自然殺傷（NK）細胞是可識別和殺死腫瘤和病毒感染的關鍵免疫細胞。 NK細胞是當今免疫治療常用的細胞。但是NK細胞培養操作難度大，對培養條件、培養技術要求很高，難以獲得高純度高效能的活性細胞。過度免疫治療所必需的離體NK細胞很難獲得。已經報道了幾種NK激活/擴增方法，其使用擴增介質加各種細胞因子（例如IL-2，IL-7和IL-15）。然而，這些需要研究人員的進一步優化。

Corning NK細胞活化擴增培養基套裝NK細胞體外擴增試劑盒包含預先涂覆的T-75燒瓶，50mL NK原代培養基，1.8mL NK Primary補充劑和1L NK擴增培養基。 NK細胞體外擴增試劑盒前三個組分是專為NK細胞激活設計的，而NK擴增培養基用于NK細胞的擴增。 NK初級培養基和NK膨脹培養基都使用高品質試劑和cGMP級原料制造。 NK細胞體外擴增試劑盒存在于培養基中的*蛋白質是血清白蛋白和重組人胰島素的可注射水平。

Corning NK細胞活化/擴增培養基套裝由多個組分組成，為NK細胞培養提供簡便的操作方案。使用Corning NK細胞活化擴增培養基套裝所獲得細胞數量產量高，活性高，具有很高的殺傷效應。

關于使用該試劑盒激活和擴增的細胞的增殖率，免疫表型和CCK-8細胞毒性測試的信息，請參閱Corning應用筆記使用康寧NK擴增試劑盒（CLS-CG-AN-427）進行人類天然殺傷細胞擴增。

提供產品：華雅干細胞致力于為中國干細胞領域的干細胞科學研究、臍帶血/造血干細胞/臍帶/脂肪/胎盤干細胞庫、干細胞臨床轉化醫學、干細胞美容及干細胞抗衰老、免疫細胞治療、T細胞治療、臨床干細胞治療等再生醫學和個體化治療產業鏈的相關客戶提供：MSC間充質干細胞培養基、iPS/ES胚胎干細胞培養基、血清替代物、羊水培養基、基底膜基質膠細胞支架、干細胞生長分化調控因子、BioMarker 流式檢測抗體、支原體檢測試劑盒、內毒素檢測試劑盒、干細胞存儲細胞凍存袋、細胞培養瓶、USP級DMSO二甲基亞砜細胞凍存液、細胞培養相關試劑耗材儀器等種類豐富的產品，以“Focus on improving the quality of Stem Cells專注干細胞研究，提升干細胞品質”為己任，我們與客戶攜手為中國干細胞產業的發展而前行。

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Corning NK細胞活化擴增培養基套裝供貨信息

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細胞活化擴增培養基套裝 NK細胞無血清培養基

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

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Corning NK細胞活化擴增培養基套裝 NK細胞體外擴增試劑盒Introduction

Natural killer (NK) cells are crucial immune cells that can recognize and kill tumors and virus infection. It is difficult to obtain the large numbers of NK cells ex vivo that are necessary for adoptive immu-notherapy. Several methods of NK activation/expansion have been reported that use expansion medium plus various cytokines (e.g., IL-2, IL-7, and IL-15). However, these require further optimization by researchers.

The Corning NK Expansion kit provides a complete set of materials and a standard operating procedure to expand NK cells with high expansion capacity, high viability, and high cytotoxicity. Researchers have no need to further optimize it, and the procedure has mini-mal consumables cost. In this application note, we provide com-petitive data, including cell yields, cell viability, and positive NK ratio and cytotoxicity capability in vitro, that compares the Corning NK expansion kit with other commercially available expansion media.

健康人的PBMC細胞接種數量為3x107 ，根據廠家建議進行細胞培養。使用Corning培養基時，0-5天使用NK活化培養基 (Primary Medium)，6-14天使用NK 擴增培養基 (NK Expansion Medium )添加1000 IU/mL IL-2 進行培養。細胞培養至14天收獲，進行細胞計數和腫瘤殺傷能力檢測。腫瘤殺傷實驗使用的靶細胞為K562細胞。

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NK細胞激活和擴增

在室溫（RT）下將抗凝血液以400xg離心10分鐘，然后將等離子體（頂層）轉移到新的管中。

熱自動等離子體在56℃滅活30分鐘，然后以800xg離心20分鐘以除去沉淀物。上清血漿應儲存在4℃，這將在隨后的14個培養日內消耗。

加入等體積的PBS（不含鈣和鎂的磷酸鹽緩沖鹽水，Corning Cat。編號21-040-CV），作為去除的自身血漿的替代物，以保持恒定體積并輕輕地重懸血細胞。

注意：在PBS中預先加入0.1％人血清白蛋白（HSA）有助于維持血細胞活力。

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根據制造商的指示，使用淋巴細胞分離培養基（LSM，Corning目錄號25-072-Cl）從上述血液樣品中制備外周血單核細胞（PBMC）。

注意：使用新鮮收集的人體血液（收集后2小時內），以獲得更好的表現;在使用前不要使用超過24小時吸取的血液。

用不含鈣和鎂的至少5倍PBS洗滌PBMC。離心機為500 xg

在室溫下10分鐘收集PBMC。

使用含有1.8mL NK Primary補充劑和10％自動等離子體的NK Primary培養基將PBMC稀釋至1×10 6個細胞/ mL。

用于激活和擴展人類天然殺傷細胞的康寧®NK試劑盒

協議

保修/免責聲明：除非另有規定，所有產品僅供研究使用。不適用于診斷或治療程序。不適用于人類。康寧生命科學不對這些產品在臨床或診斷應用中的性能提出任何索賠。

健康人的PBMC細胞根據廠家建議進行細胞培養。細胞培養至14天收獲，進行細胞計數和檢測。細胞使用CD3 FITC /CD56 PE進行染色，之后使用BDAccuriTM C6 flow cytometer進行檢測。

重慶市華雅干細胞技術有限公司主營：干細胞、細胞凍存、細胞治療、試劑耗材相關板塊的產品，以“傳遞科學價值，服務科學研究”為宗旨，經過近五年的努力推廣，重慶市華雅干細胞技術有限公司在全中國代理的BD科研產品、OriGen干細胞凍存袋等產品已經成功銷往：上海市、北京市、重慶市、天津市、廣東省的廣州市、深圳市、珠海市、江西省的南昌市、四川省的成都市、綿陽、浙江省的杭州市、寧波、溫州、江蘇省的南京市、蘇州、無錫、泰州、湖南省的長沙、陜西省的西安市、楊凌、甘肅省的蘭州、寧夏的銀川、新疆的烏魯木齊、廣西的南寧市、山東省的濟南、青島、東北三省遼寧省的沈陽市、大連市、吉林省的長春市、黑龍江省的哈爾濱市等中國絕大部分省市和自治區。

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NK細胞活化擴增培養基英文使用方法介紹

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.

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干細胞無血清培養基

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[供應]Corning NK-Corning NK細胞活化擴增培養基套裝 NK細胞體外擴增試劑盒