X'inhuma t-tubi krijo?
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. Tubi krijo, magħruf ukoll bħala kunjetti tal-ħażna krijoġeniċi, are a special type of tubu tat-test. 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.
Mhux it-tubi krijo kollha jistgħu jintużaw għan-nitroġenu likwidu, krijoġeniku kunjetti għan-nitroġenu likwidu huma ddisinjati biex jissodisfaw it-talbiet tat-temperaturi tan-nitroġenu likwidu. Huma krijotubi speċjali b'siġillar għoli u reżistenza għat-temperatura baxxa.
Daqsijiet tat-tubi krijo
M'hemm l-ebda klassifikazzjoni speċifika ta ' tubu krijo, huwa ġeneralment maqsum skond il-kapaċità kif ġej:
| Daqsijiet tat-tubi krijo |
|---|
| Kunjetti krijoġeniċi ta' 0.5 ml |
| 1 ml tubi krijoġeniċi / 1 ml krijotubi |
| 1.5 ml tubi krijoġeniċi / kunjetti krijoġeniċi 1.5 ml / tubi krijoġeniċi 1.5 ml |
| 1.8 ml cryotube / cryotube 1.8 ml |
| 2 ml tubi krijoġeniċi / krijoġeniċi 2ml / krijotubi 2ml / 2ml kunjetti krijoġeniċi / kunjetti krijoġeniċi 2ml / tubu krijoġeniku 2ml |
| Tubi krijoġeniċi ta’ 4 ml |
| 5ml krijovjali / 5ml tubi krijoġeniċi / 5 ml krijotubi / krijotubi 5ml / krijovjali 5ml |
| Tubi krijoġeniċi ta’ 7 ml |
| Tubi krijoġeniċi ta’ 10 ml |
Karatteristiċi tat-tubi krijo
Sigurtà ta' tubi krijo
Tubi krijo huma tubi tal-laboratorju użati biex jaħżnu kampjuni f'temperaturi baxxi. Użu mhux xieraq ta' krijotubu jista 'jikkawża splużjonijiet, li jirriżulta fi tnixxija ta' materjali perikolużi u, f'każijiet severi, anke korrimenti. Biex jitnaqqas ir-riskju ta tubu krijo splużjoni, jekk jogħġbok oqgħod attent għal-linji gwida li ġejjin meta tużaha.
a. Tubu krijo huwa ġeneralment magħmul minn polyethylene ta 'densità għolja jew polypropylene. Jista 'jinqasam f'żewġ kategoriji skont xenarji ta' użu differenti. Wieħed huwa krijotubi għall-kultura taċ-ċelluli u l-ieħor huwa krijotubi għan-nitroġenu likwidu. Dawn iż-żewġ tipi ta krijotubi jeħtieġ li jiġu distinti bir-reqqa qabel ix-xiri u l-użu, peress li t-temperaturi applikabbli tagħhom huma partikolarment differenti.
| Daqsijiet tat-tubi krijo |
|---|
| Kunjetti krijoġeniċi ta' 0.5 ml |
| 1 ml tubi krijoġeniċi / 1 ml krijotubi |
| 1.5 ml tubi krijoġeniċi / kunjetti krijoġeniċi 1.5 ml / tubi krijoġeniċi 1.5 ml |
| 1.8 ml cryotube / cryotube 1.8 ml |
| 2 ml tubi krijoġeniċi / krijoġeniċi 2ml / krijotubi 2ml / 2ml kunjetti krijoġeniċi / kunjetti krijoġeniċi 2ml / tubu krijoġeniku 2ml |
| Tubi krijoġeniċi ta’ 4 ml |
| 5ml krijovjali / 5ml tubi krijoġeniċi / 5 ml krijotubi / krijotubi 5ml / krijovjali 5ml |
| Tubi krijoġeniċi ta’ 7 ml |
| Tubi krijoġeniċi ta’ 10 ml |
b. All kunjetti tal-krijotubi għandu jkollu siegla tas-silikonju, siegla tal-gomma, jew siegla O-ring biex tiżgura siġill issikkat. Jekk jogħġbok innota li l-issikkar żżejjed tal-għatu jista 'jidforma s-siġill.
c. Tubi krijo għandha tinħażen fil-fażi tal-gass 'il fuq min-nitroġenu likwidu u mhux mgħaddsa fil-likwidu biex tiġi evitata l-penetrazzjoni tan-nitroġenu likwidu fit-tubi. L-iffullar ta' kontenituri tal-ħażna għandu jiġi evitat. Jekk il-kampjuni jridu jinħażnu fil-fażi likwida, huwa rakkomandat li tuża kmiem protettiv professjonali minn fornitur affidabbli li jipprovdi siġill estern addizzjonali.
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 tubu krijo limitu.
f. All krijo tubi għandhom il-possibbiltà ta’ splużjoni. Speċjalment, krijotubi użat biex ikun fih kampjuni maħżuna taħt nitroġenu likwidu jista’ jisplodi mingħajr twissija. L-isplużjoni tat-tubu huwa maħsub li kienet ikkawżata minn nitroġenu likwidu li daħal fit-tubu minn xquq ċkejkna u mbagħad jespandi malajr hekk kif it-tubu jinħall. Għalhekk, tagħmir ta 'sikurezza għandu jintuża matul l-operazzjoni ta' kunjett krijo ice-out. You should wear a Tarka tal-wiċċ, goggles, heavy gloves, and a buttoned Kowt tal-laboratorju 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.
Għotjien ta' tubi krijo
L-għatu tal- tubu krijo is divided into two types — internal screw cap and external screw cap.
Kap tal-kamin intern tat-tubi krijo
Tubi krijo b'tappijiet bil-kamin interni jintużaw biex jiffriżaw kampjuni bijoloġiċi fin-nitroġenu likwidu. Is-siegla tas-silikonju fil-bokka tat-tubu ttejjeb is-siġillar tat-tubu, li jista 'effettivament iwaqqaf l-infiltrazzjoni tan-nitroġenu likwidu.
Kap tal-kamin estern tat-tubi krijo
Tubi krijo b'tappijiet bil-kamin esterni jintużaw biex jiffriżaw kampjuni fil-friġġ. L-għatu bil-kamin b'għatu bil-kamin estern inaqqas il-probabbiltà ta 'kontaminazzjoni meta timmaniġġja l-kampjuni.
Vantaġġi tat-tubi krijo
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.
Użu ta' tubi krijo
Il-funzjoni ta ' krijotubi huwa li tippreserva l-kampjuni f'temperaturi baxxi. Għandu reżistenza aħjar għat-temperatura baxxa minn tubi EP. Ġeneralment, il-materjal isir fraġli hekk kif it-temperatura tonqos, u t-temperatura tal-fraġilità tat-tubu iffriżat hija aktar baxxa.
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 lajofilizzazzjoni 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 ℃.
L-użu ta ' tubu krijo is not a simple process like first, opening the tank tan-nitroġenu likwidu; second, putting in the tubes; third, closing the tank. There are certain safety risks when using krijo tubi, sabiex l-użu xjentifiku u korrett tagħhom jista 'jevita t-telf ta' kampjuni u jipproteġi s-sigurtà tal-persunal tat-test.
Passi għall-iffriżar taċ-ċelluli
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. Iċċentrifuga 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.)
Passi ta 'rkupru taċ-ċelluli
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 garafina, dilute it at 10×, and incubate it in a CO2 inkubatur. The next day, the freezing medium should be replaced with a fresh culture medium.