Corning NK细胞活化扩增培养基套装

Corning NK细胞活化扩增培养基套装 NK细胞体外扩增试剂盒

Corning NK细胞活化扩增培养基套装产品组成
1)  NK细胞活化培养基
2)  NK细胞活化添加剂
3)  NK细胞扩增培养基
4)  抗体预包被T75细胞培养瓶

1. 细胞培养袋

 
中文名称：康宁细胞活化扩增培养基套装 NK细胞体外扩增试剂盒
英文名称：Corning® NK Kit for Activation and Expansion of Human Natural Killer Cells
 
Corning NK细胞活化扩增培养基套装产品特色
v  扩增能力强，在14天内可达到100倍扩增
v  效应细胞比例高，可达80％

1. 操作方便

 
Corning NK细胞活化扩增培养基套装产品介绍
自然杀伤（NK）细胞是可识别和杀死肿瘤和病毒感染的关键免疫细胞。 NK细胞是当今免疫ZL常用的细胞。但是NK细胞培养操作难度大，对培养条件、培养技术要求很高，难以获得高纯度GX能的活性细胞。过度免疫ZL所必需的离体NK细胞很难获得。 已经报道了几种NK激活/扩增方法，其使用扩增介质加各种细胞因子（例如IL-2，IL-7和IL-15）。 然而，这些需要研究人员的进一步优化。
Corning NK细胞活化扩增培养基套装NK细胞体外扩增试剂盒包含预先涂覆的T-75烧瓶，50mL NK原代培养基，1.8mL NK Primary补充剂和1L NK扩增培养基。 NK细胞体外扩增试剂盒前三个组分是专为NK细胞激活设计的，而NK扩增培养基用于NK细胞的扩增。 NK初级培养基和NK膨胀培养基都使用高品质试剂和cGMP级原料制造。 NK细胞体外扩增试剂盒存在于培养基中的WY蛋白质是血清白蛋白和重组人胰岛素的可注射水平。
Corning NK细胞活化/扩增培养基套装由多个组分组成，为NK细胞培养提供简便的操作方案。使用Corning NK细胞活化扩增培养基套装所获得细胞数量产量高，活性高，具有很高的杀伤效应。
 关于使用该试剂盒激活和扩增的细胞的增殖率，免疫表型和CCK-8细胞毒性测试的信息，请参阅Corning应用笔记使用康宁NK扩增试剂盒（CLS-CG-AN-427）进行人类天然杀伤细胞扩增。
 
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Corning NK细胞活化扩增培养基套装供货信息

Product	Cat. No.	Qty	Storage	Shelf-life Duration
Corning NK Expansion kit	88-570-kit	1 kit	2°C to 8°C	9 months
Kit includes:
Pre-coated T-75 flask	Included	1 flask
NK Primary medium	Included	1 x 50 mL
NK Primary supplement	Included	1 x 1.8 mL
NK Expansion medium	Included	1 x 1,000 mL
Gas-permeable culture bag	Included	1 x 640 cm2

 
Corning NK细胞活化扩增培养基套装 NK细胞无血清培养基
Related Products

Corning Cat. No.	Product	Qty/Pk	Qty/Cs
25-072-CL	Lymphocyte separation medium	1	1
21-040-CV	Phosphate Buffer Saline (PBS) [-] Calcium and magnesium, 1x (500 mL)	6	6
354043	Interleukin-2 (IL-2), human recombinant, 10,000 BRMP units	1	1
431082	225 cm² cell culture flask, angled neck, vented cap	5	25
88-610-20	Gas-permeable cell culture bag	1	1

健康人的PBMC细胞接种数量为3x107 ，根据厂家建议进行细胞培养。使用Corning培养基时，0-5天使用NK活化培养基 (Primary Medium)，6-14天使用NK 扩增培养基 (NK Expansion Medium )添加1000 IU/mL IL-2 进行培养。细胞培养至14天收获，进行细胞计数和肿瘤杀伤能力检测。肿瘤杀伤实验使用的靶细胞为K562细胞。

NK细胞激活和扩增
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在室温（RT）下将抗凝血液以400xg离心10分钟，然后将等离子体（顶层）转移到新的管中。
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热自动等离子体在56℃灭活30分钟，然后以800xg离心20分钟以除去沉淀物。上清血浆应储存在4℃，这将在随后的14个培养日内消耗。
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加入等体积的PBS（不含钙和镁的磷酸盐缓冲盐水，Corning Cat。编号21-040-CV），作为去除的自身血浆的替代物，以保持恒定体积并轻轻地重悬血细胞。
注意：在PBS中预先加入0.1％人血清白蛋白（HSA）有助于维持血细胞活力。

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根据制造商的指示，使用淋巴细胞分离培养基（LSM，Corning目录号25-072-Cl）从上述血液样品中制备外周血单核细胞（PBMC）。
注意：使用新鲜收集的人体血液（收集后2小时内），以获得更好的表现;在使用前不要使用超过24小时吸取的血液。
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用不含钙和镁的至少5倍PBS洗涤PBMC。离心机为500 xg
在室温下10分钟收集PBMC。
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使用含有1.8mL NK Primary补充剂和10％自动等离子体的NK Primary培养基将PBMC稀释至1×10 6个细胞/ mL。
用于激活和扩展人类天然杀伤细胞的康宁®NK试剂盒
协议
保修/免责声明：除非另有规定，所有产品仅供研究使用。不适用于诊断或ZL程序。不适用于人类。康宁生命科学不对这些产品在临床或诊断应用中的性能提出任何索赔。
 

健康人的PBMC细胞根据厂家建议进行细胞培养。细胞培养至14天收获，进行细胞计数和检测。细胞使用CD3 FITC /CD56 PE进行染色，之后使用BDAccuriTM C6 flow cytometer进行检测。

NK细胞活化扩增培养基英文使用方法介绍

 Materials and Methods
Peripheral blood mononuclear cells (PBMCs) were isolated from healthy donor whole blood using lymphocyte separation medium (Corning Cat. No. 25-072-CI) according to the manufacturer’s instructions. The wash buffer was PBS (Corning Cat. No. 21-040-CV), and the culture vessel was a gas-permeable cell culture media bag (Corning Cat. No. 88-610-20). Interleukin 2 (IL-2, Corning Cat. No. 354043) was used as an in vitro cytokine for NK cell activation and expansion.
Corning NK Cell Activation and Expansion
      On Day 0, the auto-plasma was inactivated by heating at 56°C for 30 min. and centrifuged at 800 xg for 20 min. to remove the precipitates. The supernatant was collected and stored at 4°C for further use.
      The PBMCs were washed once with at least 5-fold PBS buffer, and the sample was centrifuged at 500 xg for 10 min. at room temperature. The density of PBMCs was adjusted to 1 x 106 cells/mL using KBM Corning NK Primary medium containing 1.8 mL KBM Corning NK Primary supplement and 10% auto-plasma.
      The pre-coated T-75 flask was washed carefully twice with PBS just before use. Thirty milliliters of the suspension of PBMCs were seeded into the pre-coated T-75 flask and incubated at 37°C in a humidified atmosphere of 5% CO2 in air.
Note: Before Day 6 when the medium turned yellow and the cell density was above 2 x 106 cells/mL, fresh KBM NK Primary medium plus 10% auto-plasma were added to the cell suspension to ensure that cell density stayed within the range of 2.5 x 105 to 2.0 x 106 cells/mL.
      On Day 6, all cells were centrifuged and re-suspended with an appropriate volume of KBM NK Expansion medium containing 1,000 IU/mL IL-2 and 10% auto-plasma and then transferred to a new T-225 flask or gas-permeable culture bag.
      In the following days, KBM Corning NK Expansion medium containing 1,000 IU/mL IL-2 was added every 2 to 3 days based on the cell proliferation status to ensure that cell density stayed within 2.5 x 105 to 2.0 x 106 cells/mL.
      The cells were harvested for CCK8 cytotoxic and immunophenotyping testing when the total cell number exceeded 2 x 109.
 
Note: The cells were gently dissociated, and the culture vessels were tapped softly to avoid cell damage and maintain cell viability throughout the entire experimental process.