BioVector? MYC 1-9E10.2 雜交瘤細胞株文本版說明書

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

1 產(chǎn)品基本信息與遺傳背景 / Product Identification and Genetic Background

• 產(chǎn)品名稱 (Product Name)：BioVector? MYC 1-9E10.2 雜交瘤細胞株 (BioVector? MYC 1-9E10.2 Hybridoma Cell Line)

• 常用別名 (Synonyms)：BioVector? 1-9E10.2，BioVector? 9E10，BioVector? Anti-Myc Hybridoma

• 生物學來源 (Organism Source)：體細胞雜交株（小鼠 Mus musculus 免疫脾細胞 融合 小鼠 Sp2/0-Ag14 骨髓瘤細胞）

• 組織器官定位 (Tissue and Organ Site)：免疫系統(tǒng) / 雜交瘤懸浮細胞 (Immune System / Suspension Hybridoma)

• 生長特性 (Growth Properties)：懸浮生長，細胞呈圓形，常以單個細胞或松散的小細胞團塊形式在培養(yǎng)基中懸浮。(Suspension growth, cells present as spherical entries suspended in the medium either individually or in loose mini-clusters.)

• 雜交永生化機制 (Immortalization Mechanism)：利用聚乙二醇（PEG）技術，將經(jīng)過人類 c-Myc 蛋白特異性合成肽片段免疫的小鼠原代 B 淋巴細胞（脾細胞），與具有永生化不死性的 Sp2/0-Ag14 小鼠骨髓瘤細胞進行體細胞雜交融合，通過 HAT 培養(yǎng)基篩選與多次有限稀釋克隆化，最終獲得高效穩(wěn)定分泌單克隆抗體的細胞系。(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.)

• 核心科研價值 (Core Research Significance)：BioVector? MYC 1-9E10.2 是全球分子生物學、蛋白質(zhì)組學與生物工程領域極其重要的基礎工具株。該細胞系專門用于體外大規(guī)模生產(chǎn)和分泌高特異性的 anti-Myc 單克隆抗體（克隆號 9E10）。該抗體能極其精準地識別重組蛋白中的 Myc 標簽（Myc-tag，氨基酸序列為 EQKLISEEDL）。它被全球?qū)嶒炇覐V泛用于工程蛋白的免疫印跡（WB）、免疫沉淀（IP）、免疫組化（IHC）、染色質(zhì)免疫沉淀（ChIP）以及親和層析純化，是現(xiàn)代重組蛋白表達檢測產(chǎn)業(yè)的核心底盤。(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 細胞生物學特征與核心產(chǎn)物表型 / Cellular Properties and Product Profiles

BioVector? MYC 1-9E10.2 作為標準的雜交瘤分泌細胞，具備高效的抗體表達放大效應：The BioVector? MYC 1-9E10.2 line operates as a standard manufacturing hybridoma, unlocking high-efficiency antibody upregulation pathways:

懸浮生長狀態(tài) (Suspension Proliferation State)

細胞不依賴任何基質(zhì)包被。在常規(guī)懸浮或轉(zhuǎn)瓶培養(yǎng)中，細胞維持著活躍的非貼壁有絲分裂，倍增時間通常在 22 到 28 小時之間。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.

抗體分泌特征 (Antibody Secretion Profile)

細胞在對數(shù)生長期與平臺期會源源不斷地向胞外培養(yǎng)基中分泌單克隆抗體，其物理與生物學功能特征包括：The culture continuously secretes monoclonal antibodies into the extracellular media during logarithmic and stationary frames. Its biological attributes include:

• 抗體同種型 (Antibody Isotype)：小鼠免疫球蛋白 G1 亞型（Mouse IgG1, kappa 鏈）。(Classified as Mouse Immunoglobulin G1, kappa light chain.)

• 靶向特異性 (Target Epitope)：精準靶向人 c-Myc 蛋白第 408 到第 439 位氨基酸區(qū)段（即由 EQKLISEEDL 組成的連續(xù)表位），對其他不相關蛋白質(zhì)序列完全無交叉反應。(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.)

• 產(chǎn)量表現(xiàn) (Yield Optimization)：在常規(guī)轉(zhuǎn)瓶或高密度生物反應器培養(yǎng)下，上清液中的抗體效價高，純化回收率優(yōu)異。(Delivers excellent raw antibody titers within standard culture supernatants or bioreactor systems, ensuring superb downstream yield extraction.)

3 專用培養(yǎng)基配方規(guī)范 / Culturing Medium Formulations

警告 / Critical WarningBioVector? MYC 1-9E10.2 作為雜交瘤細胞，對培養(yǎng)基中的血清質(zhì)量以及補劑濃度極其敏感。日常擴增時必須避免細胞密度過低（低于 1比10 的細胞初始密度），否則細胞會迅速失去自分泌因子的支持并引發(fā)大規(guī)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 階段：常規(guī)常規(guī)擴增完全培養(yǎng)基（用于常規(guī)擴增與維持）

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

• 基礎培養(yǎng)基 (Basal Medium)：BioVector? High-Glucose DMEM 高糖培養(yǎng)基（含 4.5 g/L 葡萄糖，含 L-谷氨酰胺，含丙酮酸鈉 / containing 4.5 grams per liter Glucose, with L-Glutamine and sodium pyruvate）或選用 BioVector? RPMI-1640 培養(yǎng)基。

• 血清添加 (Serum Supplement)：10% 到 15% BioVector? Premium Fetal Bovine Serum 優(yōu)質(zhì)胎牛血清（新復蘇時建議使用 15% 比例以增強細胞修復能力 / 15% concentration is highly recommended post-thaw to maximize cell recovery profiles）。

• 雙抗補劑 (Antibiotics)：1% BioVector? Penicillin-Streptomycin Solution 青霉素-鏈霉素溶液。

B 階段：高產(chǎn)抗體收集培養(yǎng)基（用于無血清或低血清大規(guī)模抗體收集）

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

• 基礎骨架 (Basal Matrix)：BioVector? Hybridoma-SFM 雜交瘤專用無血清培養(yǎng)基（或者將常規(guī)培養(yǎng)基中的血清比例降至 1% 到 2% 的低內(nèi)毒素血清，以便于后期的親和柱純化層析）。(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 物理環(huán)境與懸浮空間控制參數(shù) / Environmental Controls and Handling Parameters

• 工作溫度 (Incubation Temperature)：37.0 攝氏度（Constant temperature at 37.0 degrees Celsius）。

• 氣相環(huán)境 (Gas Phase)：5% 二氧化碳 (CO2)，加濕平衡空氣 (5% Carbon Dioxide balanced with humidified atmospheric air)。

• 剪切力控制與容器規(guī)范 (Vessel and Agitation Parameters)：

  • 常規(guī)擴增：細胞在未包被的普通 T25 或 T75 靜置培養(yǎng)瓶中懸浮生長即可，培養(yǎng)瓶應保持水平靜置，切勿隨意搖晃。(For routine expansion, maintain cells horizontally within standard, uncoated T-flasks under completely static tracks.)

  • 規(guī)?；a(chǎn)：在進行抗體大規(guī)模收集時，可將細胞轉(zhuǎn)入轉(zhuǎn)瓶（Roller Bottle）或搖瓶（Erlenshaker）中，攪拌速度或振蕩速度應嚴格控制在每分鐘 40 到 60 轉(zhuǎn)（40 到 60 rpm）的低剪切力范圍內(nèi)。轉(zhuǎn)速過快會導致懸浮雜交瘤的細胞膜發(fā)生物理撕裂并大面積死亡。 (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 細胞傳代與抗體收集操作規(guī)范 / Subculturing and Antibody Harvesting Protocols

懸浮期常規(guī)傳代操作 / Routine Passaging (Suspension State)

• 傳代臨界點 (Passaging Threshold)：當懸浮細胞密度達到 1.0乘以10的6次方 cells/mL 時必須進行傳代。由于雜交瘤細胞分裂代謝極為旺盛，一旦細胞密度超過 1.5乘以10的6次方 cells/mL，培養(yǎng)基會迅速變黃并累積大量有毒代謝廢物（如乳酸），導致細胞活力發(fā)生斷崖式下跌。(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.)

• 建議傳代稀釋比例 (Splitting Ratio)：通常采取半量換液或直接稀釋法，維持接種密度在 2.0乘以10的5次方 cells/mL 左右，每 2 到 3 天傳代一次。

1. 直接從孵箱中取出培養(yǎng)瓶，輕輕搖勻，使其形成分布均勻的細胞懸液（由于是完全懸浮細胞，傳代過程絕對不需要使用胰酶進行消化）。(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. 用無菌移液管吸出全部細胞懸液，轉(zhuǎn)入 15 mL 離心管中。

3. 在 200乘以g（低速離心力）下離心 5 分鐘，徹底泵干變黃的陳舊培養(yǎng)基上清。(Centrifuge at 200 x g for 5 minutes and thoroughly vacuum away the spent yellowing supernatant.)

4. 加入 10 mL 新鮮預熱的 BioVector? 增殖期完全培養(yǎng)基，用移液管輕柔吹打沉淀，配置成均勻懸液。(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 的稀釋比例，將細胞分裝接種至全新的培養(yǎng)瓶中。(Distribute the suspension into fresh vessels at a 1:4 or 1:5 dilution ratio to maintain correct seeding limits.)

大規(guī)?？贵w收集純化流程 / Bulk Antibody Harvesting Protocol

1. 將處于對數(shù)生長期的 BioVector? MYC 1-9E10.2 細胞擴大培養(yǎng)，并逐步將其調(diào)整接種至 BioVector? 雜交瘤專用無血清培養(yǎng)基 中，起始接種密度設定為 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. 置于大容量轉(zhuǎn)瓶或低速搖床中持續(xù)培養(yǎng)，讓其自然生長并分泌抗體。(Maintain within large-volume roller bottles or low-shear shaker platforms to allow continuous execution of antibody secreting pathways.)

3. 密切監(jiān)測培養(yǎng)基顏色。當細胞密度達到頂峰并開始進入衰退期（通常在接種后 5 到 7 天，鏡下觀察活細胞率降至 40% 左右時，此時胞內(nèi)抗體釋放最徹底），即可終止培養(yǎng)。(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. 將全部培養(yǎng)液收集進大容量離心管中，在 1000乘以g 的離心力下高速離心 15 分鐘，徹底沉淀細胞碎片，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. 將澄清上清液通過 0.22 微米的無菌無紡濾膜進行過濾，隨后可直接輸入裝有 BioVector? Protein A 或 Protein G 的親和層析柱中進行單克隆抗體純化洗脫。將純化后的 9E10 抗體進行透析脫鹽，即可獲得高純度的工程級 Anti-Myc 單克隆抗體。(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 復蘇與凍存恢復流線 / Thawing and Post-Cryo Recovery Workflow

1. 提前在標準 T25 培養(yǎng)瓶中注入 6.0 mL 預熱的 BioVector? 增殖期完全生長培養(yǎng)基（含 15% 優(yōu)質(zhì)血清），置于孵箱中平衡氣體與溫控。(Pre-warm 6.0 mL of Phase A growth medium fortified with 15% premium serum in a T25 flask, equilibrating inside the incubator.)

2. 從液氮中取出 BioVector? MYC 1-9E10.2 凍存管，立刻全量投入 37 攝氏度恒溫水浴箱中快速水平搖晃，在 60 秒內(nèi)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凈臺，用移液槍吸出胞懸液，緩慢逐滴注入盛有 4.0 mL 預熱增殖培養(yǎng)基的 15 mL 離心管中。(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 下低速離心 5 分鐘，徹底吸干含有毒 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 新鮮的 15% 血清培養(yǎng)基輕彈懸起細胞沉淀，隨后全量接種到預熱平衡的 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. 復蘇后 24 小時必須在顯微鏡下進行活細胞計數(shù)并補充 2 到 3 mL 新鮮培養(yǎng)基，以稀釋復蘇死亡細胞釋放的毒性碎片。(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、長期保存與質(zhì)控規(guī)范 / Biosafety, Storage and Quality Controls

• 生物安全級別 (Biosafety Level)：BSL-1。屬于安全的常規(guī)小鼠雜交瘤細胞系，無已知人類病原體傳染風險，實驗廢棄物按標準生物醫(yī)療廢棄物高壓滅菌消殺即可。(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.)

• 超低溫長期保存 (Long-Term Banking)：凍存保質(zhì)配方為 90% BioVector? 增殖期完全生長培養(yǎng)基 + 10% 細胞級 DMSO（或采用 BioVector? Premium 無血清高效凍存液）。凍存管必須永久存放于液氮罐氣相或液相中（零下 150 攝氏度至零下 196 攝氏度）。嚴禁在零下 80 攝氏度普通機械冰箱內(nèi)長期擱置超過 1 個月，否則會發(fā)生抗體表達質(zhì)粒丟失或復蘇存活率斷崖式暴跌。(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.)

• 抗體分泌效價與特異性核心質(zhì)控紅線 / Antibody Secretion Quality Control Threshold：隨著傳代次數(shù)的增加（連續(xù)傳代超過 35 到 40 代以上），雜交瘤細胞極易發(fā)生非整倍體染色體丟失、基因重排或非分泌型克隆逃逸。嚴格質(zhì)控表現(xiàn)為：每隔 10 代，應收集一次細胞無血清培養(yǎng)上清，通過 ELISA 測試或常規(guī) Western Blot 測試驗證其對標準 Myc 標簽重組蛋白（如帶有 Myc-tag 的 GFP 蛋白）的結合能力。如果測得的上清液抗體效價出現(xiàn)數(shù)量級下降，或者出現(xiàn)非特異性雜帶，證實該雜交瘤細胞已經(jīng)發(fā)生功能退化。必須立即停止該代次生產(chǎn)，全盤銷毀細胞，并重新復蘇超低代次的種子庫進行恢復。(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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