To evaluate the effect of anticancer drugs, 1 × 104 cells were seeded into 96‐well culture plates and incubated for 24 h at 37°C with 5% CO2. We subsequently added cisplatin (Sigma‐Aldrich, St. Louis, MO, USA) (0, 25, 50, 100, 200 or 400 μM) or carboplatin (Sigma‐Aldrich) (0, 31.25, 62.5, 125, 250 or 500 μM). After 72 h (cisplatin) or 48 h (carboplatin) of incubation, 10 μL of Premix WST‐1 reagent (Takara Bio, Kyoto, Japan) were added to each well. After 1 h incubation, absorbance was measured. The absorbance at 450 nm was subtracted from the background absorbance (690 nm).
Premix wst 1 reagent
Manufactured by Takara Bio
Sourced in Japan
Premix WST-1 reagent is a colorimetric assay kit developed by Takara Bio for the quantitative measurement of cell viability and proliferation. The reagent contains tetrazolium salt WST-1, which is reduced by metabolically active cells, resulting in the formation of a colored formazan dye. The intensity of the color produced is directly proportional to the number of viable cells.
Automatically generated - may contain errors
Lab products found in correlation
Thermomixer, by Eppendorf (2 mentions)
Ultra low attachment surface, by Corning (2 mentions)
Tryple express, by Corning (2 mentions)
Epoch microreader, by Agilent Technologies (2 mentions)
Cisplatin, by Merck Group (1 mentions)
Carboplatin, by Merck Group (1 mentions)
Cisplatin, by Takara Bio (1 mentions)
Carboplatin, by Takara Bio (1 mentions)
Spectramax 190 microplate reader, by Molecular Devices (1 mentions)
Dmem without phenol red, by Thermo Fisher Scientific (1 mentions)
Non essential amino acids (neaa), by Thermo Fisher Scientific (1 mentions)
Glutamine, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Cytiva (1 mentions)
Infinite 200 pro, by Tecan (1 mentions)
Pyruvate, by Fujifilm (1 mentions)
Hepes naoh, by Merck Group (1 mentions)
Protein thermal shift starter kit, by Thermo Fisher Scientific (1 mentions)
Gtx1 15, by Alomone (1 mentions)
L glutamine, by Fujifilm (1 mentions)
Rpmi 1640, by Fujifilm (1 mentions)
7 protocols using premix wst 1 reagent
1
Anticancer Drug Cytotoxicity Evaluation
2
Transduction of Cells with AAV2
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The cells were seeded in 96-well plates at 1×103 cells per well. After 12 h, the cells were transduced with AAV2-shE6E7 at 0–1×107 vg/cell or AAV2-shNC at 1×105 vg/cell. After 72 h, 10 μl premix WsT-1 reagent (Takara Bio, Otsu, Japan) was added, and the absorbance at 450 nm was measured using a SpectraMax 190 microplate reader (Molecular Devices, Sunnyvale, CA, USA) after 1 h.
3
Cytochrome P450 Reporter Assay in HepG2 Cells
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ELuc-HepG2 or CYPs-ELuc-HepG2 cells were seeded in 96-well black clear bottom plates (Nunc, Wiesbaden, Germany) at a density of 1 × 104 cells per well. After an overnight incubation, the medium was replaced with DMEM without phenol red (Gibco-BRL) but with 10% FBS (HyClone Laboratories), 4 mM glutamine, non-essential amino acids (Gibco-BRL), 1 mM pyruvate (FUJIFILM Wako Pure Chemical Corporation), 25 mM HEPES/NaOH (pH 7.0, Sigma–Aldrich), and 300 μM D-luciferin potassium salt (Resem B.V., Lijnden, The Netherlands) with or without compound. After incubation for 3 days in a CO2 incubator, ELuc luminescence intensity in the cells was measured. The 96-well plate was set in a microplate-type luminometer (AB-2350 Phelios; ATTO, Tokyo, Japan) and luminescence was measured without disrupting the cells for 5 s. For the WST-1 assay, Premix WST-1 reagent (Takara Bio Inc., Shiga, Japan) was added to the remaining HepG2 cells and incubation was carried out for 1 to 2 h at 37 °C. Absorbance was measured at 450 nm (reference 620 nm) using the microplate reader Infinite 200 PRO (Tecan).
4
Evaluating Epigenetic Drug Toxicity in 2D Tumor Cells and Organoids
5
Neuro2a Cell Viability Assay
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Neuro2a cells were cultured in complete medium for 24 h under an atmosphere of 5% CO2/air at 37 °C. Cells were transfected with the LNPs. It was then replaced with complete medium and incubated for 1 h under an atmosphere of 5% CO2/air at 37 °C. The medium was replaced with complete medium containing PremixWST-1 reagent (TAKARA Bio Inc., Shiga, Japan) and incubated for 2 h under an atmosphere of 5% CO2/air at 37 °C. Absorbance was detected at 440 and 650 nm using a microplate reader. Relative cell viability was calculated by normalizing the cell viability of the control.
where Vt and Vu represent the cell viability for treated and untreated cells with samples, respectively.
where Vt and Vu represent the cell viability for treated and untreated cells with samples, respectively.
6
Thermal Shift Assay for THP-1 Cells
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GTx1-15 was obtained from Alomone Labs (Jerusalem, Israel). Human plasma was purchased from KOJIN BIO (Saitama, Japan). The protein thermal shift starter kit was from Thermo Fisher Scientific (Tokyo, Japan). RPMI-1640 with L-glutamine and phenol red, sodium hydrogen carbonate, and Tris-HCl were purchased from Wako Chemicals (Osaka, Japan). A human monocytic leukemia cell line, THP-1 (No. JCRB0112), was from the Japanese Collection of Research Bioresources Cell Bank (Osaka, Japan). Penicillin-streptomycin solution was obtained from Life Technologies Japan Ltd. (Tokyo, Japan). HyClone fetal bovine serum was purchased from GE Healthcare Japan (Tokyo, Japan). Premix WST-1 reagent was purchased from Takara Bio (Shiga, Japan).
7
Evaluating Epigenetic Drug Toxicity in 2D Tumor Cells and Organoids
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For 2D tumour cell culture, tumour cells were seeded into 96 well plates at 2,000 cells per well and cultured for 72 h. For tumour organoids, tumour organoids with diameter between 70 and 150 μm were seeded at 50 organoids per well into 96-well microplates with ultra-low attachment surface (Corning). Cells or tumour organoids were treated with indicated compound at the final concentration of 1μM. After 3-day culture, tumour organoids were dissociated into single cells using TrypLE Express (Corning) at 37°C with a shaking velocity of 500 rpm on a thermomixer (Eppendorf) for 15 min. The 96-well microplates were added with 10 μl/well of Premix WST-1 reagent (TaKaRa) and incubated for 1 h. The microplates were read at absorbance of 450 nm on the Epoch microreader (BioTek). The toxicity of the epigenetic drugs was quantified using the Premix WST-1 reagent according to the manufacturer’s protocol.
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