Atorvastatin calcium, Mevalonic acid, GGPP, FPP, Propidium iodide (PI) and 2′,7′-dichlorofluorescein diacetate (DCFH-DA) were purchased from Sigma-Aldrich (St. Louis, MO, USA). 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) was from Amresco (Solon, OH, USA). 5,5′,6,6′-tetrachloro-1, 1′,3,3′-tetraethylbenzimidazole-carbocyanide iodine (JC-1) and cell mitochondria isolation kit were obtained from Beyotime Biotechnology (Shanghai, China). Annexin V-FITC Apoptosis Detection Kit, as well as antibodies against p-pRb (S780) (1:1,000), p27 (1:1,000) and cyclinD1 (1:1,000) were from BD Biosciences Pharmingen (San Jose, CA, USA). Antibodies against pRb (1:1,000), cyclinB1 (1:2,000), cdc2 (1:1,000), caspase-3 (1:1,000), caspase-9 (1:1,000), poly (ADP-ribose) polymerase (PARP) (1:1,000), Cytochrome c (1:1,000), YAP (1:1,000), p-YAP (Ser127) (1:1,000), Rho A (1:1,000), β-actin (1:1,000), anti-mouse (1:2,000), anti-rabbit HRP-conjugated and anti-rabbit Alexa Fluor 488-conjugated secondary antibodies (1:2,000) were from Cell Signaling Technology (Danvers, MA, USA). Antibodies against Bcl-2 (1:1,000), Bax (1:1,000) and Lamin B (1:500) were from Santa Cruz Biotechnology (Santa Cruz, CA, USA).
Mevalonic acid
Manufactured by Merck Group
Sourced in United States
Mevalonic acid is a chemical compound that serves as a precursor in the biosynthesis of various important molecules, including cholesterol, steroid hormones, and certain vitamins. It is a key intermediate in the mevalonate pathway, which is responsible for the production of these essential biological compounds. As a lab equipment product, mevalonic acid is utilized in research and analysis related to these metabolic processes.
Automatically generated - may contain errors
Lab products found in correlation
Lovastatin, by Merck Group (5 mentions)
Simvastatin, by Merck Group (3 mentions)
Cholesterol, by Merck Group (3 mentions)
Mevastatin, by Merck Group (3 mentions)
Squalene, by Merck Group (3 mentions)
Hplc grade solvents, by Thermo Fisher Scientific (2 mentions)
Sphingosine 1 phosphate (s1p), by Merck Group (2 mentions)
Leukotriene c4, by Cayman Chemical (2 mentions)
Geranylgeranyl pyrophosphate, by Cayman Chemical (2 mentions)
Ibuprofen, by Merck Group (2 mentions)
Glutathione (gsh), by Merck Group (2 mentions)
Indomethacin, by Merck Group (2 mentions)
Lovastatin, by LKT Laboratories (2 mentions)
Azadirachtin, by Merck Group (2 mentions)
Acetyl coa, by Merck Group (2 mentions)
Ro 3306, by Merck Group (2 mentions)
Cytochrome c, by Cell Signaling Technology (1 mentions)
Atorvastatin calcium, by Merck Group (1 mentions)
3 4 5 dimethyl 2 thiazolyl 2 5 diphenyl 2h tetrazolium bromide mtt, by Avantor (1 mentions)
Cyclin d1, by BD (1 mentions)
25 protocols using mevalonic acid
1
Atorvastatin Inhibits Proliferation and Induces Apoptosis in Cancer Cells
2
Coculture of HepG2 cells with PBMCs or B-cell leukemia cells
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For cocultures, 125 000 HepG2 cells per well were plated in 24-well plates, allowed to proliferate for ≈70 hours, washed with PBS, and subsequently cultured in coculture medium (DMEM with 4.5 g/L glucose, GlutaMAX, and pyruvate [Gibco] supplemented with 10% lipoprotein-depleted human serum, 100 IU/mL penicillin, 100 µg/mL streptomycin, 5 μM simvastatin [Sigma-Aldrich, Zwijndrecht, the Netherlands], and 10 μM mevalonic acid [Sigma-Aldrich]). PBMCs were isolated from whole blood and resuspended in the coculture medium at a concentration of 1.7 million cells/mL. Of this suspension, 350 µL was added to a 6.5-mm diameter transwell insert with a 0.4-um pore size (Corning, Corning, NY) that were placed on top of the HepG2 cells in the 24-well plate. After 24 hours of coculture, HepG2 cells were either collected for gene expression analysis, used for LDL uptake studies, or analyzed for LDLR protein expression. Using a similar setup, HepG2 cells were cocultured with B-cell precursor acute lymphoblastic leukemia cells Kasumi-2 and Nalm6 instead of isolated human PBMCs.
3
Preparation of Stock Solutions
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Simvastatin (Sigma, NJ, USA, S6196) was dissolved in dimethyl sulfoxide (DMSO) (Sigma, D2650-5X) to make concentrations of 1 and 2 mg/mL as stock solutions. Mevalonic acid (Sigma, 50838) and cholesterol (Sigma, C4555-1G) were dissolved in deionized water (DW) to make stock concentrations of 10 and 50 mg/mL, respectively. The stock solution was sterilized by filtration using a 0.22 µm-pore-size membrane. FPP (Sigma, F6892) and GGPP (Sigma, G6025) were stored at −20 °C and directly diluted in PBS before use.
4
LXR-Dependent Signaling Pathway Activation
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Isolated cells were treated with agonists to stimulate LXR-dependent signaling. GW3965 (GW) (Sigma) was used at a concentration of 1 or 2 μM for 6 h for RNA isolation and 16 h for protein collection and lipidomic profiling. 22R-hydroxycholesterol was dissolved in ethanol and used at a concentration of 5 μM for 16 h in combination with sterol-depleted medium [RPMI with 10% lipoprotein-deficient serum, penicillin (100U/ml), streptomycin (100ug/ml) (Gibco), 2.5 μg/ml simvastatin (Calbiochem), and 100 μM mevalonic acid (Sigma)]. T0901317 (T09) (Cayman Chemical) was used at a concentration of 10 μM for 24 h. Acetylated LDL (AcLDL) (Kalen Biochemicals) was added to the culture medium at a final concentration of 50 μg/ml for 24 h.
5
Lipid Synthesis and Signaling Assay
6
Activation of Statin Compounds
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Atorvastatin (Pfizer Pharmaceuticals, Inc, Groton, Connecticut, USA), cerivastatin (Bayer Corp, West Haven, Connecticut, USA), fluvastatin (Novartis Pharma AG, Basel, Switzerland), lovastatin, and simvastatin (Merck, Sharp & Dohme Res Lab, Rahway, New Jersey, USA) were used throughout the experiments. Lovastatin and simvastatin were obtained in the inactive lactone form and were activated by dissolving in 100% ethanol, heating for 2 h at 50°C in 0.1N NaOH and neutralizing to physiological pH with 1N HCl. Distilled water was added to obtain the final stock concentration of 10 mM. Stock solution was aliquoted and stored frozen at −20°C. Puromycin, mevalonic acid (MA), methyl-β-cyclodextrin (MβCD), and water-soluble cholesterol were purchased from Sigma.
7
Synchronizing HCT116 Cells in G1 Phase
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HCT116 cells were synchronized in the G1 phase as described previously (Javanmoghadam-Kamrani and Keyomarsi 2008 (link)). Briefly, asynchronously growing HCT116 cells were treated with 20 µM lovastatin (LKT Laboratories) for 24 h to arrest them in G1 phase. Following that, the cells were washed twice with fresh medium and then grown in medium containing 2 mM mevalonic acid (Sigma-Aldrich).
8
Lipid Synthesis and Signaling Assay
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Indomethacin, ibuprofen, mevalonic acid, mevastatin, S1P, and glutathione were purchased from Sigma. HPLC-grade solvents were purchased from Fisher. Leukotriene C4, D4, and geranylgeranyl-pyrophosphate was purchased from Cayman Chemical. LPI was purchased from Avanti Polar Lipids. ETYA and AACOCF3 were purchased from Biomol. Ggg was chemically synthesized using the protocol specified in the section below.
9
Synchronizing HCT116 cells for Repli-Seq
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HCT116-RAD21-mAC cells were synchronized in G1 with Lovastatin following a protocol of ref. 71 (link). Cells were incubated with 20 µM Lovastatin (Mevinolin) (LKT Laboratories M1687) for 24 h to synchronize in G1. Five-hundred micromolar auxin or DMSO was added 6 h before release from Lovastatin block. To release from G1 block, Lovastatin was washed away with three washes of PBS and warm media plus 2 mM Mevalonic acid (Sigma-Aldrich M4667) and 500 μM auxin or DMSO. Cells were released for 10, 14, 18, and 22 h. Two hours before the time point 100 μM BrdU was added to label nascent replication. After fixation, equal numbers of cells from each release time point were pooled together for early/late repli-seq processing36 (link). Repli-Seq data were processed as described in ref. 36 (link). In brief, data were aligned to the hg19 reference genome using bowtie2, deduplicated with samtools, and the log-2 ratio between early and late timepoints was calculated.
10
Isolation and Stimulation of Naïve CD4+ T Cells
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CD4+ T cells from the spleen and peripheral LNs were positively selected with CD4 microbeads (L3T4; Miltenyi Biotec), and naive CD4+ T cells were further sorted as CD4+CD25−CD62LhighCD44low cells with the FACSAria III cell sorter (Becton, Dickinson and Company (BD) Biosciences) and stimulated with plate-coated anti-CD3 (1 μg/mL, 145-2C11: BioXCell) and soluble anti-CD28 (1 μg/mL, 37.51; BioXCell) for 3 d in RPMI 1640 (Sigma-Aldrich) supplemented with 10% FBS (Sigma-Aldrich). To measure TCR-mediate protein phosphorylation, flow-sorted naïve CD4+ T cells were stimulated with plate-coated anti-CD3 and anti-CD28 (2 μg/mL) before being analyzed at indicated time points. In some experiments, cells were treated with either DMSO vehicle or 1 µM GW3965 (Tocris), 1 µM BIO (Sigma-Aldrich), or mevalonic acid (Sigma-Aldrich).
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