Rapamycin

Manufactured by Cell Signaling Technology

Sourced in United States, United Kingdom

Rapamycin is a potent immunosuppressant and anti-cancer drug. It functions as an inhibitor of the mammalian target of rapamycin (mTOR) signaling pathway, which is a key regulator of cell growth, proliferation, and survival.

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Lab products found in correlation

Ly294002, by Cell Signaling Technology (17 mentions) Wortmannin, by Cell Signaling Technology (7 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (7 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (6 mentions) U0126, by Cell Signaling Technology (5 mentions) Dimethyl sulfoxide (dmso), by Merck Group (5 mentions) 3 methyladenine, by Merck Group (4 mentions) Cycloheximide (chx), by Merck Group (4 mentions) Chloroquine, by Merck Group (3 mentions) Anti p70s6k, by Cell Signaling Technology (3 mentions) Sp600125, by Cell Signaling Technology (3 mentions) 3 methyladenine 3 ma, by Merck Group (3 mentions) Lipofectamine 3000, by Thermo Fisher Scientific (3 mentions) Sb203580, by Cell Signaling Technology (3 mentions) Actinomycin d, by Merck Group (3 mentions) Torin 1, by Cell Signaling Technology (3 mentions) U73122, by Bio-Techne (3 mentions) Torin1, by Bio-Techne (3 mentions) Mg132, by Merck Group (3 mentions) B27 supplement, by Thermo Fisher Scientific (3 mentions)

127 protocols using rapamycin

Irradiation and Cytokine Secretion Assay

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For TPCA-1 treatment, cells were detached using EDTA, irradiated using a ¹³⁷Cs source, seeded in 25-cm² tissue culture flasks, and incubated at 37°C 5% CO₂. TPCA-1 (1 or 10 μM; S2824, Selleckchem) was added to cell cultures after irradiation.
For rapamycin treatment, cells were passaged, placed into growth medium containing 5 nM of rapamycin (9904S, Cell Signaling Technology), and incubated at 37°C 5% CO₂ for 1 day. Cells were detached and irradiated as indicated above, placed in 25-cm² tissue culture flasks in growth media containing 5 nM rapamycin, and incubated at 37°C 5% CO₂.
On day 2 after irradiation, cells were placed in serum-free, TPCA-1/rapamycin-free growth media containing 0.5% bovine serum albumin (BSA). Cell-free supernatants were harvested on day 3 after irradiation and used for migration assays or enzyme-linked immunosorbent assay (ELISA).

Walle T., Kraske J.A., Liao B., Lenoir B., Timke C., von Bohlen und Halbach E., Tran F., Griebel P., Albrecht D., Ahmed A., Suarez-Carmona M., Jiménez-Sánchez A., Beikert T., Tietz-Dahlfuß A., Menevse A.N., Schmidt G., Brom M., Pahl J.H., Antonopoulos W., Miller M., Perez R.L., Bestvater F., Giese N.A., Beckhove P., Rosenstiel P., Jäger D., Strobel O., Pe’er D., Halama N., Debus J., Cerwenka A, & Huber P.E. (2022). Radiotherapy orchestrates natural killer cell dependent antitumor immune responses through CXCL8. Science Advances, 8(12), eabh4050.

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Rapamycin and Oseltamivir Carboxylate Modulate H1N1 Infection

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Pre-treated MLE12 cells with rapamycin (100 nM) (Cell Signaling Technology, MA) for 1 h, were re-stimulated with H1N1 virus at multiplicity of infection (MOI) = 0.01 for 2 h and co-cultured with rapamycin (100 nM) with or without oseltamivir carboxylate (10 μg/ml) for 22 hours. After 24 h of culture, cells were harvested for analyzing the protein expressions.

Jia X., Liu B., Bao L., Lv Q., Li F., Li H., An Y., Zhang X., Cao B, & Wang C. (2018). Delayed oseltamivir plus sirolimus treatment attenuates H1N1 virus-induced severe lung injury correlated with repressed NLRP3 inflammasome activation and inflammatory cell infiltration. PLoS Pathogens, 14(11), e1007428.

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Establishing Venetoclax-Resistant AML Cell Lines

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Human AML cell lines, U937 and MV4–11 (CRL-9591) were purchased from the ATCC (Manassas, Virginia, USA), MOLM-13 (DSMZ Cat# ACC-554) and OCI-AML3 (DSMZ Cat# ACC-582) cells were purchased from DSMZ (Brunswick, Lower Saxony, Germany) and maintained as described previously [25 (link)]. All cell lines were tested for Mycoplasma contamination using the MycoAlert Mycoplasma Detection Kit (Lonza) routinely. All experiments utilized logarithmically growing cells (3–4×10⁵ cells/ml).
Venetoclax-resistant MV4–11 and MOLM-13 cells were obtained by culturing in the presence of increasing venetoclax (from 1nM to 1μM) concentrations over a period of 2 months.
Venetoclax was a gift from AbbVie (Chicago, IL, USA). INK128, AZD2014 was purchased from ChemieTek (Indianapolis, IN, USA). copanlisib was purchased from AdooQ (Irvine, CA, USA). MK2206 was purchased from MedChemExpress (Monmouth Junction, NJ, USA). Rapamycin was purchased from Cell Signaling (Danvers, MA, USA). All drugs were dissolved in DMSO, aliquoted, and stored at −80. Final DMSO concentrations did not exceed 0.1%.

Satta T., Li L., Chalasani S.L., Hu X., Nkwocha J., Sharma K., Kmieciak M., Rahmani M., Zhou L, & Grant S. (2023). Dual mTORC1/2 inhibition synergistically enhances AML cell death in combination with the BCL2 antagonist venetoclax. Clinical cancer research : an official journal of the American Association for Cancer Research, 29(7), 1332-1343.

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Monitoring Autophagy Dynamics in C17.2 Cells

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C17.2 cell were transfected with mRFP-GFP-LC3 plasmid obtained from Addgene (Cambridge, MA) or GFP-LC3 vector using Lipofectamine 2000 (Invitrogen, Carlsbad, CA) as previously described [30 (link),32 ], and treated for 48 h with trehalose (100 mM, Sigma) or for 24 h with rapamycin (100 μM, Cell Signaling Technology). Bafilomycin (10 nM, Sigma) was used to block autophagy for 4 h at 37 °C and analyze autophagosome accumulation. The cells were fixed with 4% paraformaldehyde in PBS for 15 min, and the nuclei were stained with DAPI (Invitrogen).

Xu C., Chen X., Sheng W.B, & Yang P. (2019). Trehalose restores functional autophagy suppressed by high glucose. Reproductive toxicology (Elmsford, N.Y.), 85, 51-58.

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Peptide-mediated Cell Signaling Modulation

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The TAT and TAT-vFLIP peptides were custom-synthesized with purity of more than 95% (Biomatik). Rapamycin and MG132 were purchased from Cell Signaling Technology. PU-H71 was purchased from Selleckchem. EBSS was purchased from Thermo Fisher Scientific. Recombinant TRAIL was purchased from BioLegend. Staurosporin, CCCP, 3-methyladenine, rotenone, bafilomycin A1, chloroquine, and cycloheximide were purchased from Sigma.

Choi Y.B., Choi Y, & Harhaj E.W. (2018). Peroxisomes support human herpesvirus 8 latency by stabilizing the viral oncogenic protein vFLIP via the MAVS-TRAF complex. PLoS Pathogens, 14(5), e1007058.

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Ventricular Cardiomyocyte Ablation in Zebrafish

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To perform ventricular CM ablation, Tg(vmhc:mCherry-NTR) larvae were treated with 6 mM MTZ (metronidazole, Sigma-Aldrich) in E3 water for 4 h at 3 dpf as previously described [11 (link)]. To modulate signaling pathways, larvae were incubated with the following chemicals for the indicated time period: 100 μM DAPT (Sigma-Aldrich), 12 μM AG1478 (Sigma-Aldrich), 5 μM cardiomogen-1 (Sigma-Aldrich), 7.5 μM dorsomorphin (Sigma-Aldrich), 5 μM LDN193189 (Selleck), or 10 μM rapamycin (Cell Signaling Technology). To stop blood flow, larvae were treated with 1.8 mM tricaine (3-aminobenzoic acid ethyl ester, Sigma-Aldrich) or 10 mM BDM (2,3-butanedione monoxime, Sigma-Aldrich) in E3 water for the indicated time period, and then washed with fresh E3 water.

Liang S., Zhou Y., Chang Y., Li J., Zhang M., Gao P., Li Q., Yu H., Kawakami K., Ma J, & Zhang R. (2024). A novel gene-trap line reveals the dynamic patterns and essential roles of cysteine and glycine-rich protein 3 in zebrafish heart development and regeneration. Cellular and Molecular Life Sciences: CMLS, 81(1), 158.

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Cytotoxicity Assay with LZ-101

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LZ-101 was dissolved in DMSO to make a 100-mM stock and stored at −20 °C until needed. The final concentration of DMSO did not exceed 0.1% throughout the study. 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), cycloheximide, 3-MA and diamidino-phenyl-indole (DAPI) were from Sigma (St. Louis, USA). Rapamycin was purchased form Cell Signaling Technology (Danvers, USA). Bovine serum albumin (BSA) was purchased from Roche (Mannheim, Germany).

Guo Y., Zhao Y., Zhou Y., Tang X., Li Z, & Wang X. (2019). LZ-101, a novel derivative of danofloxacin, induces mitochondrial apoptosis by stabilizing FOXO3a via blocking autophagy flux in NSCLC cells. Cell Death & Disease, 10(7), 484.

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Rapamycin and IL-1β Modulate Chondrocyte Behavior

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The third generation of normal human chondrocytes was cultured in serum-free-Dulbecco's modified Eagle media (DMEM) for 24 h. The rapamycin (CellSignaling Technology, Danvers, MA, USA) pre-treated cells were cultured with 10 nmol/L rapamycin for 2 h, and so did with 3-MA (Sigma, MO, USA). And then the cells were treated with IL-1β (Sigma) at a concentration of 5 ng/mL for 0, 12, 24, and 48 h. The control cells were only treated with IL-1β. Changes of the chondrocytes were observed by optical microscopy.

Li X., Yang X., Maimaitijuma T., Cao X.Y., Jiao Y., Wu H., Meng Z.C., Liu H., Guan Z.P, & Cao Y.P. (2019). Plant homeodomain finger protein 23 inhibits autophagy and promotes apoptosis of chondrocytes in osteoarthritis. Chinese Medical Journal, 132(21), 2581-2587.

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Bovine IFN-γ Modulation Assay

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Bovine IFN-γ was purchased from the Kingfisher Group (King-fisher Biotech, Inc., USA). Bafilomycin A1 and E64d were purchased from Abcam (UK). 3-Methyladenine (3-MA) was purchased from Santa Cruz Biotechnology (Santa Cruz Biotechnology Inc., Germany). Chloroquine and rapamycin were purchased from Cell Signaling Technology (USA). The amino acids were purchased from Nanjing Keygen Biotech. Co., Ltd.

Xia X., Che Y., Gao Y., Zhao S., Ao C., Yang H., Liu J., Liu G., Han W., Wang Y, & Lei L. (2016). Arginine Supplementation Recovered the IFN-γ-Mediated Decrease in Milk Protein and Fat Synthesis by Inhibiting the GCN2/eIF2α Pathway, Which Induces Autophagy in Primary Bovine Mammary Epithelial Cells. Molecules and Cells, 39(5), 410-417.

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Steroidogenic Pathway Analysis in Cells

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McCoy's 5A medium and 0.4% trypan blue were purchased from Invitrogen/GIBCO (Carlsbad, CA). Penicillin-streptomycin was purchased from Roche Diagnostics (Indianapolis, IN). BSA and anti-β-tubulin antibody were purchased from Sigma Chemical Co. (St. Louis, MO). Purified hCG was purchased from Dr. A. F. Parlow (National Hormone and Peptide Program, Torrance, CA). Forskolin was obtained from BIOMOL Research Laboratories (Plymouth Meeting, PA). Rapamycin, anti-mouse or anti-rabbit IgG horseradish peroxidase conjugates, anti-p-S6K1, and anti-S6K1 antibodies were purchased from Cell Signaling Technology (Beverly, MA). Anti-CYP11A1 was obtained from Abcam (Cambridge, MA). Femto Super Signal Chemiluminescence reagent and Restore stripping buffer were purchased from Pierce (Rockford, IL). Progesterone Enzyme Immunoassay (EIA) kit was purchased from Cayman Chemical (Ann Arbor, MI). Real-time PCR reagents, as well as the primers and probes for STAR (assay id: Hs00264912_m1), CYP11A1 (assay id: Hs00167984_m1), HSD3B1 (assay id: Hs01084547_gH), and 18S rRNA (assay id: Hs99999901_s1) were purchased from Applied Biosystems (Foster City, CA). All other reagents used were conventional commercial products.

Moravek M.B., Shang M., Menon B, & Menon K. (2016). HCG-Mediated Activation of mTORC1 Signaling Plays a Crucial Role in Steroidogenesis in Human Granulosa Lutein Cells. Endocrine, 54(1), 217-224.

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