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HK Mad2-LAP/H2B-mCherry
Tissue
子宫颈
Disease
癌症
Synonyms
HeLa Kyoto Mad2-LAP and H2B-mCherry, HeLa Kyoto Mad2-LAP

Documents

HK Mad2-LAP/H2B-mCherry

Cell line · Product number 300920

The HK Mad2-LAP/H2B-mCherry cell line is a genetically engineered cell model extensively utilized for studying chromosome segregation and the spindle asse...

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1

01General information

Description

HK Mad2-LAP/H2B-mCherry 细胞系是一种基因工程细胞模型,广泛用于研究染色体分离和有丝分裂过程中纺锤体组装检查点。这些细胞来源于 HeLa Kyoto 细胞,这是一种强健的人类细胞系,最初取自宫颈癌。该细胞系的 HK Mad2-LAP(LAP 标记的 Mad2)有助于对 Mad2 蛋白进行可视化和功能分析,Mad2 蛋白是纺锤体组装检查点的一个重要组成部分,它能阻止无丝分裂期的开始,直到所有染色体在移子板上正确排列。

加入 H2B-mCherry 后,组蛋白 H2B 被标记为 mCherry 荧光蛋白,可对细胞分裂过程中的染色质动态进行实时成像。这一特性使 HK Mad2-LAP/H2B-mCherry 细胞系成为高分辨率活细胞成像技术的绝佳工具,用于观察各种实验条件下人类细胞的染色体运动和有丝分裂进程。荧光标签的使用有助于精确跟踪和量化,从而为细胞周期调控和染色体稳定性的分子机制提供了宝贵的见解。

Tissue
子宫颈
Disease
癌症
Synonyms
  • HeLa Kyoto Mad2-LAP and H2B-mCherry
  • HeLa Kyoto Mad2-LAP

02Classification

Organism
Human
Sex
Female
Growth properties
Adherent
GMO status
GMO-S1
GMO status details
GMO-S1: This HeLa Kyoto line contains Mad2-LAP and H2B-mCherry constructs enabling visualization of spindle checkpoint dynamics. This classification applies only within Germany and may differ elsewhere.
Biosafety level
1

03Characteristics

Morphology
具有镶嵌石形状的上皮样细胞

04Identifiers

Citation
HK Mad2-LAP/H2B-mCherry(Cytion 目录号 300920)
NCBI TaxID
9606
Cellosaurus
CVCL_1D65
Depositor
艾伦伯格实验室(EMBL)

05Expression and susceptibility

Protein expression
Mad2-LAP/H2B-mCherry

06Handling and culture

Culture medium
DMEM,w:4.5 克/升葡萄糖,w:4 毫摩尔 L-谷氨酰胺,w:3.7 克/升 NaHCO3,w:1.0 毫摩尔丙酮酸钠(Cytion 编号 820300a)。
Incubation atmosphere

37°C, 5%CO2, 加湿环境。

Supplements
在培养基中添加 10% FBS
Dissociation reagent
Accutase
Subculturing
去除粘附细胞上的旧培养基,用不含钙和镁的 PBS 冲洗。T25 烧瓶用 3-5 毫升 PBS,T75 烧瓶用 5-10 毫升。然后用 Accutase 完全覆盖细胞,T25 烧瓶用 1-2 毫升,T75 烧瓶用 2.5 毫升。让细胞在室温下孵育 8-10 分钟,使其分离。孵育后,用 10 毫升培养基轻轻混合细胞使其重悬,然后用 300xg 离心 3 分钟。弃去上清液,用新鲜培养基重悬细胞,并将其转移到已装有新鲜培养基的新烧瓶中。
Seeding density
1 × 10⁴ 个细胞/平方厘米
Fluid renewal
每周 2 至 3 次
Post-thaw recovery
解冻后,将细胞以5×10⁴个细胞/平方厘米的密度接种于培养皿中,使细胞从冷冻过程中恢复并至少贴壁24小时。
Thawing and culturing
  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.

07Quality control and cryopreservation

Sterility

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。

Freeze medium
我们使用完全生长培养基(包括 FBS)+10% DMSO 或 CM-1(Cytion 商品目录编号 800100)作为冷冻保存培养基,以获得足够的解冻后存活率,CM-1(Cytion 商品目录编号 800100)含有优化的渗透保护剂和代谢稳定剂,可提高恢复能力并减少冷冻引起的应激。
Storage conditions

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。

Shipping conditions

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。

08Donor

Donor age
30年
Ethnicity
非洲裔美国人