove the supernatant and resuspend the cells with an appropriate volume of medium containing serum.

g. Cells and lyophilization solution (60% medium, 20% fetal bovine serum, 20% DMSO) are mixed in a 1:1 volume ratio and transferred to cryogenic tubes. The density of frozen cells is 1-5×106 cells/ml.

h. Cryo tubes containing cells are recommended to be cooled down at a rate of -1 K/min and can be placed in a container containing isopropanol at -70°C. If the tubes are used to store other samples, they can be placed directly at -20°C, -70°C, or in the gas phase of liquid nitrogen. To ensure uniform freezing of samples, 4 mL and 5 mL cryo tubes need to be placed in a -20°C refrigerator overnight before transferring to -70°C or liquid nitrogen in the gas phase.

i. Then transfer the tubes to a liquid nitrogen tank. To avoid contamination and for safety reasons, it is better to place cryo tubes in the gas phase of liquid nitrogen, do not place them directly in the liquid phase. (Caution: if you use incorrect storage methods, liquid nitrogen remaining in the cryo tube will lead to explosion or leakage of biohazardous substances due to vaporization.)

a. Take the cryo tube out of liquid nitrogen with large forceps and put it into the water at a temperature of 37℃ to 42 ℃. Shake it from time to time to make it pass through -5 to 0 ℃ quickly. Constantly shaking to make it quickly through -5 to 0 ℃.

b. In the ultra-clean table, the neck of the cryotube should be rubbed with an abrasive wheel and sterilized with alcohol cotton balls. Then, open the cryo tube, aspirate the cell suspension into a culture flask, dilute it at 10×, and incubate it in a CO2 incubator. The next day, the freezing medium should be replaced with a fresh culture medium.

If you are interested in our cryo tubes or have any questions, please write an e-mail to [email protected], we will reply to you as soon as possible.