Features
- ・Natural killer (NK) cells can be expanded from human peripheral blood mononuclear cells(PBMCs) without using feeder cells.
- ・NK cells can be expanded several hundred to several thousand-fold in 2 to 3 weeks of culture.
- ・One kit is sufficient to expand NK cells patients using 20-50 mL each of the whole blood.
- ・Research use only.
Contents
- NK Cell Initial Medium, Cocktail
- NK Cell Initial Flask
- NK Cell Subculture Medium
BINKIT specification sheet (PDF file)
Options
- Culture bag(small, large)
- Culture bag connection tube
MSDS
Application Data
Procedure overview
Procedures
- ・Preparing reagentsNK Cell Initial Medium and NK Cell Subculture Medium should be supplemented with 5 % (v/v) of heat-inactivated FBS or autologous plasma.
- ・Preparing peripheral blood mononuclear cells (PBMCs)
- PBMCs should be isolated from the whole blood by density gradient centrifugation using Ficoll-Paque.
- ・Washing an NK cell Initial Flask
- 10 ml PBS should be added to an NK Cell Initial Flask. Slant the flask to cover the entire surface with PBS. Aspirate the liquid completely from the flask. Care should be taken not to scratch the inner surface of the flask. Repeat the washing process two more times.
- ・Culturing NK cells from PBMCs
- PBMCs should be suspended in the NK Cell Initial Medium at 1 x 106 cells/ml. Add 40 μl of NK Cell Initial Cocktail to the 1 ml of the cell suspension. Seed the cells onto the pre-washed NK Cell Initial Flask, and incubate under 5 % CO2 at 37 ℃ for 3 days.
- ・Changing medium and sub-culturing
- Transfer floating as well as adherent cells to conical centrifuge tube and centrifuge at 200 x g for 8 minutes. Remove the supernatant and re-suspended the cells at 1 x 106 cells/ml in the NK Cell Subculture Medium supplemented with 5 % (v/v) of heat- inactivated FBS or autologous plasma. The cell suspension is transferred to conventional culture flasks and incubated under 5 % CO2 at 37 ℃. Cells should be sub-cultured every 2 - 3 days by suspending cells in the completed NK Cell Subculture Medium at 0.8 x 106 cells/ml.
- ・Suggested culturing period: 2 to 3 weeks.
Other supplies required
- Ficoll-Paque (GE Healthcare, Sweden)
- Sterile PBS
- FBS or autologous plasma (It is desirable to have them heat-inactivated at 56 ℃ for 30 minutes.)
- Sterile conical centrifuge tubes
- Sterile culture flasks
Precautions
- NK Cell Initial Flask may carry condensation on the surface, which does not adversely affect the performance of the kit.
This product contains human plasma albumin.
References
- Ming-Ju Hsieh, Jen-Tsun Lin, Yi-Ching Chuang, Yu-Sheng Lo, Chia-Chieh Lin, Hsin-Yu Ho, Mu-Kuan Chen: Raddeanin A augments the cytotoxicity of natural killer cells against chronic myeloid leukaemia cells by modulating MAPK and Ras/Raf signalling pathways., J. Cell. Mol. Med., 28(16): e70016, 2024
https://doi.org/10.1111/jcmm.70016 - Ming-Ju Hsieh, Jen-Tsun Lin, Yi-Ching Chuang, Chia-Chieh Lin, Yu-Sheng Lo, Hsin-Yu Ho, Mu-Kuan Chen: Limocitrin increases cytotoxicity of KHYG-1 cells against K562 cells by modulating MAPK pathway., Environ. Toxicol., 38(12): 2939-2951, 2023
https://doi.org/10.1002/tox.23929 - Nickolas Chelyapov, Toai T Nguyen, and Rafael Gonzalezb., Autologous NK cells propagated and activated ex vivo decrease senescence markers in human PBMCs. Biochem Biophys Rep. 32:101380, 2022
PMID: 36386442 - Jen-Tsun Lin, Yi-Ching Chuang, Mu-Kuan Chen, Yu-Sheng Lo, Chia-Chieh Lin, Hsin-Yu Ho, Yen-Tze Liu, Ming-Ju Hsieh., Shuterin Enhances the Cytotoxicity of the Natural Killer Leukemia Cell Line KHYG-1 by Increasing the Expression Levels of Granzyme B and IFN-γ through the MAPK and Ras/Raf Signaling Pathways. International Journal of Molecular Sciences. 23(21):12816, 2022
PMID: 36361609 - Chu Thi Thao, Bui Viet Anh, Nguyen Van Phong, Nguyen Trung Kien, Nguyen Dac Tu, Doan Trung Hiep, Nguyen Thanh Liem, Hoang Thi My Nhung., Immune Cells Expansion from Peripheral Blood of Some Vietnamese Cancer Patients. VNU Journal of Science: Natural Sciences and Technology. 37(3):2588-1140, 2021
https://doi.org/10.25073/2588-1140/vnunst.5291 - Hui Menga, Yi Ding, Enjie Liu, Wencai Li, Liang Wang., ZG16 regulates PD-L1 expression and promotes local immunity in colon cancer. Translational Oncology. 14(2):101003, 2021
PMID: 33360840 - Ikumi Katano,Ryoji Ito,Kenji Kawai,and Takeshi Takahashi., Improved Detection of in vivo Human NK Cell-Mediated Antibody-Dependent Cellular Cytotoxicity Using a Novel NOG-FcγR-Deficient Human IL-15 Transgenic Mouse. Frontiers in Immunology. 11:532684, 2020
PMID: 33117338 - Nguyen Thanh Liem, Nguyen Van Phong, Nguyen Trung Kien, Bui Viet Anh, Truong Linh Huyen, Chu Thi Thao, Nguyen Dac Tu, Doan Trung Hiep, Do Thi Hoai Thu, and Hoang Thi My Nhung., Phase I Clinical Trial Using Autologous Ex Vivo Expanded NK Cells and Cytotoxic T Lymphocytes for Cancer Treatment in Vietnam. Int J Mol Sci. 20(13):3166, 2019
PMID: 31261712 - Chu Thi Thao, Do Thi Hoai Thu, Bui Viet Anh, Truong Linh Huyen, Nguyen Van Phong, Nguyen Thanh Liem, and Hoang Thi My Nhung., Efficient Expansion of Human Umbilical Cord Blood- Derived NK Cells Ex Vivo without Requiring Feeder Layers. Biomed J Sci & Tech Res. 8:6739-6744, 2018
DOI: 10.26717/BJSTR.2018.08.001711 - Hoang Thi My Nhung, Bui Viet Anh, Truong Linh Huyen, Doan Trung Hiep, Chu Thi Thao, Phung Nam Lam, Nguyen Thanh Liem., Ex vivo expansion of human peripheral blood natural killer cells and cytotoxic T lymphocytes from lung cancer patients. Oncol Lett. 15:5730-5738, 2018
PMID: 29552207 - 照沼裕、芦葉恵介、鄧学文、茂野淳平、高根翼、鮫島葉月、照沼篤, 活性化NK細胞の大量培養法の開発とその有効利用の可能性. BIO Clinica. 31:55-61, 2016 [Japanese].
- 鄧学文、芦葉恵介、照沼篤、高根翼、鮫島葉月、贄田美江、照沼裕, 培養NK細胞の投与による体内NK細胞の活性化. Medical Science Digest. 41, 42-45, 2015.
- Deng X, Terunuma H, Terunuma A, Takane T, Nieda M., Ex vivo-expanded natural killer cells kill cancer cells more effectively than ex vivo-expanded gd T cells or ab T cells. Int Immunopharmacol. 22:486-491, 2014
PMID: 25131561 - Terunuma H, Deng X, Nishino N, Watanabe K: NK cell-based autologous immune enhancement therapy (AIET) for cancer. Stem Cells Regenerative Med. 9:9-13, 2013
PMID: 24693203 - Deng X, Terunuma H, Nieda M, Xiao W, Nicol A., Synergistic cytotoxicity of ex vivo expanded natural killer cells in combination with monoclonal antibody drugs against cancer cells. Int Immunopharmacol. 14:593-605, 2012
PMID: 23063974
Orders
- For ordering BINKIT, please fill an order form and send it to info@bij-net.com.