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HK EGFP-LaminB1/H2B-mCherry
Tissue
子宮頸
Disease
癌
Metastatic site
原發性腫瘤部位(子宮頸)
Synonyms
HeLa Kyoto EGFP-LaminB1 and H2B-mCherry

Documents

HK EGFP-LaminB1/H2B-mCherry

Cell line · Product number 300919

The HK EGFP-LaminB1/H2B-mCherry cell line is an Hela Kyoto derived in vitro model designed for real-time visualization of chromatin dynamics and nuclear a...

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01General information

Description

HK EGFP-LaminB1/H2B-mCherry 細胞系是一種源自 Hela Kyoto 的體外模型,專為即時觀察活細胞中的染色質動態與核結構而設計。 該細胞系表達兩種螢光蛋白融合蛋白:EGFP(增強型綠色螢光蛋白)與 Lamin B1 的融合蛋白,以及 mCherry(一種紅色螢光蛋白)與組蛋白 H2B 的融合蛋白。 EGFP 與 Lamin B1 的融合蛋白可讓研究人員觀察核膜與核層,這些結構對於維持細胞核的完整性與功能至關重要。Lamin 蛋白屬於 V 型中間絲蛋白,會在內核膜下方形成網狀結構,在細胞核穩定性、染色質組織及基因調控方面扮演關鍵角色。

另一方面,帶有 mCherry 標記的組蛋白 H2B 則能使細胞核內的染色質顯影。組蛋白是核小體的基本組成部分,參與將 DNA 組織成染色質的過程,因此對於 DNA 複製、修復及轉錄至關重要。 H2B 上的 mCherry 標籤會發出鮮豔的紅色螢光,與 EGFP 的綠色螢光形成鮮明對比,使研究人員能在活細胞實驗中同時對核結構與染色質進行雙重成像。 此細胞系常被用於研究核力學、有絲分裂及基因組穩定性,能提供動態視角來觀察那些難以即時觀察的細胞過程。

Tissue
子宮頸
Disease
癌
Metastatic site
原發性腫瘤部位(子宮頸)
Applications
核膜基質與染色質組織;Lamin B1 動態;H2B 染色質成像;雙色活細胞螢光;核力學;有絲分裂;基因組穩定性;核膜生物學
Synonyms
  • HeLa Kyoto EGFP-LaminB1 and H2B-mCherry

02Classification

Organism
Human
Sex
Female
Growth properties
Adherent
GMO status
GMO-S1
GMO status details
GMO-S1: This HeLa Kyoto line contains EGFP-Lamin B1 and H2B-mCherry constructs for imaging nuclear envelope and chromatin organization. This classification applies only within Germany and may differ elsewhere.
Biosafety level
1

03Characteristics

Morphology
呈馬賽克狀石塊形狀的類上皮細胞
Cell type
上皮細胞

04Identifiers

Citation
HK EGFP-LaminB1/H2B-mCherry(Cytion 型號 300919)
NCBI TaxID
9606
Cellosaurus
CVCL_UR41
Depositor
艾倫伯格實驗室(EMBL)

05Expression and susceptibility

Protein expression
EGFP-LaminB1/H2B-mCherry
Products
組蛋白 H2B

06Handling and culture

Culture medium
DMEM,含 4.5 g/L 葡萄糖、4 mM L-谷氨醯胺、3.7 g/L NaHCO₃ 及 1.0 mM 丙酮酸鈉(Cytion 產品編號 820300a)
Incubation atmosphere

37°C,5% CO₂,加濕環境。

Supplements
在培養基中加入 10% 的胎牛血清(FBS)
Dissociation reagent
Accutase
Subculturing
從貼壁細胞中移除舊培養基,並用不含鈣和鎂的PBS洗滌細胞。T25培養瓶使用3-5毫升PBS,T75培養瓶則使用5-10毫升。 接著,用 Accutase 完全覆蓋細胞,T25 培養瓶使用 1–2 毫升,T75 培養瓶則使用 2.5 毫升。將細胞於室溫下孵育 8–10 分鐘,使其脫附。 孵育後,將細胞與 10 毫升培養基輕柔混合以使其重新懸浮,然後以 300xg 離心 3 分鐘。棄去上清液,將細胞重新懸浮於新鮮培養基中,並轉移至已預先加入新鮮培養基的新培養瓶中。
Seeding density
1 × 10⁴ 個細胞/cm²
Fluid renewal
每週 2 至 3 次
Post-thaw recovery
解凍後,將細胞以 5 × 10⁴ 個細胞/cm² 的密度接種於培養皿中,並讓細胞從冷凍過程中恢復,並附著至少 24 小時。
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

透過基於 PCR 的檢測及基於發光原理的支原體檢測方法,排除支原體污染的可能性。

為確保無細菌、真菌或酵母菌污染,每日均會對細胞培養物進行目視檢查。

Freeze medium
作為冷凍保存培養基,我們使用完全生長培養基(含 FBS)+ 10% DMSO 以確保解凍後的存活率,或使用 CM-1 (Cytion 型號 800100),該培養基含有經優化的滲透保護劑和代謝穩定劑,可提升細胞恢復率並減輕冷凍誘導的壓力。
Storage conditions

若需長期保存,請將試管置於溫度約為 −150 至 −196 °C 的氣相液氮中。僅在轉移至液氮之前,可將試管暫時存放於 −80 °C 環境中,作為短期過渡步驟。

Shipping conditions

冷凍保存的細胞系會以乾冰冷藏,並裝於經驗證的隔熱包裝中,包裝內含有足夠的冷媒,以確保在運輸過程中溫度維持在約 −78 °C。收到貨物後,請立即檢查容器,並盡快將試管轉移至適當的儲存場所。

08Donor

Donor age
30年
Ethnicity
非裔美國人