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View additional product information for Synth-a-Freeze™ Cryopreservation Medium - FAQs (A1254201)
18 product FAQs found
我们推荐使用含10% FBS和10% DMSO的DMEM，或者Recovery细胞培养冷冻培养基。您可以尝试常规冷冻细胞的操作流程，仅更换冷冻溶液。
没有，但是有FDA备案的设备管理档案。请通过outlicensing@lifetech.com联系我们的许可证团队获取设备管理档案备案说明
Synth-a-Freeze冻存培养基适用于任何标准冷冻方案。其性能可媲美我们标准含有血清的冻存培养基，适合低温储存各种类型的细胞，例如人类角质细胞、胚胎干细胞、神经干细胞和间充质干细胞等。
不可以，不能使用冻存的组织。
当从Thermo Fisher Scientific购买了Gibco或Invitrogen冻存或正处于增殖期的细胞，可对其进行扩增和重新冻存。不过，冻存操作可能会对细胞的生长状态有所影响。下列实验方案为您提供了一份使用Synth-a-Freeze培养基冻存细胞的基础指南，Synth-a-Freeze培养基是Thermo Fisher Scientific旗下的一款成分确定，无蛋白成份的冻存培养基。
请注意：由于冻存设备与个人技术之间的差异，我们无法保使用本方案冻存的细胞在复苏后能够保持活力，我们也无法为研究用户实验室冻存细胞的效果进行担保。
1. 使用37°C水浴化冻Synth-a-Freeze培养基(https://www.thermofisher.com/order/catalog/product/A1254201)或4°C条件下过夜化冻。
2. 如果在水浴中进行化冻，请确保温度不要超过37°C，也勿将该产品延长时间置于37°C。
3. Synth-a-Freeze培养基在使用前应置于4°C条件下彻底平衡。为获得最优结果，推荐客户使用能够控制变温速率的冰箱。如果缺少能够控制变温速率的冰箱，也可使用细胞冻存盒（如Thermo Scientific Mr Frosty container)。
4. 如需用酶试剂解离培养器皿表面的细胞，则需使用对应的终止溶液重悬细胞以中和酶的效果。
5. 通过离心沉淀细胞。
6. 去除上清液后，使用预冷的Synth-a-Freeze培养基以5x10E5至3x10E6细胞/毫升的密度重悬细胞。
7. 将细胞悬液分装至合适数量的冻存管中。
8. 将细胞尽快冷却至4°C。
9. 如果使用控制变温速率的冰箱：以每分钟降低1°C的速率冰冻样品，直至–40°C。之后按照每分钟降低2°C的速率降至–90°C左右。
10. 如果使用细胞冻存盒：请按照说明书来准备冻存盒。
为获得最佳结果，我们推荐您在细胞温度降至–80°C后，尽快将冻存管转移至液氮储存设备的气相中。
作为Synth-a-Freeze培养基的替代品，可使用该冻存细胞推荐的基础培养基，添加10%胎牛血清（FBS）和10% DMSO进行细胞冻存。请注意不推荐将Synth-a-Freeze培养基用于人表皮黑素细胞的冻存操作。
我们不推荐在种板前离心细胞以去除冻存培养基。尤其是在不适当的高速条件下，离心对细胞有损伤。以我们Invitrogen细胞培养实验室目前经验的来看，如果DMSO的浓度足够低，不会对细胞造成伤害。因此，我们的产品说明中提供了一份详细方案，其中包括如何使用推荐的接种密度和培养基的体积稀释细胞使得DMSO的终浓度低于0.4%（v/v）。
下列步骤以单管冻存细胞进行培养的实验方案为例。
1.准备一烧杯37°C的水。
2. 从液氮储存中取出一管细胞，注意保护手和眼睛。
3. 拧松管盖1/4圈，等待10秒钟以释放螺纹中可能残留的液氮，再重新拧紧管盖。
4. 将冻存管的下半部分置于37°C水浴中进行解冻，直至冻存管内仅剩余一小块冰芯，使细胞迅速解冻。
5. 用消毒液擦拭管体外表面，再将其移至II级A型层流细胞培养通风橱中。
6. 打开管盖，使用1 mL移液器上下吹打细胞悬液以分散细胞。
7. 从管中吸取20 uL细胞悬液，并将其稀释于20 uL台盼蓝溶液中（如：Gibco台盼蓝，货号 15250-061）。
8. 使用血细胞计数板来确定每毫升悬液中的活细胞数。
9. 将管中悬液（1 mL）稀释至产品说明中的推荐浓度（例如1.25 x 10E4活细胞/毫升，Gibco 新生人表皮角质形成细胞）。
10. 将5 mL细胞悬液加入25 cm2培养瓶，或将15 mL细胞悬浮液加入75cm2培养瓶中。
11. 摇匀培养瓶中的培养基以彻底分散细胞。许多类型的细胞都会迅速贴壁，如果没有在传代后即刻摇匀细胞，细胞可能生长不均匀。
12. 在37°C、5% CO2/95%空气、湿度细胞培养箱中培养细胞。为了获得最佳结果，培养开始后至少在24小时之内不要扰动细胞。
请点击此处(https://www.thermofisher.com/cn/zh/home/references/gibco-cell-culture-basics/cell-culture-protocols/thawing-cells.html)查看我们的细胞复苏方案。
请点击此处（https://www.thermofisher.com/us/en/home/references/gibco-cell-culture-basics/cell-culture-protocols/freezing-cells.html）查看我们的细胞冻存方案。
We recommend using DMEM plus 10% FBS and 10% DMSO, or the Recovery Cell Culture Freezing Medium. You can try the same general recommendations for freezing cells, just changing the freezing solution.
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
There isn't a Drug Master File, but there is a Device Master File on file with the FDA. Please contact our Licensing team at outlicensing@thermofisher.com in order to obtain instructions for referencing this Device Master File.
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
Synth-a-Freeze Cryopreservation Medium can be used with any standard freezing protocol. It offers performance comparable to that of our standard, serum-containing cryopreservation medium for cyropreserving a variety of cell types including human keratinocytes, embryonic stem cells, neural stem cells, and mesenchymal stem cells.
Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.
No, the tissue was not cryopreserved.
When either Gibco or Invitrogen cryopreserved or proliferating cultures are purchased from us, they may be expanded and cryopreserved again. However, the cryopreservation process may result in altered growth performance of the cells. The following protocol provides a basic guideline for the cryopreservation of cells using Synth-a-Freeze medium, a defined, protein-free cryopreservation medium available from us.
Please note: Due to differences in cryopreservation equipment and individual techniques, we cannot guarantee that cells cryopreserved using this protocol will be viable upon recovery from cryopreservation, and we do not provide a warranty for cells cryopreserved in an investigator's laboratory.
1. Thaw Synth-a-Freeze medium in a 37 degrees C water bath or overnight at 4 degrees C.
2. If thawed in a water bath, do not exceed 37 degrees C and do not leave the product at 37 degrees C for an extended period of time.
3. Synth-a-Freeze medium should be equilibrated to 4 degrees C prior to use. For optimal results, the use of a controlled-rate freezer is recommended. In the absence of a controlled-rate freezer, a cell cryopreservation container (e.g., Thermo Scientific Mr Frosty container) may be useful.
4. If enzymatic agents are used to remove the cells from a culture surface, resuspend the cells in a solution that will neutralize the effects of the enzyme.
5. Pellet the cells by centrifugation.
6. After removing the supernatant, resuspend the cell pellet in cold Synth-a-Freeze medium at a concentration of 5 x 10E5 to 3 x 10E6 cells/mL.
7. Distribute the cell suspension in an appropriate number of cryopreservation vials.
8. Cool the vials of cells to 4 degrees C as quickly as possible.
9. If using a controlled-rate freezer: freeze the material by reducing the temperature 1degrees C per minute until the temperature reaches -40 degrees C. Then reduce the temperature at a rate of 2 degrees C per minute until the temperature reaches approximately -90 degrees C.
10. If using a cell cryopreservation container: Prepare the container according to the manufacturer's instructions.
For best results we recommend transferring the vials to the vapor phase of a liquid nitrogen storage facility as soon as possible after the cells have reached -80 degrees C.
As a substitute for Synth-a-Freeze medium, the recommended basal medium for the cell type being cryopreserved, supplemented with 10% fetal bovine serum (FBS) and 10% DMSO, may be used. Please note that Synth-a-Freeze medium is NOT recommended for the cryopreservation of human epidermal melanocytes.
Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.
We do not recommend spinning cells out of cryopreservation medium prior to plating. Centrifugation can be harmful to cells, particularly if inappropriately high speeds are used. Experience in our in-house cell culture laboratory has shown that cells do not suffer deleterious effects if the DMSO concentration is sufficiently low. Therefore, our product instructions include a detailed protocol that involves diluting the cells into culture medium such that the final DMSO concentration is less than 0.4% (v/v) at the recommended seeding density and volume of medium.
Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.
The procedure given below is a sample protocol for establishing cultures from the contents of one vial.
1. Prepare a beaker of water at 37 degrees C.
2. Remove a vial of cells from liquid nitrogen storage, taking care to protect hands and eyes.
3. Loosen the cap on the vial 1/4 turn for 10 seconds to release any liquid nitrogen that may be trapped in the threads, then re-tighten the cap.
4. Dip the lower half of the vial into the 37 degrees C water to thaw.
5. When the contents of the vial have thawed, wipe the outside of the vial with disinfecting solution and move to a Class II, type A laminar flow culture hood.
6. Open the vial and pipette the suspension up and down with a 1 mL pipette to disperse the cells.
7. Remove 20 µL from the vial and dilute the cell suspension in 20 µL of trypan blue solution (for example: Gibco Trypan Blue, Cat. No. 15250-061).
8. Use a hemacytometer to determine the number of viable cells per mL.
9. Dilute the contents of the vial (1 mL) to the concentration recommended by the product instructions (for example 1.25 X 10E4 viable cells/mL for Gibco neonatal human epidermal keratinocytes ).
10. Add 5 mL of cell suspension to each 25 cm2 culture flask or 15 mL of cell suspension to each 75 cm2 culture flask.
11. Following inoculation, swirl the medium in the flasks to distribute the cells. Many cell types attach to culture surfaces quickly, and if the medium is not distributed immediately following inoculation, the cells may grow in uneven patterns.
12. Incubate the cultures in a 37 degrees C, 5% CO2/95% air, humidified cell culture incubator. For best results, do not disturb the culture for at least 24 hours after the culture has been initiated.
Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.
Please see our protocol here for thawing frozen cells (https://www.thermofisher.com/us/en/home/references/gibco-cell-culture-basics/cell-culture-protocols/thawing-cells.html).
Please see our protocol here for freezing cells (https://www.thermofisher.com/us/en/home/references/gibco-cell-culture-basics/cell-culture-protocols/freezing-cells.html).