Nci h1650

Manufactured by American Type Culture Collection

Sourced in United States

The NCI-H1650 is a human lung adenocarcinoma cell line. It is a widely used model for the study of lung cancer.

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H1650 (273 citations)

94 protocols using nci h1650

Metabolic Reprogramming in Lung Cancer

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The normal human bronchial epithelioid cell (HBE) and the non-small-cell lung cancer cell lines including NCI-H1650, NCI-H1299, A549, NCI-H460, HCC4006, NCI-H1975, and NCI-H358 were obtained from ATCC and cultured with recommended culture medium. The primary antibodies for western blotting including anti-TRAF6 (#8028), hexokinase-1 (#2024), hexokinase-2 (#2867), GLUT1 (#12939), PKM2 (#4053), LDHA (#35 82), VDAC-1 (#4661), phosphor-Akt (#4060), Akt (#8596), phosphor-S6 (#4858), and ubiquitin (#58395) as well as the secondary anti-rabbit IgG HRP (#7074) were products of Cell Signaling Technology Inc. (Danvers, MA). β-Actin (A5316) was obtained from Sigma-Aldrich. In immunohistochemistry staining, the primary antibodies against hexokinase-2 (ab227198) and Ki67 (ab15580) were products of Abcam. TRAF6 shRNA#1 (TRCN0000007350) and shRNA#2 (TRCN0000007351) were purchased from the Sigma Mission shRNA library. The constitutively active Akt (CA-Akt) plasmid (Cat. #10841) and pLKO.1 GFP shRNA (Cat. #30323) were purchased from Addgene (Cambridge, MA, USA). Recombinant human insulin-like growth factor 1 (IGF-1) was a product of R&D (Cat. 291-G1-200). Lipofectamine 2000 was obtained from Invitrogen (Carlsbad, CA).

Feng L., Feng S., Nie Z., Deng Y., Xuan Y., Chen X., Lu Y., Liang L, & Chen Y. (2021). TRAF6 Promoted Tumor Glycolysis in Non-Small-Cell Lung Cancer by Activating the Akt-HIFα Pathway. BioMed Research International, 2021, 3431245.

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Lung Adenocarcinoma Cell Lines: Cultivation and Characterization

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Lung adenocarcinoma cell lines, including NCI-H1650, NCI-H1299, LTEP-a 2, NCI-H1975, CaLu-3, A549, PG49, NCI-H358, NCI-H1299 and HEY-293T, were obtained from ATCC. The cell lines were maintained in Roswell Park Memorial Institute (RPMI) -1640 containing 10% fetal bovine serum (FBS; Invitrogen, USA).
The lung carcinoma tissues and clinical data obtained from our institute were approved by the Institutional Review Board of China (approval ID 81470137). We also subjected the cell lysates from the clinical specimens to Western blot analysis.

Zhang L., Wang Y., Zhang B., Zhang H., Zhou M., Wei M., Dong Q., Xu Y., Wang Z., Gao L., Qu Y., Shi B., Zhu J., Yin Y., Chen Y., Sun L., Zhang W., Xu S., Ying G, & Wang C. (2017). Claudin-3 expression increases the malignant potential of lung adenocarcinoma cells: role of epidermal growth factor receptor activation. Oncotarget, 8(14), 23033-23047.

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Generation of Null Alleles Using CRISPR

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The Hap1 cell line (8 (link)) was kindly provided by Dr. Thijn Brummelkamp, Netherlands Cancer Institute. The 293FT cell line used to generate high-titer lentiviruses was obtained from ThermoFisher Scientific; the A549, PC3, MDA-MB-361, HCC-1954 and NCI-H1650 cell lines were purchased from ATCC, where they were validated by STR profiling, and used at passage numbers <5. The KOPN8 cell line was a generous gift from Professor Michael Cleary, Stanford University. The Phoenix-Ampho cell line used for retrovirus production was purchased from Allele Biotechnology (San Diego, CA).
Null alleles for genes were constructed using the CRISPR/Cas9 system. For Hap1 cells the oligos encoding the guide RNAs were cloned into pSpCas9(BB)-2A-GFP (PX458, Addgene Plasmid #48138 from Dr. Feng Zhang) (9 (link)). Single cells were sorted using flow cytometry, expanded and clones bearing null alleles were identified by Sanger sequencing and immunoblotting. For gene disruption in cancer cell lines, the gRNAs validated in the Hap1 cells were introduced into LentiCRISPR v2 (Addgene Plasmid #52961 from Dr. Feng Zhang) (10 (link)) for lentiviral-mediated delivery. The oligo sequences for guide RNAs are provided in the supplementary methods.

Dubey R., Lebensohn A., Bahrami-Nejad Z., Marceau C., Champion M., Gevaert O., Sikic B.I., Carette J.E, & Rohatgi R. (2016). Chromatin-remodeling complex SWI/SNF controls multidrug resistance by transcriptionally regulating the drug efflux pump ABCB1. Cancer research, 76(19), 5810-5821.

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Sal-B Inhibits NSCLC Metastasis

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To investigate the therapeutic effect of Sal-B on NSCLC metastasis, two NSCLC cell lines (NCI-H2030 and NCI-H1650; ATCC, USA) were selected. All cells were grown in DMEM (Hyclone, USA) plus 10% fetal bovine serum (FBS; Hyclone) at 37°C in a humidified environment of 5% CO₂. Sal B (purity ≥98%; Hongqiao Pharmaceutical Technology Research Institute Co., Ltd., Nanjing, China) was dissolved in absolute ethanol to 50 mM and stored at −80°C. Moreover, TEPP-46 was acquired from MedChemExpress Company (USA). NSCLC cell lines were treated with 10 μM TEPP-46 for selectively activating pyruvate kinase M2 (PKM2).

Zhang H., Tang J., Cao Y, & Wang T. (2022). Salvianolic Acid B Suppresses Non-Small-Cell Lung Cancer Metastasis through PKM2-Independent Metabolic Reprogramming. Evidence-based Complementary and Alternative Medicine : eCAM, 2022, 9302403.

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Characterization of NSCLC Cell Lines

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NSCLC cell lines were obtained from the H. Lee Moffitt Lung Cancer Center of Excellence Cell Line Core and maintained in RPMI medium supplemented with 10% heat-inactivated fetal bovine serum and 1% Streptomycin/Penicillin. The cell lines used were the following: NCI-H1666 (ATCC CRL-5885, human adenocarcinoma), NCI-H520 (ATCC HTB-182, human, squamous cell carcinoma), NCI-H1650 (ATCC CRL-5883, human adenocarcinoma), NCI-1975 (ATCC CRL-5908, human adenocarcinoma), NCI-H596 (ATCC HTB-178, human adenosquamous carcinoma), and NCI-H60 (ATCC CRL-5821, human small cell lung carcinoma). Cell lines were cultured in 100 mm plates and kept in a humidified water-jacketed incubator at 37°C and 5% CO₂ and passaged every 5–7 days to maintain a confluence of approximately 80%. Cell lines tested negative for mycoplasma contamination and were authenticated by short tandem repeat (STR) analysis of loci TH01, TPOX, vWA, CSF1PO, D16S539, D7S820, D13S317 and D5S818. Genotypes of both cell lines were based on the Cell Line Encyclopedia as previously described [20 (link)].

Pérez-Morales J., Mejías-Morales D., Rivera-Rivera S., González-Flores J., González-Loperena M., Cordero-Báez F.Y., Pedreira-García W.M., Chardón-Colón C., Cabán-Rivera J., Cress W.D., Gordian E.R., Muñoz-Antonia T., Cabrera-Ríos M., Isidro A., Coppola D., Rosa M., Boyle T.A., Izumi V., Koomen J.M, & Santiago-Cardona P.G. (2018). Hyper-phosphorylation of Rb S249 together with CDK5R2/p39 overexpression are associated with impaired cell adhesion and epithelial-to-mesenchymal transition: Implications as a potential lung cancer grading and staging biomarker. PLoS ONE, 13(11), e0207483.

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Culturing BEAS-2B, HCC827, and NCI-H1650 Cell Lines

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The human nontumorigenic lung epithelial cell line BEAS-2B was purchased from the American Type Culture Collection (ATCC) and was cultured in BEBM complete medium supplemented with 10% FBS and incubated at 37°C and 5% CO₂ atmosphere along with penicillin (100 U/ml) and streptomycin (100 mg/ml, HyClone, USA). The adenocarcinoma lung cancer cell lines HCC827 and NCI-H1650 were purchased from ATCC and were cultured in RPMI-1640 medium supplemented with 10% FBS and incubated at 37°C and 5% CO₂ atmosphere along with penicillin (100 U/ml) and streptomycin (100 mg/ml, HyClone, USA) at 37°C with 5% CO₂.

Xue Y., Zhang J., Hou J, & Wang X. (2022). EGFR-AS1 Promotes Nonsmall Cell Lung Cancer (NSCLC) Progression via Downregulating the miR-524-5p/DRAM1 Axis and Inhibiting Autophagic Lysosomal Degradation. Journal of Oncology, 2022, 4402536.

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NSCLC Cell Lines and Patient Tissues

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Five NSCLC cell lines (SK‐MES‐1, NCI‐H1650, A549, NCI‐H1975, 95D) and normal lung epithelial cells (NLEC) were purchased from ATCC. The cells were cultured in Dulbecco's modified eagle's medium (DMEM) containing 10% fetal bovine serum (FBS) at 37°C in 5% CO2. The tumor tissues and patient data were obtained from The Second Hospital of Jilin University. All of the patients were provided written informed consent.

Jia X., Wang Z., Qiu L., Yang Y., Wang Y., Chen Z., Liu Z, & Yu L. (2016). Upregulation of LncRNA‐HIT promotes migration and invasion of non‐small cell lung cancer cells by association with ZEB1. Cancer Medicine, 5(12), 3555-3563.

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Culturing Diverse Lung Cell Lines

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Human lung bronchial epithelial cell line (BEAS-2B) and human non-small cell lung cancer cell lines (A549, NCI-H1299, NCI-H1650) were all available from ATCC (Manassas, VA, USA) and maintained at 37 °C in an incubator supplied with 5% CO₂. Human non-small cell lung cancer cell line PC-9 was purchased from COBIOER Company (Nanjing, China). BEAS-2B cell line was cultivated in BEGM medium (Gibco) with LHC-9 media (Gibco, USA). F-12 K medium (Gibco) was utilized to cultivate A549 cells with 10% FBS. NCI-H1299, NCI-H1650 and PC-9 cells were routinely cultured with 10% FBS in RPMI-1640 medium (Gibco).

Weng L., Qiu K., Gao W., Shi C, & Shu F. (2020). LncRNA PCGEM1 accelerates non-small cell lung cancer progression via sponging miR-433-3p to upregulate WTAP. BMC Pulmonary Medicine, 20, 213.

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NSCLC Tumor Characterization Protocol

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40 patients with pathologically confirmed NSCLC were enrolled for the study from The Affiliated Hospital of Qingdao University from Jan, 2015 to Feb, 2017. The informed written consent had been signed by all patients and the hospital had approved the study. Tumors were histologically graded according to the American Joint Committee on Cancer (AJCC, version 8) [10 (link)]. The human lung epithelial cell BEAS-2B, and lung cancer cell lines NCI-H441, PC-9, NCI-H1650, A549 were purchased from ATCC. All cell lines were cultured in DMEM supplemented with 10% FBS at 37°C under 5% CO₂.

Yu W., Li D., Ding X., Sun Y., Liu Y., Cong J., Yang J., Sun J., Ning X., Wang H, & Xu T. (2019). LINC00702 suppresses proliferation and invasion in non-small cell lung cancer through regulating miR-510/PTEN axis. Aging (Albany NY), 11(5), 1471-1485.

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Sourcing and Culturing Cancer Cell Lines

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Human prostate carcinoma PPC-1, PC-3, DU145, pancreatic carcinoma SU.86.86, MIA-PaCa-2, PANC-1, BxPC-3, Panc 10.05, Capan-1, lung carcinoma A549, NCI-H1650, mammary epithelial 184A1, MCF10A, and melanoma MDA-MB-435 cells were obtained from ATCC. Breast carcinoma cell lines were obtained from either ATCC or from the laboratories of Drs. Steve Ethier and Adi Gazdar [14 (link)]. Bone marrow aspirates or peripheral blood samples were collected from acute myeloid leukemia (AML) patients under the OHSU Institutional Review Board (IRB) approved 4422 research collection protocol which covers in vitro drug testing of leukemia cells and genetic studies. Patients signed an IRB-approved written consent form after verbal consent was obtained. Mononuclear cells were isolated from the aspirate using a Ficoll gradient and viability assayed using GuavaNexin reagent. All patients were treated in accordance with the ethical guidelines laid out in the Declaration of Helsinki. AML cells were cultured in RPMI medium supplemented with 10% fetal bovine serum. Human primary hepatocytes were from Lonza. All cells were cultured according to the provider's guidelines.

Rozanov D., Cheltsov A., Sergienko E., Vasile S., Golubkov V., Aleshin A.E., Levin T., Traer E., Hann B., Freimuth J., Alexeev N., Alekseyev M.A., Budko S.P., Bächinger H.P, & Spellman P. (2015). TRAIL-Based High Throughput Screening Reveals a Link between TRAIL-Mediated Apoptosis and Glutathione Reductase, a Key Component of Oxidative Stress Response. PLoS ONE, 10(6), e0129566.

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