BioVector? TK-1B 人急性髓系白血病細胞株

BioVector? TK-1B Human Acute Myeloid Leukemia Cell Line

第一部分 中文說明

一 產品基本信息與遺傳學背景

• 細胞名稱：BioVector? TK-1B

• 保藏機構貨號：DSMZ ACC 946

• 物種來源：人類 (Homo sapiens)

• 性別與年齡：男性，22歲（Japanese 種群背景）

• 組織與疾病背景：

  • 該細胞株是在 1984 年建立的。源自一名 22 歲日本男性患者的外周血。該患者最初被診斷為 T 淋巴母細胞淋巴瘤（T-lymphoblastic lymphoma），隨后病情發(fā)生惡性白血病轉變，演變?yōu)?/span>急性髓系白血?。ˋcute Myeloid Leukemia，具體分型為伴有骨髓單核細胞特征的 AML-M4 型）。

  • 構建來源：TK-1B 是通過有限稀釋法（Limited dilution）從其親本細胞株 TK-1 中分離純化出來的單克隆亞株（Subclone）。

• 細胞譜系與分子特征：

  • 髓單核細胞雙重表型特性：TK-1B 展現(xiàn)出與親本 TK-1 高度相似的急性髓單核細胞（Myelomonocytic）的生物學和免疫學特征。

  • EBV 陰性特征：經分子生物學驗證，該克隆細胞系不攜帶愛潑斯坦-巴爾病毒（EBV）核抗原，為內源性非病毒轉化系。

• 免疫表型譜（Immunophenotype）：

  • 陽性表面標記：CD4+、CD7+、CD10+、CD13+、CD15+、CD33+、HLA-DR+。

  • 陰性表面標記：CD2-、CD3-、CD5-、CD8-、CD14-、CD19-、CD20-、CD34-。

• 生物安全級別：1級（BSL-1）。經檢測常見病毒如 EBV, HBV, HCV, HIV-1/2, HTLV-1/2 以及 MLV 均為陰性。

二 細胞形態(tài)學與培養(yǎng)環(huán)境

• 形態(tài)學特征：展現(xiàn)典型的急性髓系白血病細胞特征。在顯微鏡下，細胞呈圓形，在懸浮培養(yǎng)基中主要以單個散在生長為主，部分會聚集形成微小的細胞團塊（Round cells growing singly and partly in small clumps）。

• 生長模式：完全懸浮生長。

• 群體倍增時間（Doubling Time）：大約 48 至 72 小時，屬于生長速度相對溫和的髓系細胞。

• 標準完全培養(yǎng)基配方：

  • 基礎培養(yǎng)基：80% 至 90% BioVector? RPMI-1640 培養(yǎng)基。

  • 維持添加：10% 至 20% 優(yōu)質熱滅活胎牛血清（h.i. FBS）。

• 物理培養(yǎng)參數：37攝氏度恒溫、5% 二氧化碳、空氣飽和濕度。

三 細胞傳代與復蘇標準操作步驟

1. 初始接種與復蘇早期極度敏感特性：

   • 前一周平臺期警告：TK-1B 細胞對冷凍解凍過程以及傳代初期極其敏感。在凍存管初次復蘇接種后的第一周內，細胞通常處于適應期，幾乎沒有明顯的數量增長（No cell growth during first week）。這是該細胞株的固有生物學特性，無需頻繁更換培養(yǎng)基或棄液，此時應保持耐心，靜置培養(yǎng)。

   • 高密度啟動參數：復蘇或初始接種時，必須采用高濃度、高血清的配置。強烈推薦在 24 孔培養(yǎng)板中進行接種，且初始接種活細胞密度必須直接設定在每毫升約 1000000 個活細胞（1 x 10^6 cells/ml）的高水平，同時使用 20% 濃度的胎牛血清 進行加壓維持。

2. 常規(guī)日常傳代執(zhí)行：

   • 當細胞度過復蘇早期的停滯期并進入對數生長期后，細胞開始穩(wěn)定擴增。

   • 日常維持密度窗口：在常規(guī)維持循環(huán)中，應將懸液活細胞密度嚴密控制在 每毫升 500000 至 1000000 個活細胞（0.5-1 x 10^6 cells/ml） 之間。

   • 傳代策略：當細胞收集密度達到或接近每毫升 1500000 個（1.5 x 10^6 cells/ml）的收獲上限時，需要進行分裂傳代。直接通過低速離心收集細胞并丟棄舊基，使用新鮮完全培養(yǎng)基重懸，常規(guī)傳代稀釋比例為 1比2 至 1比3，通常每 2 到 3 天處理一次。

四 核心科研應用方向

1. T淋巴瘤向髓系白血?。ˋML）惡性轉變機制研究：由于 BioVector? TK-1B 的親本起源于極具臨床研究價值的“T淋巴母細胞淋巴瘤向急性髓系白血?。ˋML-M4）惡性轉化”的特殊病例，該細胞株是研究血液腫瘤譜系轉換（Lineage switch）、白血病多克隆演進及跨譜系致癌基因網絡調控的珍貴細胞模型。

2. 急性髓單核細胞白血病（AML-M4）特異性標志物與藥物靶向研究：該細胞株穩(wěn)定表達 CD13、CD33 及 HLA-DR，同時不表達 CD14。這種獨特的免疫學表面標記表型使其常被用于急性髓單核細胞白血病靶向分子的特征性分析、抗體偶聯(lián)藥物（ADCs）的體外選擇性殺傷評價，以及誘導髓系細胞定向成熟分化的表觀遺傳學干預研究。

PART 2 ENGLISH SECTION

I General Information and Genetic Background

• Cell Line Name: BioVector? TK-1B

• Repository Catalog Number: DSMZ ACC 946

• Species Origin: Human (Homo sapiens)

• Sex and Age of Donor: Male, 22 years old (Japanese population profile)

• Tissue and Disease Background:

  • Established in 1984 from the peripheral blood mononuclear cells of a 22-year-old Japanese male patient. The donor was initially diagnosed with T-lymphoblastic lymphoma, which subsequently underwent a malignant leukemic conversion into Acute Myeloid Leukemia (AML M4 subtype).

  • Clonal Origin: TK-1B represents an authentic single-cell clone isolated from its parent cell line, TK-1, via the limited dilution method.

• Lineage & Molecular Characteristics:

  • Myelomonocytic Signature: TK-1B preserves myelomonocytic phenotypic traits that are highly analogous to the primary parental line.

  • EBV-Negative Profile: Molecular validation confirmed that these cloned cells lack Epstein-Barr virus nuclear antigens (EBNA), confirming a virus-free endogenous transformation etiology.

• Immunophenotypic Profile:

  • Positive Cell Surface Markers: CD4+, CD7+, CD10+, CD13+, CD15+, CD33+, and HLA-DR+.

  • Negative Cell Surface Markers: CD2-, CD3-, CD5-, CD8-, CD14-, CD19-, CD20-, and CD34-.

• Biosafety Level: BSL-1.Certified negative via multiplex PCR assays for EBV, HBV, HCV, HIV-1/2, HTLV-1/2, and MLV viral genomes.

II Morphological Attributes and Cultivation Media

• Morphology: Displays characteristic features of acute myelomonocytic leukemia blastic variants. Under standard microscopic evaluation, the culture presents as spherical individual suspension cells growing cleanly singly, with a minor tendency to aggregate into loose, small suspension clumps.

• Growth Mode: Strict suspension growth.

• Population Doubling Time: Approximately 48 to 72 hours, reflecting a moderate expansion velocity.

• Standard Complete Growth Medium Formulation:

  • Basal Medium: 80% to 90% BioVector? RPMI-1640 medium.

  • Routine Supplements: 10% to 20% premium BioVector? Heat-Inactivated Fetal Bovine Serum (h.i. FBS).

• Physical Incubation Parameters: Regulated strictly at 37 degrees Celsius under a humidified atmosphere containing 5% Carbon Dioxide.

III Subculturing and Thawing Protocols

1. Initial Seeding and Delayed Post-Thaw Recovery Dynamics:

   • First-Week Lag Phase Warning: TK-1B cells display critical sensitivity to physical cryorecovery stresses.In routine thawing procedures, no measurable cell proliferation or growth occurs during the first week of culture (no cell growth during first week). This is an intrinsic biological lag phase characteristic of the clone; cultures must be left undisturbed without aggressive media handling during this adaptation window.

   • High-Density Initiation Protocol: When reviving a cryovial or starting fresh cultures, cells must be initiated with elevated serum support and strict volume restriction. Seed the cell suspension out at a minimum high-density of approximately 1000000 viable cells/mL (1 x 10^6 cells/ml) inside a standard 24-well plate setup utilizing 20% FBS complete medium.

2. Routine Continuous Passaging Schedule:

   • Once the culture successfully transits the initial lag phase and establishes stable logarithmic kinetics, enter routine handling.

   • Target Cultivation Density Window: Maintain the active expanding suspension cell profile strictly between a baseline floor of 500000 and an upper ceiling of 1000000 viable cells/mL (0.5-1 x 10^6 cells/ml).

   • Passaging Execution: Subculture or split the vessel when total suspension density approaches the maximum harvest concentration of 1500000 cells/mL (1.5 x 10^6 cells/ml).Because cells grow strictly unattached, pellet the biomass by spinning at 150 g for 5 minutes, decant the spent supernatant, and redistribute in fresh complete medium at recommended split sequences of 1:2 to 1:3 every 2 to 3 days.

IV Strategic Research Applications

1. Investigating Lineage Switch and Malignant Lymphoma-to-AML Conversion: Since BioVector? TK-1B originates from an clinically rare presentation of a T-lymphoblastic lymphoma converting into an aggressive acute myeloid leukemia, this clone serves as a valuable tool to explore hematopoietic clonal evolution, multiclonal chemotherapy escape, and the transcriptomic plasticity driving lineage switches.

2. Targeting Myelomonocytic Epitopes and Differentiation Kinetics: The stable presentation of CD13 and CD33 alongside a negative CD14 status makes this cell line highly useful for investigating target engagement profiles for myeloid-directed antibody-drug conjugates (ADCs). It is also employed to track the transcriptional blockades in acute myelomonocytic leukemia (AML-M4) under treatment with epigenetic modifiers or targeted differentiation-inducing small molecules.

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