Cell Counter Kit-8 (CCK-8, Dojindo Laboratories, Kumamoto, Japan) assays were used to assess the cytotoxicity of different formulations. Briefly, HepG2 cells were added to 96 well plates (4×104 cells well−1) and allowed to adhere. Then, they were treated with fresh media containing free drugs (different formulations, DOX: β-Lapachone = 1:0, 8:2, 6:4, 5:5, 4:6, 2:8, and 0:1) along with free DOX, BDOX-NCS, BDOX-CCS, β-Lapachone/BDOX-NCS, and β-Lapachone/BDOX-CCS. The equivalent DOX/BDOX concentrations were 0.01, 0.1, 1.0, 10.0, and 100.0 mg L−1, respectively. Subsequently, the cells were incubated for 24 h. After washing with PBS, the CCK-8 solution was added for 4 h. Finally, the absorbance was measured at 450 nm using a microplate reader (ELX 800, Bio Tec, USA). Percent viability was normalized based on untreated cells.
Cell counter kit 8
Manufactured by Dojindo Laboratories
Sourced in Japan
The Cell Counter Kit-8 is a colorimetric assay that can be used to measure the number of viable cells in a sample. The kit utilizes a water-soluble tetrazolium salt that is reduced by cellular dehydrogenase enzymes, producing a formazan dye that can be quantified using a spectrophotometer.
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Lab products found in correlation
Dulbecco s modified eagle s medium f12 dmem f12, by Thermo Fisher Scientific (1 mentions)
First strand cdna synthesis kit, by Takara Bio (1 mentions)
Trizol rna isolation reagent, by Thermo Fisher Scientific (1 mentions)
Sybr green reagent, by Takara Bio (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
N acetyl l cysteine, by Merck Group (1 mentions)
2 7 dichlorofluorescein diacetate, by Merck Group (1 mentions)
Sh sy5y cell, by American Type Culture Collection (1 mentions)
Crystal violet, by Merck Group (1 mentions)
6 protocols using cell counter kit 8
1
Cytotoxicity of DOX and β-Lapachone Formulations
2
Cell Proliferation Assay Using CCK-8
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Cell proliferation was assessed by using the Cell Counter Kit-8 (CCK-8; Dojindo, Kumamoto, Japan) as described in our previous studies 29 (link), 30 (link). In brief, cells were plated (6.0×103 cells per well for MDA-MB-231 and MCF-7; 5.0×103 cells per well for BT549; 4×104 cells per well for T-47-D) in 96-well plates and allowed to adhere overnight. The cells were then treated with metformin at various concentrations (0, 2.5, 5, 10, 15, or 20 mM) for 12, 24, 36, or 48 h before CCK-8 assay reagent (10µL) was added to each well. After that the OD values were determined at 450 nm, which were used to evaluate the relative cell viability. Transfected MDA-MB-231 and MCF-7 cells stably expressing pre-miR-200c, miR-200c inhibitor or empty vector and untransfected cells were seeded at a density of 4.5×103 cells in 96-well plates and incubated for various periods of time (0 to 5 days). Relative cell proliferation rate also was assessed by the OD values at 450 nm.
3
Sodium Hydrosulfide Modulates NLRP3 Inflammasome
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Sodium hydrosulfide (NaHS) was purchased from Chuandong Chemical Group Co. Ltd. (Chongqing, China). N-Acetyl-L-cysteine (NAC) and 2,7-dichlorofluorescein diacetate (DCFH-DA) were manufactured by Sigma-Aldrich (St. Louis, MO, USA). Cell Counter Kit-8 (CCK-8) was purchased from Dojindo (Kumamoto, Japan). Dulbecco's modified Eagle's medium F12 (DMEM/F12) and fetal bovine serum (FBS) were purchased from Gibco-BRL (Carlsbad, CA). TRIzol RNA isolation reagent was purchased from Invitrogen Life Technologies (New York, New York, USA). The first strand cDNA synthesis kit and SYBR green reagents were purchased from Takara (Dalian, China). siRNA of NLRP3 was obtained from Hanbio Biotechnology Co. Ltd. (Shanghai, China).
4
Assessing Cell Viability with CCK-8
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Cell viability of SH-SY5Y cells (ATCC, USA) was assessed by the Cell Counter Kit-8 (CCK-8) assay (Dojindo Laboratories, Japan) according to the manufacturer's instruction. SH-SY5Y cells purchased from the Chinese Type Culture Collection (Shanghai, China) were seeded in a 96-well plate and left to attach overnight. After the indicated treatments, 10 μM CCK-8 solution dissolved by serum-free medium was added to each well. The cells were incubated for 1 h in an incubator, and absorbance was quantified on an automated microplate reader (Bio-Tec, CA, USA).
5
Cell Viability and Colony Formation Assay
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Cell viability was detected by adding 10% Cell Counter Kit 8 (CCK-8, DOJINDO, Japan) into the cells plated in 96-well plates and incubated at 37 °C for 2 H at 0, 24, 48, 72 and 96 H. The absorbance of each well was measured by a microplate reader set at 450 and 630 nm. All experiments were performed in triplicates.
For colony formation assay, 500 KYSE30 and KYSE450 cells were maintained in each well of six-well plates and the medium was refreshed every 3 days. Colonies were fixed with methanol after 10 days for KYSE30 and 14 days for KYSE450, then stained with 0.1% crystal violet (Sigma-Aldrich, USA) for 30 min and washed with PBS. The numbers of colonies were counted.
For colony formation assay, 500 KYSE30 and KYSE450 cells were maintained in each well of six-well plates and the medium was refreshed every 3 days. Colonies were fixed with methanol after 10 days for KYSE30 and 14 days for KYSE450, then stained with 0.1% crystal violet (Sigma-Aldrich, USA) for 30 min and washed with PBS. The numbers of colonies were counted.
6
Cell Viability Assay using CCK-8
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Cell viability was assessed with the Cell Counter Kit 8 (CCK-8) assay (Dojindo Laboratories., Japan) according to the manufacturer's instruction. SH-SY5Y cells were seeded on 96-well plates and left to attach overnight. After treatments, 10 mM of the CCK-8 solution was dissolved by serum free DMEM/F12 medium and added to each well of the plate. Then, cells were incubated in the incubator for 1 h, and the absorbance at 450 nm was quantified by automated microplate reader (Synergy 2, BioTec, CA, USA).
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