HK EGFP-alpha-tubulin/H2B-mCherry

HK EGFP-α-tubulin/H2B-mCherry HeLa Kyoto 细胞系是一种精心设计的模型，用于详细观察细胞过程。该克隆系经稳定转染后可表达两种荧光蛋白融合体，从而实现染色质和微管网络的实时成像。红色荧光蛋白 mCherry 与核心组蛋白 H2B 融合，形成 H2B-mCherry。这种融合蛋白由 pH2B-mCherry-IRES-neo3 质粒表达，可作为染色质标记，在活细胞成像中突出显示核 DNA，促进染色质动力学和核结构研究。

此外，该细胞系通过 pmEGFP-α-tubulin-IRES-puro2b 质粒表达融合了α-微管蛋白的单体增强型 GFP（绿色荧光蛋白）。GFP-α-微管蛋白融合物发出鲜艳的绿色荧光，勾勒出细胞内的微管结构。这一特性对于研究微管的组织、动力学及其在细胞分裂和细胞内运输中的作用至关重要。这些构建体的稳定整合可对这些细胞成分进行连续、长期的观察，而无需重复转染，从而减少了变异性，提高了实验结果的可靠性。转染后的耐药性选择确保了该品系细胞表达的稳定性和一致性。

37°C, 5%_CO2, 加湿环境。

1. Confirm that the vial remains deeply frozen upon delivery, as cells are shipped on dry ice to maintain optimal temperatures during transit.

2. Upon receipt, either store the cryovial immediately at temperatures below -150°C to ensure the preservation of cellular integrity, or proceed to step 3 if immediate culturing is required.

3. For immediate culturing, swiftly thaw the vial by immersing it in a 37°C water bath with clean water and an antimicrobial agent, agitating gently for 40-60 seconds until a small ice clump remains.

4. Perform all subsequent steps under sterile conditions in a flow hood, disinfecting the cryovial with 70% ethanol before opening.

5. Carefully open the disinfected vial and transfer the cell suspension into a 15 ml centrifuge tube containing 8 ml of room-temperature culture medium, mixing gently.

6. Centrifuge the mixture at 300 x g for 3 minutes to separate the cells and carefully discard the supernatant containing residual freezing medium.

7. Gently resuspend the cell pellet in 10 ml of fresh culture medium. For adherent cells, divide the suspension between two T25 culture flasks; for suspension cultures, transfer all the medium into one T25 flask to promote effective cell interaction and growth.

8. Adhere to established subculture protocols for continued growth and maintenance of the cell line, ensuring reliable experimental outcomes.

使用基于 PCR 的检测方法和基于发光的支原体检测方法排除支原体污染。

为确保没有细菌、真菌或酵母菌污染，每天都要对细胞培养物进行目视检查。

如需长期保存，可将样品瓶放入气相液氮中，温度约为 -150 至 -196 ℃。在-80 °C下保存只能作为转入液氮前的短暂过渡。

冷冻保存的细胞系用干冰装运，并用经过验证的隔热包装和足够的制冷剂在整个运输过程中保持约 -78 °C 的温度。收到货后，应立即检查容器，并立即将小瓶转移到适当的储藏室。

在Cytion，我们保证产品的质量和活力。 我们保证，从我们的工厂发货后 60 天内，我们的细胞在开始培养时都是存活的。 我们保证所有材料均符合分析证书和/或产品说明书中描述的规格。

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