Mts reagent

Manufactured by Promega

Sourced in United States, United Kingdom, China, Switzerland, France

The MTS reagent is a tetrazolium compound used in cell viability and proliferation assays. It is a colorimetric assay that measures the reduction of MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) to a formazan product by metabolically active cells. The absorbance of the formazan product is proportional to the number of living cells in the culture.

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Close spelling variants of this product

RealTime-Glo™ MT Cell Viability Assay reagent 3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS reagent) MTS/MPS Cell Titer 96® One solution Reagent MTS labeling reagent CellTiter 96® AQueous MTS Reagent Solution MTS assay reagent 3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-tetrazolium (MTS) reagent MTS reagent powder (G1111) MTS labeling reagent solution CellTiter 96® AQueous MTS Reagent Powder

397 protocols using mts reagent

Comparative Efficacy of Targeted Fusion Antibodies

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Example 12

Fusion Abs of the disclosure were tested in several cell lines to compare the effectiveness of targeted fusion Abs versus non-targeted fusion Abs. Briefly, 1×10e4 cells/well (50 uL/well) of each cell line were seeded into 96-well tissue culture plates (Becton Dickinson). The next day cells were treated with 50 uL/well of each indicated Ab at the indicated concentrations. Six (6) days later, 20 uL/well MTS reagent (Promega) was added to the plates. Absorbance changes are read at 490 nm with a Biotek EPOCH reader. The results show that the Targeted Fusion Abs are more effective than the Non-Targeted Fusion Abs. (See, FIG. 15(A) & FIG. 15(B)).

In another experiment, 1.5×10e4 U266 cells/well (50 uL/well) or 1×10e4 CAPAN-2 cells/well (50 uL/well) were seeded into 96-well tissue culture plates (Becton Dickinson). U266 cells were treated the same day, while CAPAN-2 cells were treated the next day with 50 uL/well of each indicated Ab at the indicated concentrations. Six (6) days after treatment, 20 uL/well MTS reagent (Promega) was added to the plates. Absorbance changes are read at 490 nm with a Biotek EPOCH reader. The results show that the Targeted Fusion Abs are more effective than the Non-Targeted Fusion Abs. In addition, the masked Fusion Ab is less effective than the cleaved version. (See, FIG. 16(A) & FIG. 16(B)).

US11136353B2. Fusion protein composition(s) comprising masked type I interferons (IFNA and IFNB) for use in the treatment of cancer and methods thereof (2021-10-05). Qwixel Therapeutics LLC [US]. Inventors: David Stover [US], Sherie Morrison [US], Alex Vasuthasawat [US], Kham Trinh [US], George Ayoub [US].

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Assessing FRG1 Impact on Cell Viability

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HEK293T cells (0.1 × 10³) were seeded in a 96-well plate. Cells were transfected with FRG1 expression and silencing vectors, along their respective controls, using Xtremegene9. Transfected cells were grown for 96 h. Afterward, the medium was replaced with fresh medium and 20 µl of MTS reagent (Promega). Absorbance was recorded at 490 nm wavelength using iMark Microplate Reader (Bio–Rad).
HUVECs (0.5 × 10³) were seeded in a 96-well plate in HiEndoXL Endothelial Cell Growth Medium (HiMedia). Post 24 h of seeding, endothelial cell growth medium was replaced with conditioned medium obtained from HEK293T cells transfected with FRG1 expression vector and respective vector control. HUVECs were grown in the respective conditioned medium for 96 h, followed by addition of fresh medium and 20 µl of MTS reagent (Promega).

Tiwari A., Pattnaik N., Mohanty Jaiswal A, & Dixit M. (2017). Increased FSHD region gene1 expression reduces in vitro cell migration, invasion, and angiogenesis, ex vivo supported by reduced expression in tumors. Bioscience Reports, 37(5), BSR20171062.

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Comparative Analysis of Targeted Fusion Antibodies

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Example 12

US11795198B2. Fusion protein composition(s) comprising masked type I interferons (IFNA and IFNB) for use in the treatment of cancer and methods thereof (2023-10-24). Qwixel Therapeutics LLC [US]. Inventors: David Stover [US], Sherie Morrison [US], Alex Vasuthasawat [US], Kham Trinh [US], George Ayoub [US].

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Evaluating Cytotoxic and Antiproliferative Effects of rHct-S3

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To determine cytotoxic activity of rHct-S3, cells (1.0 × 10⁴) were seeded in 96-well plates (“Jet Biofil”, Guangzhou, China) and cultured in 200 µL of complete culture medium for 24 h at 37 °C in a 5% CO₂ incubator. The cell monolayer was washed with PBS and treated either with PBS (control) or various concentrations of rHct-S3 (0.01 µM–10 µM) in fresh appropriate culture medium for 24 h. Subsequently, the cells were incubated with 15 µL MTS reagent (“Promega”, Madison, WI, USA) for 3 h, and the absorbance of each well was measured at 490/630 nm using Power Wave XS microplate reader (“BioTek”, Wynusky, VT, USA).
To determine the antiproliferative activity of rHct-S3, cells (0.7 × 10⁴) were seeded in 96-well plates and cultured in 200 µL of complete culture medium for 24 h at 37 °C in a 5% CO₂ incubator. The cell monolayer was washed with PBS and treated either with PBS (control) or various concentrations of rHct-S3 (1, 2, and 4 µM) in fresh appropriate culture medium for 24, 48, 72, 96 h. Subsequently, the cells were incubated with 15 µL MTS reagent (“Promega”, Madison, WI, USA) for 3 h, and the absorbance of each well was measured at 490/630 nm using Power Wave XS microplate reader (“BioTek”, Wynusky, VT, USA).

Kvetkina A., Malyarenko O., Pavlenko A., Dyshlovoy S., von Amsberg G., Ermakova S, & Leychenko E. (2020). Sea Anemone Heteractis crispa Actinoporin Demonstrates In Vitro Anticancer Activities and Prevents HT-29 Colorectal Cancer Cell Migration. Molecules, 25(24), 5979.

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Quantitative Cell Viability Assays

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For IC50 assay, DU145 (1000/well), PC-3 (1000/well), C4-2 (2000/well) or LNCaP cells (2000/well) were seeded in 96-well plates overnight. Cells were then treated with different doses of the indicated compounds. At 72 h after treatment, MTS reagent (Promega) was added to each well and the plates were incubated at 37 °C for 2 h. Optical density (OD) of cells was measured at 490 nm using microtiter reader (Biotek). For cell growth curve assay, PC-3 cells (800/well) or C4-2 cells (1500/well) were seeded in 96-well plates overnight and treated with indicated chemicals. At the indicated time points, MTS reagent (Promega) was added to each well and the plates were incubated at 37 °C for 2 h. Optical density (OD) of cells was measured at 490 nm using microtiter reader (Biotek).

Ding D., Zheng R., Tian Y., Jimenez R., Hou X., Weroha S.J., Wang L., Shi L, & Huang H. (2022). Retinoblastoma protein as an intrinsic BRD4 inhibitor modulates small molecule BET inhibitor sensitivity in cancer. Nature Communications, 13, 6311.

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Cisplatin Sensitivity Assay in Bladder Cancer

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Experiments were performed in 6-well plates after cell density reached to about 90% (2 × 10³/well). TCCSUP or T24 cells were transfected with various constructs or siRNAs and cisplatin simultaneously for 48 h (siRNA added 2 h before cisplatin). Cells were then incubated with cisplatin containing serum-free medium (RPMI1640 or DMEM) for the indicated time. Cell viability was determined using MTS reagents, as instructed by the company’s protocol (Promega Corporation, Madison, WI).

Mansour A.M., Abdelrahim M., Laymon M., Elsherbeeny M., Sultan M., Shokeir A., Mosbah A., Abol-Enein H., Awadalla A., Cho E., Sairam V., Park T.D., Shahid M, & Kim J. (2018). Epidermal growth factor expression as a predictor of chemotherapeutic resistance in muscle-invasive bladder cancer. BMC Urology, 18, 100.

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Evaluating Cisplatin Resistance in Bladder Cancer

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For transient cell transfections of siRNA oligonucleotides, T24R cells were grown to 80% confluence in 6-well culture plates and transfected with siRNAs targeting CDK2, CHEK1, ERBB2, and control (ThermoScientific) using Lipofectamine 2000 (Invitrogen), according to the manufacturer’s instructions. Cisplatin-resistant BC cells (T24R, J82R, and RT4R) were incubated with cisplatin and/or CDK2 inhibitor for 72 hr. Post-treatment cell survival was determined by measuring cell viability using MTS reagents (Promega Corporation, Madison, WI), according to the provided protocols.

Jung J.H., You S., Oh J.W., Yoon J., Yeon A., Shahid M., Cho E., Sairam V., Park T.D., Kim K.P, & Kim J. (2018). Integrated proteomic and phosphoproteomic analyses of cisplatin-sensitive and resistant bladder cancer cells reveal CDK2 network as a key therapeutic target. Cancer letters, 437, 1-12.

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Evaluating Gefitinib's Effect on NSCLC Cell Proliferation

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Cell proliferation was assessed using MTS and clonogenic assays. For the MTS assay, stably transfected NSCLC cell lines were seeded (2‐5 × 10³ cells/well in 200 μL) into 96‐well plates and divided into the gefitinib group (four wells) and the control group (four wells). The cells were later treated with gefitinib (1 μmol/L) or vehicle (DMSO) according to their respective groups. After incubation for an additional 0, 24, 48, or 72 hours, 20 μL MTS reagents (Promega, Madison, WI, USA) were added to each well. The absorbance at 490 nm of each well was measured on a spectrophotometer after incubation for 1 hour. For the clonogenic assay, stably transfected NSCLC cell lines were seeded (1 × 10³ cells/well) in a 6‐well plate and divided into the gefitinib and control groups and then treated with gefitinib (1 μmol/L) or vehicle (DMSO). After 1‐2 weeks of culture, colonies were stained with crystal violet and analyzed using Image‐Pro Plus 7.0 software.

Feng W., Xie Q., Liu S., Ji Y., Li C., Wang C, & Jin L. (2018). Krüppel‐like factor 4 promotes c‐Met amplification‐mediated gefitinib resistance in non‐small‐cell lung cancer. Cancer Science, 109(6), 1775-1786.

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Cisplatin Cytotoxicity in T24 Cells

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T24S and T24R cell lines were incubated with 10 mM cisplatin for 2 days. Cell survival was determined by measuring cell viability using MTS reagents, according to the company’s protocols (Promega Corporation, Madison, WI).

Lee M.Y., Yeon A., Shahid M., Cho E., Sairam V., Figlin R., Kim K.H, & Kim J. (2018). Reprogrammed lipid metabolism in bladder cancer with cisplatin resistance. Oncotarget, 9(17), 13231-13243.

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Evaluating Drug Resistance in Breast Cancer Cells

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Tamoxifen- or fulvestrant-resistant T47D or MCF7 cells were plated in triplicate in 96-well plates (3,000 cells/well) in the appropriate growth medium. At the indicated time points, the cells were placed in 90 μl of fresh growth medium supplemented with 10 μl of MTS reagents (Promega) and incubated at 37°C for 2 h. The absorbance value was measured at 490 nm using a 96-well plate reader. The experiments were repeated three times with similar results.

Yan C., Gao R., Gao C., Hong K., Cheng M., Liu X., Zhang Q, & Zhang J. (2023). FDXR drives primary and endocrine-resistant tumor cell growth in ER+ breast cancer via CPT1A-mediated fatty acid oxidation. Frontiers in Oncology, 13, 1105117.

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