Cell Meter™ Colorimetric WST-8 Cell Quantification Kit

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Telephone	1-800-990-8053
Fax	1-800-609-2943
Email	sales@aatbio.com
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Storage, safety and handling

Certificate of Origin	Download PDF
H-phrase	H303, H313, H333
Hazard symbol	XN
Intended use	Research Use Only (RUO)
R-phrase	R20, R21, R22
Storage	Freeze (< -15 °C); Minimize light exposure
UNSPSC	12352200

Overview SDS Protocol

Our Cell Meter™ assay kits are a set of tools for monitoring cell viability. There are a variety of parameters that can be used for monitoring cell viability. Cell Meter™ Colorimetric WST-8 Cell Quantification Kit uses water-soluble WST-8 tetrazolium salt to quantify the number of live cells. The water-soluble WST-8 tetrazolium salt produces a water-soluble orange formazan dye upon bioreduction in the presence of an electron carrier, 1-methoxy-5-methylphenazinium methyl sulfate. The kit is convenient and robust with a mix and read format. WST-8 solution is added directly to the test cells, no pre-mixing of components is required. WST-8 tetrazolium salt is reduced by cellular dehydrogenases to an orange formazan product that is soluble in tissue culture medium. The amount of formazan produced is directly proportional to the number of living cells by monitoring absorbance increase at 460 nm. The excellent stability and little cytotoxicity of WST-8 solution make the kit useful for the assays that require long incubation (such as 24 to 48 hours). Cell Meter™ Colorimetric WST-8 Cell Quantification Kit provides a sensitive colorimetric assay for the determination of the number of viable cells in the proliferation and cytotoxicity assays. The detection sensitivity is higher than any other tetrazolium salt-based assays such as MTT, XTT or MTS etc.

Platform

Absorbance microplate reader

Absorbance	460 nm
Recommended plate	Clear bottom

Example protocol

AT A GLANCE

Protocol Summary

Prepare cells in a 96-well plate (100 µL/well).
Add 10 µL of WST-8™ Solution to each well.
Incubate at 37°C for 1 - 4 hours.
Monitor absorbance at OD = 460 nm.

Important Note

The WST-8™ Solution can be stable for up to one year if stored at 4°C and protected from light. Store it at <-20°C for longer storage.

CELL PREPARATION

For guidelines on cell sample preparation, please visit:

https://www.aatbio.com/resources/guides/cell-sample-preparation.html

SAMPLE EXPERIMENTAL PROTOCOL

Cell Proliferation and Cytotoxicity Assay:

Plate 5000 to 10,000 cells/well in a tissue culture microplate with clear bottom.
Add test compounds into the cells and incubate for a desired period of time (such as 24, 48, or 96 hours) in a 37°C, 5% CO₂ incubator. For blank wells (medium without the cells), add the same amount of test compounds. The suggested total volume is 100 µL for a 96-well plate or 50 µL for a 384-well plate.

Note: Each cell line should be evaluated on an individual basis to determine the optimal cell density for proliferation or cytotoxicity induction. For proliferation assays, use fewer cells; for cytotoxicity assays, use more cells to start with.
Add 10 µL/well (96-well plate) or 5 µL/well (384-well plate) of WST-8™ Solution to each well.
Incubate the plate at 37°C for 1 - 4 hours, protect from light.

Note: The incubation time could be from 30 minutes to overnight depending on the individual cell type and cell concentration used. Optimize the incubation time for each experiment.
Monitor the absorbance increase with an absorbance plate reader at OD =460 nm.

Cell Counting Assay:

Prepare cell culture in a tissue culture microplate with a clear bottom. The suggested total volume is 100 µL for a 96-well plate or 50 µL for a 384-well plate.

Note: We used serially diluted HeLa and Jurkat cell suspension in a clear bottom 96-well plate for the assay.
Add 10 µL/well (96-well plate) or 5 µL/well (384-well plate) of WST-8™ Solution to each well.
Incubate the plate at 37°C for 1 - 4 hours, protect from light.

Note: The incubation time could be from 30 minutes to overnight depending on the individual cell type and cell concentration used. Optimize the incubation time for each experiment.
Monitor the absorbance increase with an absorbance plate reader at OD = 460 nm.

Images

Figure 1. Cell number was determined with Cell Meter™ Colorimetric WST-8 Cell Quantification Kit. HeLa cells at 40 to 10,000 cells/well/100 µL were added in a clear bottom 96-well plate. The absorbance was measured at 460 nm using a SpectraMax reader (Molecular Devices).

Figure 2. Cytotoxicity tests of HeLa cells in response to (A) SDS and (B) Staurosporine treatment were measured with Cell Meter™ Colorimetric WST-8 Cell Quantification Kit. HeLa cells at 10,000 cells/well/100 µL were seeded overnight in a black wall/clear bottom 96-well plate. Cells were treated with serially diluted SDS for 2 hours or Staurosporine for 4 hours. The absorbance was measured at 460 nm using a SpectraMax reader.

Figure 3. Cell number was determined with Cell Meter™ Colorimetric WST-8 Cell Quantification Kit. Jurkat cells at 60 to 150,000 cells/well/100 µL were added in a clear bottom 96-well plate. The absorbance was measured at 460 nm using a SpectraMax reader (Molecular Devices).

Figure 4. FGF10 increased the viability of primary mouse corneal epithelial cells. Mouse corneal epithelial cells were seeded in multi-well dishes, treated with FGF10 (30 ng/mL) or BSA (control), and cultured for 1, 3, and 7 days. Cells were processed for viability assay (tetrazolium salt, WST- 8). Dots represent the number of repeats (n = 9) for each condition/time. Statistical significance was assessed with unpaired t-tests. All bar plots are mean ± SD. *P < 0.05; ****P < 0.0001. Source: Role of FGF10/FGFR2b Signaling in Homeostasis and Regeneration of Adult Lacrimal Gland and Corneal Epithelium Proliferation by Findburgh et.al., Investigative Ophthalmology & Visual Science Jan. 2023.

Citations

View all 3 citations: Citation Explorer

Role of FGF10/FGFR2b Signaling in Homeostasis and Regeneration of Adult Lacrimal Gland and Corneal Epithelium Proliferation
Authors: Finburgh, Emma N and Mauduit, Olivier and Noguchi, Takako and Bu, Jennifer J and Abbas, Anser A and Hakim, Dominic F and Bellusci, Saverio and Meech, Robyn and Makarenkova, Helen P and Afshari, Natalie A
Journal: Investigative Ophthalmology \& Visual Science (2023): 21--21

MicroRNA-30a suppresses self-renewal and tumorigenicity of glioma stem cells by blocking the NT5E-dependent Akt signaling pathway
Authors: Peng, Lilei and Ming, Yang and Zhang, Ling and Zhou, Jie and Xiang, Wei and Zeng, Shan and He, Haiping and Chen, Ligang
Journal: The FASEB Journal (2020): 5128--5143

miR-218 inhibits the proliferation of human glioma cells through downregulation of Yin Yang 1
Authors: Gao, Yong and Sun, Laisheng and Wu, Zicheng and Xuan, Chengmin and Zhang, Junxia and You, Yongping and Chen, Xincheng
Journal: Molecular Medicine Reports (2017)

References

View all 5 references: Citation Explorer

A sensitive WST-8-based bioassay for PEGylated granulocyte colony stimulating factor using the NFS-60 cell line
Authors: Tiwari, K.; Wavdhane, M.; Haque, S.; Govender, T.; Kruger, H. G.; Mishra, M. K.; Ch and ra, R.; Tiwari, D.
Journal: Pharm Biol (2015): 849-54

Comparison of the WST-8 colorimetric method and the CLSI broth microdilution method for susceptibility testing against drug-resistant bacteria
Authors: Tsukatani, T.; Suenaga, H.; Shiga, M.; Noguchi, K.; Ishiyama, M.; Ezoe, T.; Matsumoto, K.
Journal: J Microbiol Methods (2012): 160-6

WST-8 analysis of cell viability during osteogenesis of human mesenchymal stem cells
Authors: Stoddart, M. J.
Journal: Methods Mol Biol (2011): 21-5

monosodium salt (WST-8) rapid colorimetric assays for antimicrobial susceptibility testing of staphylococci and ESBL-producing clinical isolates
Authors: Brady, A. J.; Kearney, P.; Tunney, M. M., Comparative evaluation of 2,3-bis [2-methyloxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide (XTT) and 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2, 4-disulfophenyl)-2H-tetrazolium
Journal: J Microbiol Methods (2007): 305-11

Reactivity of blood samples spotted onto filter papers in the WST-8 method for screening of G6PD deficiency
Authors: Arai, M.; Kosuge, K.; Kawamoto, F.; Matsuoka, H.
Journal: Acta Med Okayama (2006): 127-34