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ClonaCell™-HY杂交瘤试剂盒

完整的杂交瘤生成试剂盒
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产品号 #（选择产品）

产品号 #03800_C

完整的杂交瘤生成试剂盒

产品优势

  • 节省时间：将杂交瘤的选择和克隆结合为一步
  • 节省资源：单细胞来源的杂交瘤在半固体培养基中形成可见的离散菌落，易于筛选和转移到液体培养基中进行扩增
  • 克隆效率：单个细胞悬浮并固定在半固体培养基中，防止因过度生长而丢失稀有的高产克隆

产品组分包括

  • ClonaCell™-HY Medium A, 500 mL（目录#03801）
  • ClonaCell™-HY Medium B, 500 mL（目录#03802）
  • ClonaCell™-HY Medium C, 100 mL（目录#03803）
  • ClonaCell™-HY Medium D, 90ml（目录#03804）
  • ClonaCell™-HY Medium E, 500 mL（目录#03805）
  • ClonaCell™-HY PEG, 1.5 mL（目录#03806）
总览
产品说明书及文档
应用领域
相关材料与文献
相关产品

总览

使用ClonaCell™-HY杂交瘤试剂盒中包含的培养基和试剂，完成杂交瘤开发及单克隆抗体生产的所有步骤：
• ClonaCell™-HY培养基A用于骨髓瘤和杂交瘤培养
• ClonaCell™-HY培养基B用于杂交瘤融合
• ClonaCell™-HY培养基C用于杂交瘤融合后恢复
• ClonaCell™-HY培养基D用于杂交瘤筛选与克隆
• ClonaCell™-HY培养基E用于杂交瘤生长
• ClonaCell™-HY PEG用于支持杂交瘤融合

分类
半固体培养基，专用培养基
 
细胞类型
杂交瘤细胞
 
种属
小鼠，其它细胞系，大鼠
 
应用
细胞培养，杂交瘤制备
 
品牌
ClonaCell
 
研究领域
抗体​制备，细胞系制备，药物发现和毒理检测，杂交瘤制备
 

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产品说明书及文档

请在《产品说明书》中查找相关支持信息和使用说明，或浏览下方更多实验方案。

Document Type
Product Name
Catalog #
Lot #
Language
Document Type
Product Information Sheet 1
Product Name
ClonaCell™-HY Hybridoma Kit
Catalog #
03800
Lot #
All
Language
English
Document Type
Product Information Sheet 2
Product Name
ClonaCell™-HY Hybridoma Kit
Catalog #
03800
Lot #
All
Language
English
Document Type
Product Information Sheet 3
Product Name
ClonaCell™-HY Hybridoma Kit
Catalog #
03800
Lot #
All
Language
English
Document Type
Product Information Sheet 4
Product Name
ClonaCell™-HY Hybridoma Kit
Catalog #
03800
Lot #
All
Language
English
Document Type
Technical Manual
Product Name
ClonaCell™-HY Hybridoma Kit
Catalog #
03800
Lot #
All
Language
English
Document Type
Safety Data Sheet
Product Name
ClonaCell™-HY Hybridoma Kit
Catalog #
03800
Lot #
All
Language
English

应用领域

本产品专为以下研究领域设计，适用于工作流程中的高亮阶段。探索这些工作流程，了解更多我们为各研究领域提供的其他配套产品。

Research Area
Workflow Stages

相关材料与文献

技术资料 (10)

Increase the Efficiency of Your Hybridoma Workflow with EasySep™ and ClonaCell™
Brochure
Increase the Efficiency of Your Hybridoma Workflow with EasySep™ and ClonaCell™
ClonaCell™ Media for Hybridoma and Cell Line Development
Brochure
ClonaCell™ Media for Hybridoma and Cell Line Development
ClonaCell™-HY Medium D for Robust Colony Formation During Semi-Solid Cloning
Brochure
ClonaCell™-HY Medium D for Robust Colony Formation During Semi-Solid Cloning
Protocol for Semi-Solid Hybridoma Cloning Using ClonaCell™-HY Medium in a 96-Well Plate
Technical Bulletin
Protocol for Semi-Solid Hybridoma Cloning Using ClonaCell™-HY Medium in a 96-Well Plate
Integrating Electrofusion with the ClonaCell™-HY Workflow for Hybridoma Production
Protocol
Integrating Electrofusion with the ClonaCell™-HY Workflow for Hybridoma Production
Why Use Semi-Solid Medium for Cloning Hybridomas and CHO Cells
3:39
Video
Why Use Semi-Solid Medium for Cloning Hybridomas and CHO Cells
ClonaCell™-HY Procedure
8:24
Video
ClonaCell™-HY Procedure
How to Prepare and Plate Semi-Solid Medium
1:52
Video
How to Prepare and Plate Semi-Solid Medium
How to Generate Hybridomas in 96-Well Plates Using ClonaCell™-HY Semi-Solid Cloning Medium
5:56
Video
How to Generate Hybridomas in 96-Well Plates Using ClonaCell™-HY Semi-Solid Cloning Medium
A Smarter Way To Clone: What You Don't Know About Limiting Dilution Can Hurt Your Probability of Monoclonality
34:50
Webinar
A Smarter Way To Clone: What You Don't Know About Limiting Dilution Can Hurt Your Probability of Mon...

常见问题

Why is there HT (hypoxanthine, thymidine) in Medium E?

Hybridomas are selected using HAT (hypoxanthine, aminopterin, thymidine). Aminopterin blocks the de novo pathway for synthesizing nucleotide precursors for DNA synthesis. The inhibition of the de novo pathway can persist even after the cells are removed from selection. Hypoxanthine and thymidine (HT) provide the necessary nucleotide precursors for hybridoma cells to synthesize DNA using the salvage pathway. Once the cells are growing well in Medium E, they can be gradually switched to Medium A or another medium without HT.

Is the serum in ClonaCell™-HY media heat inactivated?

Yes, all serum used in ClonaCell™-HY media is heat inactivated.

Is there any IgG in clonacell™-HY media?

While we don't add IgG to the ClonaCell™-HY media, we do add serum, which contains an undefined amount of IgG. We selectively use serum lots with low IgG levels in the production of ClonaCell™-HY media, however, levels vary from lot to lot. IgG levels in a specific lot of ClonaCell™-HY medium are available in the lot-specific Certificate of Analysis.

Are there antibiotics in ClonaCell™-HY media?

These products contain gentamycin rather than penicillin/streptomycin/amphotericin B, because gentamycin is more stable and is a broad spectrum antibiotic that is non-toxic to most mammalian cells in culture.

What is the optimal number of colonies per plate?

We recommend 50-150 colonies per plate. An average fusion will result in approximately 1000 colonies per fusion (approx. 100 colonies per plate). Even if the average number of colonies per plate approaches 300, there should still be enough separation between colonies to pick easily.

Why do I have to put my fused cells into liquid medium overnight? Why can't I just plate directly into Medium D?

We recommend waiting up to 24 hours so that all of the fused cells can go through one cell cycle. This will ensure they have a chance to express HPRT (hypoxanthine guanine phosphoribosyltransferase), the enzyme necessary to survive in the presence of aminopterin (present in Medium D). Additionally, fused cells are very fragile immediately after fusion. Waiting a day before mixing the cells with the methylcellulose will improve their survival. Although it is not recommended, fused cells may be plated on the same day as fusion, but the cells should be allowed to recover for several hours in ClonaCell™-HY Medium C prior to plating.

What myeloma and mouse strains should I use?

Myeloma: There are at least two common myeloma cell lines used to generate hybridomas - SP2/0 and P3X63Ag8.683. Both are available from ATCC. Researchers should ensure that the myeloma line is from a reliable source and is negative for mycoplasma. Mycoplasma contamination of the myeloma line can result in decreased efficiency of hybridoma formation. Mouse: We suggest using BALB/c splenocytes and parental myeloma cells of BALB/c for the following reasons: they are highly immune reactive, well characterized and myeloma cells are available from the same genetic strain. Other mouse strains, however, are also compatible with cloning in ClonaCell™-HY media.

Can I grow human/rat/T cell hybridomas in ClonaCell™-HY?

Although we have not tried to generate human, rat or T cell hybridomas during in-house testing, these experiments are expected to be successful using ClonaCell™-HY. The researcher would need to ensure that the cell lines used in the fusion are sensitive to HAT selection and grow well in methylcellulose-based medium.

There are very few colonies growing in my Medium D. Why?

Low numbers of colonies is generally a result of low fusion efficiency, which can have many causes. The fusion efficiency can be affected by the presence of serum during fusion, the presence of mycoplasma, low viability of cells, overexposure to polyethylene glycol or slow-growing myeloma cells prior to fusion.

Why does the ClonaCell™-HY manual suggest two different methods for fusion (A or B)? Can one expect better results with one method over the other?

Which method chosen is a personal preference and there should not be significant differences in efficiency. Method B is faster and has less steps, but Method B requires you to remove all the PEG before the cells are diluted, so you will risk aspirating cells if not very careful. With Method A, you dilute the PEG with Medium B, so you have less opportunity to lose cells.

Why does the ClonaCell™-HY manual suggest two different methods for fusion (A or B)? Can one expect better results with one method over the other?

A: Which method chosen is a personal preference and there should not be significant differences in efficiency. Method B is faster and has less steps, but Method B requires you to remove all the PEG before the cells are diluted, so you will risk aspirating cells if not very careful. With Method A, you dilute the PEG with Medium B, so you have less opportunity to lose cells.

Once I pick the colonies and grow the cells in plates, will the residual methylcellulose interfere with characterization? For example, will I have problems doing an ELISA?

 There will likely be some residual methylcellulose contamination when colonies are picked and transferred to the 96-well plate with the liquid growth medium. The concentration of methylcellulose, however, should be low enough that it should not interfere with most assays.

文献 (68)

Structural basis for nonneutralizing antibody competition at antigenic site II of the respiratory syncytial virus fusion protein. Mousa JJ et al. Proceedings of the National Academy of Sciences of the United States of America 2016 OCT

Abstract

Palivizumab was the first antiviral monoclonal antibody (mAb) approved for therapeutic use in humans, and remains a prophylactic treatment for infants at risk for severe disease because of respiratory syncytial virus (RSV). Palivizumab is an engineered humanized version of a murine mAb targeting antigenic site II of the RSV fusion (F) protein, a key target in vaccine development. There are limited reported naturally occurring human mAbs to site II; therefore, the structural basis for human antibody recognition of this major antigenic site is poorly understood. Here, we describe a nonneutralizing class of site II-specific mAbs that competed for binding with palivizumab to postfusion RSV F protein. We also describe two classes of site II-specific neutralizing mAbs, one of which escaped competition with nonneutralizing mAbs. An X-ray crystal structure of the neutralizing mAb 14N4 in complex with F protein showed that the binding angle at which human neutralizing mAbs interact with antigenic site II determines whether or not nonneutralizing antibodies compete with their binding. Fine-mapping studies determined that nonneutralizing mAbs that interfere with binding of neutralizing mAbs recognize site II with a pose that facilitates binding to an epitope containing F surface residues on a neighboring protomer. Neutralizing antibodies, like motavizumab and a new mAb designated 3J20 that escape interference by the inhibiting mAbs, avoid such contact by binding at an angle that is shifted away from the nonneutralizing site. Furthermore, binding to rationally and computationally designed site II helix-loop-helix epitope-scaffold vaccines distinguished neutralizing from nonneutralizing site II antibodies.
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An influenza A virus (H7N9) anti-neuraminidase monoclonal antibody with prophylactic and therapeutic activity in vivo Wilson JR et al. Antiviral Research 2016 NOV

Abstract

Zoonotic A(H7N9) avian influenza viruses emerged in China in 2013 and continue to be a threat to human public health, having infected over 800 individuals with a mortality rate approaching 40%. Treatment options for people infected with A(H7N9) include the use of neuraminidase (NA) inhibitors. However, like other influenza viruses, A(H7N9) can become resistant to these drugs. The use of monoclonal antibodies is a rapidly developing strategy for controlling influenza virus infection. Here we generated a murine monoclonal antibody (3c10-3) directed against the NA of A(H7N9) and show that prophylactic systemic administration of 3c10-3 fully protected mice from lethal challenge with wild-type A/Anhui/1/2013 (H7N9). Further, post-infection treatment with a single systemic dose of 3c10-3 at either 24, 48 or 72 h post A(H7N9) challenge resulted in both dose- and time-dependent protection of up to 100% of mice, demonstrating therapeutic potential for 3c10-3. Epitope mapping revealed that 3c10-3 binds near the enzyme active site of NA, and functional characterization showed that 3c10-3 inhibits the enzyme activity of NA and restricts the cell-to-cell spread of the virus in cultured cells. Affinity analysis also revealed that 3c10-3 binds equally well to recombinant NA of wild-type A/Anhui/1/2013 and to a variant NA carrying a R289K mutation known to infer NAI resistance. These results suggest that 3c10-3 has the potential to be used as a therapeutic to treat A(H7N9) infections either as an alternative to, or in combination with, current NA antiviral inhibitors.
View publication
Neutralizing human antibodies prevent Zika virus replication and fetal disease in mice. Sapparapu G et al. Nature 2016 NOV

Abstract

Zika virus (ZIKV) is an emerging mosquito-transmitted flavivirus that can cause severe disease, including congenital birth defects during pregnancy(1). To develop candidate therapeutic agents against ZIKV, we isolated a panel of human monoclonal antibodies (mAbs) from subjects with prior ZIKV infection. A subset of mAbs recognized diverse epitopes on the envelope (E) protein and exhibited potently neutralizing activity. One of the most inhibitory mAbs, ZIKV-117, broadly neutralized infection of ZIKV strains corresponding to African, Asian, and American lineages. Epitope mapping studies revealed that ZIKV-117 recognized a unique quaternary epitope on the E protein dimer-dimer interface. We evaluated the therapeutic efficacy of ZIKV-117 in pregnant and non-pregnant mice. mAb treatment markedly reduced tissue pathology, placental and fetal infection, and mortality in mice. Thus, neutralizing human mAbs can protect against maternal-fetal transmission, infection and disease, and reveal important determinants for structure-based rational vaccine design efforts.
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种属 Mouse, Other, Rat
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