MYC 1-9E10.2 BioVector®杂交瘤细胞株9E10 Hybridoma Cell Line

BioVector® MYC 1-9E10.2 杂交瘤细胞株文本版说明书

BioVector® MYC 1-9E10.2 Hybridoma Cell Line Text-Based Datasheet

1 产品基本信息与遗传背景 / Product Identification and Genetic Background

• 产品名称 (Product Name)：BioVector® MYC 1-9E10.2 杂交瘤细胞株 (BioVector® MYC 1-9E10.2 Hybridoma Cell Line)

• 常用别名 (Synonyms)：BioVector® 1-9E10.2，BioVector® 9E10，BioVector® Anti-Myc Hybridoma

• 生物学来源 (Organism Source)：体细胞杂交株（小鼠 Mus musculus 免疫脾细胞 融合 小鼠 Sp2/0-Ag14 骨髓瘤细胞）

• 组织器官定位 (Tissue and Organ Site)：免疫系统 / 杂交瘤悬浮细胞 (Immune System / Suspension Hybridoma)

• 生长特性 (Growth Properties)：悬浮生长，细胞呈圆形，常以单个细胞或松散的小细胞团块形式在培养基中悬浮。(Suspension growth, cells present as spherical entries suspended in the medium either individually or in loose mini-clusters.)

• 杂交永生化机制 (Immortalization Mechanism)：利用聚乙二醇（PEG）技术，将经过人类 c-Myc 蛋白特异性合成肽片段免疫的小鼠原代 B 淋巴细胞（脾细胞），与具有永生化不死性的 Sp2/0-Ag14 小鼠骨髓瘤细胞进行体细胞杂交融合，通过 HAT 培养基筛选与多次有限稀释克隆化，最终获得高效稳定分泌单克隆抗体的细胞系。(Engineered via Polyethylene Glycol mediated somatic fusion between primary mouse splenocytes immunized with a synthetic peptide corresponding to the human c-Myc protein sequence and immortal Sp2/0-Ag14 myeloma cells, selected via HAT screening and single-cell cloning to establish a stable, antibody-secreting hybridoma line.)

• 核心科研价值 (Core Research Significance)：BioVector® MYC 1-9E10.2 是全球分子生物学、蛋白质组学与生物工程领域极其重要的基础工具株。该细胞系专门用于体外大规模生产和分泌高特异性的 anti-Myc 单克隆抗体（克隆号 9E10）。该抗体能极其精准地识别重组蛋白中的 Myc 标签（Myc-tag，氨基酸序列为 EQKLISEEDL）。它被全球实验室广泛用于工程蛋白的免疫印迹（WB）、免疫沉淀（IP）、免疫组化（IHC）、染色质免疫沉淀（ChIP）以及亲和层析纯化，是现代重组蛋白表达检测产业的核心底盘。(BioVector® MYC 1-9E10.2 stands as a globally pivotal tool line in molecular biology, proteomics, and biotechnology. This specific hybridoma is dedicated to the large-scale in vitro manufacture and secretion of the anti-Myc monoclonal antibody (Clone 9E10). This antibody tracks the peptide sequence EQKLISEEDL defining the human c-Myc epitope tag. It is widely utilized across international core labs for Western Blotting, Immunoprecipitation, and affinity chromatography purification of recombinant proteins.)

2 细胞生物学特征与核心产物表型 / Cellular Properties and Product Profiles

BioVector® MYC 1-9E10.2 作为标准的杂交瘤分泌细胞，具备高效的抗体表达放大效应：The BioVector® MYC 1-9E10.2 line operates as a standard manufacturing hybridoma, unlocking high-efficiency antibody upregulation pathways:

悬浮生长状态 (Suspension Proliferation State)

细胞不依赖任何基质包被。在常规悬浮或转瓶培养中，细胞维持着活跃的非贴壁有丝分裂，倍增时间通常在 22 到 28 小时之间。Cells proliferate independently of matrix anchoring. Under standard suspension or roller-bottle tracks, entries maintain active non-adherent mitosis, yielding doubling times between 22 and 28 hours.

抗体分泌特征 (Antibody Secretion Profile)

细胞在对数生长期与平台期会源源不断地向胞外培养基中分泌单克隆抗体，其物理与生物学功能特征包括：The culture continuously secretes monoclonal antibodies into the extracellular media during logarithmic and stationary frames. Its biological attributes include:

• 抗体同种型 (Antibody Isotype)：小鼠免疫球蛋白 G1 亚型（Mouse IgG1, kappa 链）。(Classified as Mouse Immunoglobulin G1, kappa light chain.)

• 靶向特异性 (Target Epitope)：精准靶向人 c-Myc 蛋白第 408 到第 439 位氨基酸区段（即由 EQKLISEEDL 组成的连续表位），对其他不相关蛋白质序列完全无交叉反应。(Exhibits absolute specificity toward the human c-Myc domain mapping amino acids 408 to 439, showing zero non-specific cross-reactivity with adjacent cellular grids.)

• 产量表现 (Yield Optimization)：在常规转瓶或高密度生物反应器培养下，上清液中的抗体效价高，纯化回收率优异。(Delivers excellent raw antibody titers within standard culture supernatants or bioreactor systems, ensuring superb downstream yield extraction.)

3 专用培养基配方规范 / Culturing Medium Formulations

警告 / Critical WarningBioVector® MYC 1-9E10.2 作为杂交瘤细胞，对培养基中的血清质量以及补剂浓度极其敏感。日常扩增时必须避免细胞密度过低（低于 1比10 的细胞初始密度），否则细胞会迅速失去自分泌因子的支持并引发大规模自溶性死亡。BioVector® MYC 1-9E10.2 hybridomas are exceptionally sensitive to serum quality and trace component density. During routine expansion, never seed below minimal viability thresholds (e.g., seeding below 100,000 viable cells per mL), as the absence of autocrine reinforcement loops will provoke widespread autolytic death.

A 阶段：常规常规扩增完全培养基（用于常规扩增与维持）

Phase A: Proliferating Complete Growth Medium (For Routine Expansion and Maintenance)

• 基础培养基 (Basal Medium)：BioVector® High-Glucose DMEM 高糖培养基（含 4.5 g/L 葡萄糖，含 L-谷氨酰胺，含丙酮酸钠 / containing 4.5 grams per liter Glucose, with L-Glutamine and sodium pyruvate）或选用 BioVector® RPMI-1640 培养基。

• 血清添加 (Serum Supplement)：10% 到 15% BioVector® Premium Fetal Bovine Serum 优质胎牛血清（新复苏时建议使用 15% 比例以增强细胞修复能力 / 15% concentration is highly recommended post-thaw to maximize cell recovery profiles）。

• 双抗补剂 (Antibiotics)：1% BioVector® Penicillin-Streptomycin Solution 青霉素-链霉素溶液。

B 阶段：高产抗体收集培养基（用于无血清或低血清大规模抗体收集）

Phase B: Antibody Production and Collection Medium (For Serum-Free Multi-Batch Harvesting)

• 基础骨架 (Basal Matrix)：BioVector® Hybridoma-SFM 杂交瘤专用无血清培养基（或者将常规培养基中的血清比例降至 1% 到 2% 的低内毒素血清，以便于后期的亲和柱纯化层析）。(BioVector® Hybridoma-SFM Serum-Free Medium, or alternative low-serum variants drop-scaled to 1% to 2% to simplify downstream Protein A/G affinity purification processing.)

4 物理环境与悬浮空间控制参数 / Environmental Controls and Handling Parameters

• 工作温度 (Incubation Temperature)：37.0 摄氏度（Constant temperature at 37.0 degrees Celsius）。

• 气相环境 (Gas Phase)：5% 二氧化碳 (CO2)，加湿平衡空气 (5% Carbon Dioxide balanced with humidified atmospheric air)。

• 剪切力控制与容器规范 (Vessel and Agitation Parameters)：

  • 常规扩增：细胞在未包被的普通 T25 或 T75 静置培养瓶中悬浮生长即可，培养瓶应保持水平静置，切勿随意摇晃。(For routine expansion, maintain cells horizontally within standard, uncoated T-flasks under completely static tracks.)

  • 规模化生产：在进行抗体大规模收集时，可将细胞转入转瓶（Roller Bottle）或摇瓶（Erlenshaker）中，搅拌速度或振荡速度应严格控制在每分钟 40 到 60 转（40 到 60 rpm）的低剪切力范围内。转速过快会导致悬浮杂交瘤的细胞膜发生物理撕裂并大面积死亡。 (For upscale production, cultures can be moved into roller bottles or shaking bioreactors, with mechanical speeds strictly limited to 40 to 60 rpm. Excess shear stress will rupture the hybridoma membranes, triggering severe culture crashing.)

5 细胞传代与抗体收集操作规范 / Subculturing and Antibody Harvesting Protocols

悬浮期常规传代操作 / Routine Passaging (Suspension State)

• 传代临界点 (Passaging Threshold)：当悬浮细胞密度达到 1.0乘以10的6次方 cells/mL 时必须进行传代。由于杂交瘤细胞分裂代谢极为旺盛，一旦细胞密度超过 1.5乘以10的6次方 cells/mL，培养基会迅速变黄并累积大量有毒代谢废物（如乳酸），导致细胞活力发生断崖式下跌。(Passage promptly when suspension density hits 1.0 x 10^6 cells/mL. Because cell metabolic pathways operate intensely, overgrowing past 1.5 x 10^6 cells/mL will exhaust carbon feeds, accumulate lactic toxins, and cause viability metrics to plunge.)

• 建议传代稀释比例 (Splitting Ratio)：通常采取半量换液或直接稀释法，维持接种密度在 2.0乘以10的5次方 cells/mL 左右，每 2 到 3 天传代一次。

1. 直接从孵箱中取出培养瓶，轻轻摇匀，使其形成分布均匀的细胞悬液（由于是完全悬浮细胞，传代过程绝对不需要使用胰酶进行消化）。(Retrieve the flask and agitate gently to create an even suspension. Because entries grow in absolute suspension, trypsin enzymes must never be applied during passaging.)

2. 用无菌移液管吸出全部细胞悬液，转入 15 mL 离心管中。

3. 在 200乘以g（低速离心力）下离心 5 分钟，彻底泵干变黄的陈旧培养基上清。(Centrifuge at 200 x g for 5 minutes and thoroughly vacuum away the spent yellowing supernatant.)

4. 加入 10 mL 新鲜预热的 BioVector® 增殖期完全培养基，用移液管轻柔吹打沉淀，配置成均匀悬液。(Introduce 10 mL of pre-warmed Phase A growth medium and pipette gently to release the hybridoma pellet.)

5. 按照 1比4 或 1比5 的稀释比例，将细胞分装接种至全新的培养瓶中。(Distribute the suspension into fresh vessels at a 1:4 or 1:5 dilution ratio to maintain correct seeding limits.)

大规模抗体收集纯化流程 / Bulk Antibody Harvesting Protocol

1. 将处于对数生长期的 BioVector® MYC 1-9E10.2 细胞扩大培养，并逐步将其调整接种至 BioVector® 杂交瘤专用无血清培养基 中，起始接种密度设定为 3.0乘以10的5次方 cells/mL。(Expand log-phase cultures and step-adapt them into BioVector® Hybridoma-SFM, with an initial seeding density targeting 3.0 x 10^5 cells/mL.)

2. 置于大容量转瓶或低速摇床中持续培养，让其自然生长并分泌抗体。(Maintain within large-volume roller bottles or low-shear shaker platforms to allow continuous execution of antibody secreting pathways.)

3. 密切监测培养基颜色。当细胞密度达到顶峰并开始进入衰退期（通常在接种后 5 到 7 天，镜下观察活细胞率降至 40% 左右时，此时胞内抗体释放最彻底），即可终止培养。(Monitor background metrics closely; harvest when the culture transits into decline, typically 5 to 7 days post-seeding, when cell viability drops toward 40%, signaling optimal antibody release into the broth.)

4. 将全部培养液收集进大容量离心管中，在 1000乘以g 的离心力下高速离心 15 分钟，彻底沉淀细胞碎片，小心吸出并保留澄清的上清液。(Harvest the total pool, centrifuge at 1000 x g for 15 minutes to clear cellular debris, and collect the clarified supernatant fluid.)

5. 将澄清上清液通过 0.22 微米的无菌无纺滤膜进行过滤，随后可直接输入装有 BioVector® Protein A 或 Protein G 的亲和层析柱中进行单克隆抗体纯化洗脱。将纯化后的 9E10 抗体进行透析脱盐，即可获得高纯度的工程级 Anti-Myc 单克隆抗体。(Pass the fluid through a 0.22-micrometer membrane filter, then load onto a BioVector® Protein A/G affinity column for monoclonal antibody purification, yields high-purity, research-grade 9E10 anti-Myc antibodies post-dialysis.)

6 复苏与冻存恢复流线 / Thawing and Post-Cryo Recovery Workflow

1. 提前在标准 T25 培养瓶中注入 6.0 mL 预热的 BioVector® 增殖期完全生长培养基（含 15% 优质血清），置于孵箱中平衡气体与温控。(Pre-warm 6.0 mL of Phase A growth medium fortified with 15% premium serum in a T25 flask, equilibrating inside the incubator.)

2. 从液氮中取出 BioVector® MYC 1-9E10.2 冻存管，立刻全量投入 37 摄氏度恒温水浴箱中快速水平摇晃，在 60 秒内使其快速完全融化。(Retrieve the cryovial from liquid nitrogen and plunge it into a 37 degrees Celsius water bath, agitating horizontally to complete the melting sequence within 60秒.)

3. 酒精表面功能消毒后移入超净台，用移液枪吸出胞悬液，缓慢逐滴注入盛有 4.0 mL 预热增殖培养基的 15 mL 离心管中。(Sanitize with 75% ethanol and dropwise introduce the suspension into a conical tube containing 4.0 mL of pre-warmed medium.)

4. 在 200乘以g 下低速离心 5 分钟，彻底吸干含有毒 DMSO 的上清液。(Centrifuge at 200 x g for 5 minutes and thoroughly vacuum away the toxic DMSO-laden supernatant.)

5. 加入 2.0 mL 新鲜的 15% 血清培养基轻弹悬起细胞沉淀，随后全量接种到预热平衡的 T25 瓶中。(Add 2.0 mL of fresh 15% serum medium, tap gently to scatter the pellet, and transfer into the pre-stabilized T25 flask.)

6. 复苏后 24 小时必须在显微镜下进行活细胞计数并补充 2 到 3 mL 新鲜培养基，以稀释复苏死亡细胞释放的毒性碎片。(Inspect 24 hours post-thaw; perform a viable cell count and top up with 2 to 3 mL of fresh medium to dilute toxicity from dead cell fragments.)

7 生物安全、长期保存与质控规范 / Biosafety, Storage and Quality Controls

• 生物安全级别 (Biosafety Level)：BSL-1。属于安全的常规小鼠杂交瘤细胞系，无已知人类病原体传染风险，实验废弃物按标准生物医疗废弃物高压灭菌消杀即可。(BSL-1 classification. Represents a secure murine hybridoma matrix with no known pathogenic viral threats to human safety; process waste items via standard biological autoclave disinfection paths.)

• 超低温长期保存 (Long-Term Banking)：冻存保质配方为 90% BioVector® 增殖期完全生长培养基 + 10% 细胞级 DMSO（或采用 BioVector® Premium 无血清高效冻存液）。冻存管必须永久存放于液氮罐气相或液相中（零下 150 摄氏度至零下 196 摄氏度）。严禁在零下 80 摄氏度普通机械冰箱内长期搁置超过 1 个月，否则会发生抗体表达质粒丢失或复苏存活率断崖式暴跌。(The freezing blend consists of 90% Phase A medium + 10% analytical grade DMSO, or BioVector® Premium Serum-Free Cryopreservation Medium. Vials must reside permanently inside liquid nitrogen repositories (minus 150 to minus 196 degrees Celsius). Storage in mechanical minus 80 degrees Celsius freezers for extended spans is strictly prohibited, as it compromises antibody plasmid stability and survival kinetics.)

• 抗体分泌效价与特异性核心质控红线 / Antibody Secretion Quality Control Threshold：随着传代次数的增加（连续传代超过 35 到 40 代以上），杂交瘤细胞极易发生非整倍体染色体丢失、基因重排或非分泌型克隆逃逸。严格质控表现为：每隔 10 代，应收集一次细胞无血清培养上清，通过 ELISA 测试或常规 Western Blot 测试验证其对标准 Myc 标签重组蛋白（如带有 Myc-tag 的 GFP 蛋白）的结合能力。如果测得的上清液抗体效价出现数量级下降，或者出现非特异性杂带，证实该杂交瘤细胞已经发生功能退化。必须立即停止该代次生产，全盘销毁细胞，并重新复苏超低代次的种子库进行恢复。(Extended passaging (exceeding 35 to 40 continuous subcultures) can trigger chromosomal dropouts, gene rearrangements, or non-secreting clonal drift in hybridomas. For functional quality control, sample the serum-free supernatant every 10 passages; evaluate binding kinetics toward a standard Myc-tagged recombinant protein via ELISA or Western Blotting. If the recorded antibody titer falls by an order of magnitude or exhibits non-specific artifact bands, the lineage has degraded. Cease manufacturing chains immediately, discard active flasks, and re-thaw a lower-passage master vial from the cryo-vault.)

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