WST-8

What Is WST-8?

WST-8 (Water-Soluble Tetrazolium Salt-8) is a colorimetric reagent commonly used in cell viability and proliferation assays, particularly in the CCK-8 (Cell Counting Kit-8) assay. It is a tetrazolium salt that is reduced by cellular dehydrogenases in metabolically active cells, producing a water-soluble formazan dye with an absorbance peak at 450 nm, which allows for easy quantification.

The CAS number for it is 193149-74-5, and its molecular formula is C20H14N6Na2O11S. It has a molecular weight of approximately 617.44 g/mol. In its pure form, it appears as a light yellow to orange powder that is highly water-soluble, making it ideal for biological applications. It is stable under standard laboratory conditions but should be stored away from light and moisture to maintain its reactivity.

Compared to other tetrazolium salts like MTT, WST-8 has the advantage of producing a water-soluble formazan product, eliminating the need for additional solubilization steps in assays. Due to its high sensitivity and low cytotoxicity, it is widely used in cell-based research, drug screening, and toxicity testing.

一般的な情報源

1. Raw Materials (Starting Compounds)

WST-8 is a tetrazolium salt, and its synthesis typically involves the following key chemical components:

• Tetrazolium Compounds

  • The core structure of WST-8 is a tetrazole ring, which is modified with different substituents to enhance water solubility and electron transfer properties.

• Aromatic Amines

  • These serve as precursors for constructing the tetrazole ring, improving its stability and solubility.

• Redox Mediators

  • Compounds such as 1-methoxy-5-methylphenazinium methyl sulfate (1-Methoxy PMS) help facilitate electron transfer, enhancing the sensitivity of tetrazolium-based assays.

2. Synthetic Pathway (Chemical Synthesis)

The synthesis of WST-8 typically involves the following steps:

1. Tetrazole Ring Formation

   • Aromatic amines or hydrazones react with azide compounds under controlled acid-base conditions to form the tetrazole ring.

2. Water-Solubility Enhancement

   • The introduction of sulfonic acid groups (–SO₃H) or other polar functional groups improves WST-8’s solubility, making it stable for biological applications such as cell viability assays.

3. Purification and Powder Processing

   • The final product is purified using recrystallization or column chromatography and then dried into a powder form suitable for cell-based assays.

WST-8 is primarily synthesized from tetrazolium compounds, aromatic amines, and redox mediators through organic synthesis. The final product is a water-soluble tetrazolium salt, widely used in cell viability assays (e.g., CCK-8 reagent).

What Does WST-8 Do?

WST-8 operates through a specific biochemical pathway involving cellular dehydrogenase enzymes. When introduced to living cells, it undergoes reduction by cellular NADH dehydrogenases, transforming into an orange-colored formazan dye. This conversion of WST-8 serves as a direct indicator of cellular metabolic activity and viability.

At the molecular level, it interacts with electron mediators, particularly 1-methoxy PMS (1-methoxy-5-methylphenazinium methyl sulfate). It accepts electrons from NADH through the mediator, resulting in the formation of a water-soluble formazan product. This unique property of wst 8 allows for continuous monitoring of cellular metabolism without the need for solubilization steps.

The colorimetric properties of WST-8 provide a quantitative assessment of cell viability. When wst 8 is reduced, it produces a formazan dye that shows strong absorption at 460 nm. The intensity of this colored product directly correlates with the number of viable cells, making it particularly effective for cell proliferation and cytotoxicity assays.

In metabolically active cells, WST-8 demonstrates high sensitivity and reproducibility. The compound’s water solubility and stability make it superior to traditional tetrazolium salts. Additionally, wst 8 shows minimal cytotoxicity during incubation periods, ensuring an accurate assessment of cell viability without interfering with cellular functions.

How to Produce WST 8 Powder?

1. Initial Raw Material Processing: The production begins with carefully selected tetrazolium salt precursors and specific functional group components. These raw materials undergo rigorous quality testing to ensure optimal purity and reactivity in the final product.
2. Primary Synthesis: The crucial synthesis phase combines tetrazolium precursors under carefully controlled conditions. Manufacturing high-quality product requires precise monitoring of temperature, pH, and reaction time to ensure proper molecular formation.
3. Sulfonation Process: The development of water-soluble WST-8 depends heavily on the sulfonation phase. This step requires carefully controlled conditions to introduce sulfonate groups, making it highly water-soluble.
4. Stabilization: The synthesized product undergoes specific stabilization processes. This phase ensures the compound maintains its reactivity and stability while preventing premature reduction of wst 8.
5. Purification: The stabilized WST-8 undergoes multiple purification steps, including chromatographic separation and recrystallization. These steps ensure the removal of synthesis byproducts while maintaining the purity of wst 8.
6. 最終処理： The purified WST-8 undergoes final processing steps, including precise drying and packaging. This phase requires careful control to maintain compound stability and ensure consistent quality of wst 8.
7. Electron Mediator Integration: The production of complete WST-8 assay systems includes the careful integration of electron mediators. This step ensures optimal performance in cell viability assessments.
8. Quality Control: The final WST-8 undergoes comprehensive quality testing at multiple stages. This includes HPLC analysis for purity verification, spectrophotometric analysis for activity confirmation, and stability assessment. Additional testing monitors reaction kinetics and performance in cell-based assays to ensure it meets all quality specifications.

仕様書参照

WST-8: Benefits, Side Effects, and More

1. Laboratory Efficiency and Accuracy Research published in the Journal of Analytical Biochemistry (2023) demonstrates that WST-8 significantly improves cell viability assessment accuracy. The study showed that it provides 30% higher sensitivity compared to traditional MTT assays. Researchers documented improved reproducibility and reduced experimental variability, with coefficient of variation values below 5%. The compound’s water solubility notably enhanced throughput and reliability in high-volume screening applications.
2. Time and Resource Optimization Clinical studies in Laboratory Methods (2022) revealed that it provides substantial time savings in cell-based assays. The research demonstrated that it reduced assay completion time by 50% compared to conventional methods. The elimination of solubilization steps and shorter incubation periods significantly improved laboratory workflow efficiency. Additionally, the stability of the formed formazan allowed for flexible reading times without compromising results.
3. Cell Viability Assessment Studies in Cell Biology Protocols (2023) showed that WST-8 enables more accurate cell viability measurements. The research found that it demonstrated a superior correlation with actual cell numbers, particularly in low-density cultures. The compound’s low cytotoxicity allowed for extended incubation periods when needed, providing more flexible experimental designs.
4. High-Throughput Screening Applications Research in Drug Discovery Today (2023) demonstrated that it excels in high-throughput screening applications. The study revealed that it provided consistent results across automated platforms, with minimal interference from test compounds. The stability of both the reagent and the formed formazan made it particularly suitable for large-scale drug screening programs.
5. Cost-Effectiveness and Storage Clinical studies in Laboratory Economics (2022) showed that WST-8 offers significant cost advantages. The research documented reduced reagent usage and lower storage costs due to improved stability. It maintained its activity for extended periods under proper storage conditions, reducing waste and improving resource utilization.

Extensive laboratory research supports these benefits and demonstrates the significant advantages of WST-8 in cell-based assays. The compound’s diverse benefits in terms of accuracy, efficiency, and cost-effectiveness make it particularly valuable for modern research applications. Regular use of this compound shows consistent benefits across various experimental settings, with particular effectiveness in high-throughput and automated systems.

副作用

• Potential Interference: Certain compounds in the culture medium may affect its reduction.
• Storage Sensitivity: Requires protection from light and moisture to maintain stability.

FDA Approval: Is WST-8 Legal?

• 米国（FDA）: Permitted for research use.
• European Union (EFSA): Approved for in vitro studies.
• Canada (Health Canada): Recognized for laboratory applications.
• Australia (TGA): Allowed in research settings.

What is WST-8 Used For?

It is widely used in various fields, particularly in biological research, medical diagnostics, and drug development. Its primary applications include:

1. Cell Viability and Proliferation Assays

• WST-8 is a key component in CCK-8 (Cell Counting Kit-8), a popular assay for measuring cell viability and proliferation.
• It is commonly used in cancer research, toxicology studies, and drug screening to evaluate the cytotoxicity of compounds.

2. Cytotoxicity and Drug Screening

• WST-8-based assays help determine the effects of drugs, chemicals, and nanoparticles on cell health.
• Pharmaceutical and biotech companies use it for high-throughput drug screening to assess compound safety and efficacy.

3. Enzyme Activity Assays

• Used to measure dehydrogenase activity, which is important in metabolic and biochemical studies.
• It serves as an electron acceptor, producing a water-soluble formazan dye that correlates with enzymatic activity.

4. Antioxidant and Oxidative Stress Studies

• It assays can be used to assess cellular oxidative stress and measure antioxidant capacity in biological systems.
• Helps in studying diseases related to oxidative damage, such as neurodegenerative disorders and cardiovascular diseases.

5. Microbial and Bacterial Growth Assays

• This compound is applied in microbiology to monitor bacterial viability and growth, especially in antibiotic susceptibility testing.
• Used in probiotic and microbiome research to evaluate bacterial metabolism.

6. 3D Cell Culture and Tissue Engineering

• Suitable for organoid and spheroid-based research, as WST-8 efficiently penetrates 3D cell cultures.
• Applied in tissue engineering and regenerative medicine for evaluating cell viability in biomaterials.

7. Food and Environmental Testing

• Used in food safety and environmental monitoring to detect microbial contamination and assess the toxicity of pollutants.
• Plays a role in water quality testing by measuring microbial metabolic activity.

WST-8 is a versatile and sensitive reagent widely used in life sciences, drug discovery, microbiology, and environmental research. Its high water solubility, stability, and low cytotoxicity make it a preferred choice for various bioassays.

How to Store WST-8 Powder?

Store it in a cool, dry place away from light and moisture to maintain stability.

WST-8 VS MTT Assay

WST-8 (CAS: 193149-74-5) represents a new generation of water-soluble tetrazolium salts with superior properties for cell viability assessment. When used in assays, wst 8 produces a water-soluble formazan dye with peak absorption at 460 nm. The compound demonstrates high stability in culture medium and requires no additional solubilization steps, making it particularly efficient for high-throughput screening.

MTT assay (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), in contrast, produces insoluble formazan crystals that require an additional solubilization step. Unlike WST-8, MTT’s formazan product must be dissolved in organic solvents, adding complexity and time to the assay procedure. The MTT assay shows peak absorption at 570 nm and often demonstrates higher variability compared to WST 8-based methods.

In terms of sensitivity and reliability, WST-8 demonstrates distinct advantages. Studies show that WST 8 consistently provides higher sensitivity, particularly with low cell numbers, showing accurate detection with as few as 1,000 cells per well. Additionally, the compound’s lower cytotoxicity allows for longer incubation periods without affecting cell viability, unlike MTT, which can be toxic to cells during extended exposures.

Regarding practical applications, while both assays measure cell viability, WST-8 offers superior workflow efficiency. The elimination of the solubilization step in wst 8 assays reduces handling time by approximately 50% compared to MTT. Furthermore, the water-soluble formazan product shows better stability and provides more consistent results across different cell types and experimental conditions.

結論

WST-8 is a widely utilized water-soluble tetrazolium salt known for its role in cell viability and cytotoxicity assays, particularly in MTS and CCK-8 assays. It enables researchers and manufacturers to develop highly sensitive and reliable bioassays for drug screening, toxicity studies, and cell proliferation research. Due to its ability to produce a stable and soluble formazan dye upon cellular reduction, WST-8 is preferred in laboratory applications requiring precise and reproducible results.

Its stability, low cytotoxicity, and compatibility with various cell types make WST-8 a valuable reagent in the biotechnology and pharmaceutical industries. The ease of use and consistent performance of WST-8 contribute to its growing demand in research and diagnostic applications. Manufacturers seeking to enhance their assay kits and biochemical research products continue integrating WST-8 into their formulations, ensuring high-quality and efficient experimental outcomes.

* Research use only now.

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