BioVector® YM 人弥漫大B细胞淋巴瘤细胞株

BioVector® YM Human Diffuse Large B-Cell Lymphoma Cell Line

第一部分 中文说明

一 产品基本信息与遗传学背景

• 细胞名称：BioVector® YM

• 保藏机构货号：DSMZ ACC 948

• 物种来源：人类 (Homo sapiens)

• 性别与年龄：男性，61岁

• 组织与疾病背景：该细胞株建立于1993年，分离自一名患有弥漫大B细胞淋巴瘤（Diffuse Large B-Cell Lymphoma, DLBCL）的61岁男性患者的腹水（Ascites fluid）组织。

• 核心致癌分子与双打击（Double-Hit）特征：

  • 双打击淋巴瘤（Double-hit lymphoma）染色体特征：该细胞株被明确定义为携带经典双打击易位重排的恶性B细胞系，同时携带染色体易位 t(2;18)(p11;q21) 和 t(3;16)(q27;p11)。

  • IL21R::BCL6 融合基因表达：经 RT-PCR 分子生物学技术验证，该细胞株内源性特异表达 IL21R::BCL6 原癌融合基因，是驱动该淋巴瘤克隆恶性转化的核心病理因子之一。

• 免疫表型谱（Immunophenotype）：

  • 阳性表面标记：CD19、CD20、CD37、CD38、CD80、HLA-DR、sIgM、sIg-kappa。

  • 阴性表面标记：CD3、CD10、CD13、CD34、CD138。

• 生物安全级别：1级（BSL-1）。经检测常见病毒如 EBV, HBV, HCV, HIV-1/2, HTLV-1/2 以及 MLV 均为阴性。

二 细胞形态学与培养环境

• 形态学特征：展现典型的恶性B细胞淋巴瘤形态特征。在倒置显微镜下，细胞呈微小的单细胞或松散成团的悬浮颗粒簇形式存在。

• 生长模式：完全悬浮生长。

• 群体倍增时间（Doubling Time）：大约 48 小时左右。

• 标准完全培养基配方：

  • 基础培养基：80% 至 90% BioVector® RPMI-1640 培养基。

  • 维持添加：10% 至 20% 优质热灭活胎牛血清（h.i. FBS）。

• 物理培养参数：37摄氏度恒温、5% 二氧化碳、空气饱和湿度。

三 细胞传代与复苏标准操作步骤

1. 初始接种与常规传代操作（Subculturing Protocol）：

   • 起始接种密度：当首次从冻存管复苏开启本细胞时，请务必将其初始接种密度设定在每毫升约 500000 个活细胞，并建议置于 24 孔板等小限制性体积器皿中以富集自分泌因子。

   • 日常维持密度窗口：在日常维持和扩增循环中，应将悬液活细胞密度严密控制在 每毫升 700000 至 1000000 个活细胞 之间。

   • 传代执行：当细胞收集密度达到每毫升 1000000 至 1500000 个的饱和峰值时需要进行分裂传代。由于是完全悬浮生长，无需胰酶消化。直接通过离心收集细胞并丢弃旧基，使用新鲜完全培养基重悬，常规传代比例为 1比2 至 1比3，通常每 2 到 3 天常规处理一次。

2. 冻存细胞复苏：

   • 快速将冻存管自液氮中取出，移入 37 摄氏度 BioVector® 水浴锅中持续摇晃，在 1 到 2 分钟内令其极速融化。

   • 将解冻的细胞浆液缓慢移入含有 5 毫升 预热高血清完全培养基的离心管中，以 150 g 离心 5 分钟。

   • 彻底吸除上清以去除残留的 DMSO 冻存液，使用新鲜的高血清完全培养基重悬，并按照上述推荐的高密度启动参数接种培养。

四 核心科研应用方向

1. 双打击淋巴瘤（DHL）分子转录调控与发病机制研究：BioVector® YM 是国际上研究携带 t(2;18) 和 t(3;16) 染色体易位的双打击淋巴瘤非常罕见的内源性标准靶向模型。主要用于阐明 IL21R::BCL6 融合基因对生发中心 B 细胞发育分化控制网络的压制机制。

2. 小分子靶向阻断剂/降解剂的药效动力学评价：由于该细胞高表达由易位产生的 BCL6 融合功能体，它是用于高通量筛选新型 BCL6 小分子抑制剂、BCL6 靶向嵌合体降解剂以及联合抑制阻断剂药效评估的理想体外底物。

3. 靶向 B 细胞表位免疫疗法（CAR-T/CAR-NK/双特异性抗体）效能测试：该细胞表面稳定且高丰度地表达 B 细胞系成熟分化抗原 CD19, CD20, CD37 及 HLA-DR。可广泛用于作为肿瘤靶细胞，在体外细胞毒性释放测定中用于评估和评价全新一代抗 CD19/CD20 双抗、抗体偶联药物以及 CAR-T、CAR-NK 细胞的靶向溶瘤能力。

PART 2 ENGLISH SECTION

I General Information and Genetic Background

• Cell Line Name: BioVector® YM

• Repository Catalog Number: DSMZ ACC 948

• Species Origin: Human (Homo sapiens)

• Sex and Age of Donor: Male, 61 years old

• Tissue and Disease Background: Established in 1993 from the malignant ascites fluid of a 61-year-old male patient diagnosed with Diffuse Large B-Cell Lymphoma (DLBCL).

• Core Oncogenic Markers & "Double-Hit" Traits:

  • Double-Hit Cytogenetic Recombination: Characterized as an authentic double-hit lymphoma model carrying concomitant chromosomal translocations t(2;18)(p11;q21) and t(3;16)(q27;p11).

  • IL21R::BCL6 Fusion Transcript: Molecular integration and persistent expression of the pathognomonic IL21R::BCL6 fusion gene was structurally confirmed via RT-PCR assays.

• Immunophenotypic Profile:

  • Positive Cell Surface Markers: CD19, CD20, CD37, CD38, CD80, HLA-DR, sIgM, and sIg-kappa.

  • Negative Cell Surface Markers: CD3, CD10, CD13, CD34, and CD138.

• Biosafety Level: BSL-1. Validated negative via targeted PCR tracking 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 malignant lymphoblastoid variants. Under standard inverted microscope setups, cells expand as small single cells or loose unattached cluster matrices.

• Growth Mode: Strict suspension growth.

• Population Doubling Time: Approximately 48 hours.

• 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 an atmosphere of 5% Carbon Dioxide with saturated air humidity.

III Subculturing and Thawing Protocols

1. Initial Seeding Setup and Routine Passaging Schedule:

   • Seeding Initialization: When opening a fresh cryovial or working with heavy dilutions, always seed the culture out at approximately 500000 viable cells/mL, preferentially inside a 24-well plate layout to maximize critical paracrine support signals.

   • Core Maintenance Density Window: During routine culture maintenance, keep the active expanding cell slurry tightly restricted between 700000 and 1000000 cells/mL.

   • Subculturing Routine: Process the vessel when the active mass density approaches the saturation ceiling of 1000000 to 1500000 cells/mL. Because it expands purely in suspension, enzymatic trypsinization is entirely unnecessary. Pellet the cells by spinning at 150 g for 5 minutes, decant the spent media, and redistribute in fresh complete medium at recommended split sequences averaging 1:2 to 1:3 every 2 to 3 days.

2. Cryovial Thawing and Recovery:

   • Re-trieve the cryovial from cryogenic storage and plunge it into a 37 degrees Celsius BioVector® water bath with continuous rapid agitation, achieving complete liquefaction within 1 to 2 minutes.

   • Transfer the cell suspension slowly into a sterile centrifuge tube carrying 5 mL of pre-warmed complete growth medium and spin at 150 g for 5 minutes.

   • Decant the supernatant to clear out toxic residual DMSO, and gently resuspend the cell pellet in fresh complete growth medium, establishing the high baseline seeding configuration of around 500000 cells/mL.

IV Strategic Research Applications

1. Elucidating Double-Hit Lymphoma (DHL) Oncogenesis: BioVector® YM serves as a valuable, scarce endogenous line co-harboring reciprocal t(2;18) and t(3;16) genetic alterations. It is utilized to map transcription disruptions driven by IL21R::BCL6 fusions that arrest somatic B-cell terminal maturation pathways.

2. Screening Targeted Small-Molecule BCL6 Inhibitors / Degraders: Given its structural presentation of a mutated, fusion-activated BCL6 transcript, this line acts as a specific screening system to discover small-molecule BCL6 inhibitors, assess targeted chimeric degraders, or test therapeutic synergy profiles.

3. Validating Anti-B-Cell Modern Immunotherapies (CAR-T, CAR-NK, or Bi-specific Platforms): Because YM cells stably present a series of mature B-lineage antigen determinants including CD19, CD20, CD37, and HLA-DR on their outer cell membranes, they function as optimized targeted cells in standard cytotoxicity assays. They are routinely deployed to track specific effector engagement and oncolytic efficiencies of multi-specific monoclonal antibodies, Antibody-Drug Conjugates, and engineered CAR-T or CAR-NK products.

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