冷凍細(xì)胞的解凍 Thawing of frozen cells
Caesar
介紹?Introduction
The thawing procedure is stressful to frozen cells, and using good technique and working quickly ensures that a high proportion of the cells survive the procedure. As with other cell culture procedures, we recommend that you closely follow the instructions provided with your cells and other reagents for best results.?解凍過(guò)程對(duì)冷凍細(xì)胞來(lái)說(shuō)是緊張的，使用好的技術(shù)和快速的工作可以確保很大比例的細(xì)胞存活下來(lái)。與其他細(xì)胞培養(yǎng)程序一樣，我們建議您嚴(yán)格按照細(xì)胞和其他試劑提供的說(shuō)明進(jìn)行操作，以獲得最佳效果
Thaw frozen cells rapidly (< 1 minute) in a 37°C water bath.?在攝氏37度的水浴中快速解凍凍住的細(xì)胞(< 1分鐘)
Dilute the thawed cells slowly, using pre-warmed growth medium.?用預(yù)先加熱的培養(yǎng)基緩慢稀釋解凍的細(xì)胞
Plate thawed cells at high density to optimize recovery.?平板高密度解凍細(xì)胞以優(yōu)化回收率
Always use proper aseptic technique and work in a laminar flow hood.?始終使用適當(dāng)?shù)臒o(wú)菌操作，并在層流罩內(nèi)工作
Always wear personal protective equipment, including a face mask or goggles. Cryovials stored in liquid-phase present a risk of explosion when thawed.?始終佩戴個(gè)人防護(hù)裝備，包括面罩或護(hù)目鏡。液相儲(chǔ)存的冰凍卵在解凍時(shí)有爆炸的危險(xiǎn)
Some freezing media contain DMSO, which is known to facilitate the entry of organic molecules into tissues. Handle reagents containing DMSO using equipment?一些冷凍介質(zhì)含有 DMSO，這是眾所周知的促進(jìn)有機(jī)分子進(jìn)入組織。使用設(shè)備處理含有 DMSO 的試劑
實(shí)驗(yàn)材料?Experimental material
Cryovial containing frozen cells?含有冷凍細(xì)胞的冷凍瓶
Complete growth medium, pre-warmed to 37°C?全生長(zhǎng)培養(yǎng)基，預(yù)熱至攝氏37度
Disposable, sterile centrifuge tubes?一次性無(wú)菌離心管
Water bath at 37°C?攝氏37度水浴
70% ethanol?70% 乙醇
Tissue-culture treated flasks, plates, or dishes?經(jīng)組織培養(yǎng)處理的燒瓶、盤子或培養(yǎng)皿
Protocol for Thawing Frozen Cells?冷凍細(xì)胞解凍方案
視頻 5: 復(fù)蘇細(xì)胞?Video 5: resuscitating cells
視頻5介紹了細(xì)胞解凍的最佳方法，且在此過(guò)程中不會(huì)對(duì)細(xì)胞造成不利影響。 Our scientists demonstrate how to carefully transfer cells from storage in liquid nitrogen to the incubator.?Video 5 shows the best way to thaw cells without adversely affecting them in the process. Our scientists demonstrate how to carefully transfer cells from storage in liquid nitrogen to the incubator
The following protocol describes a?下面的協(xié)議描述了一個(gè)general procedure?一般程序?for thawing cryopreserved cells.?用來(lái)解凍凍存的細(xì)胞For detailed protocols, always refer to the cell-specific product insert.?對(duì)于詳細(xì)的協(xié)議，始終參考特定于單元格的產(chǎn)品插入
Remove the cryovial containing the frozen cells from liquid nitrogen storage and?將裝有冷凍細(xì)胞的冷凍小瓶從液氮貯存器中取出immediately?立即?place it into a 37°C water bath.?把它放入37 ° c 的水浴中
Quickly thaw the cells (< 1 minute) by gently swirling the vial in the 37°C water bath until there is just a small bit of ice left in the vial.?快速解凍細(xì)胞(小于1分鐘) ，輕輕旋轉(zhuǎn)小瓶在37 ° c 的水浴，直到只有一點(diǎn)點(diǎn)冰留在小瓶
Transfer the vial it into a laminar flow hood. Before opening, wipe the outside of the vial with 70% ethanol.?將小瓶放入層流罩中，開(kāi)放前用70% 乙醇擦拭小瓶外部
Transfer the desired amount of pre-warmed complete growth medium appropriate for your cell line?轉(zhuǎn)移適合細(xì)胞系的預(yù)熱完全生長(zhǎng)培養(yǎng)基的所需量dropwise?逐層的?into the centrifuge tube containing the thawed cells.?進(jìn)入含有解凍細(xì)胞的離心管
Centrifuge the cell suspension at approximately 200 × g for 5–10 minutes. The actual centrifugation speed and duration varies depending on the cell type.?將細(xì)胞懸浮液以大約200 × g 離心5-10分鐘。實(shí)際離心速度和持續(xù)時(shí)間取決于電池類型
After the centrifugation, check the clarity of supernatant and visibility of a complete pellet. Aseptically decant the supernatant without disturbing the cell pellet.?離心后，檢查上清液的透明度和完整顆粒的可見(jiàn)度。無(wú)菌傾倒上清液，不干擾細(xì)胞顆粒
Gently resuspend the cells in complete growth medium, and transfer them into the appropriate culture vessel and into the recommended culture environment.?將細(xì)胞在完全培養(yǎng)基中輕輕復(fù)蘇，并將其轉(zhuǎn)移到適當(dāng)?shù)呐囵B(yǎng)容器和推薦的培養(yǎng)環(huán)境中
注：?Note:The appropriate flask size depends on the number of cells frozen in the cryovial, and the culture environment varies based on the cell and media type.?合適的燒瓶大小取決于冰凍細(xì)胞的數(shù)量，培養(yǎng)環(huán)境因細(xì)胞和培養(yǎng)基類型的不同而不同
