Product Description
Cell Counting Kit-8 (CCK-8) 允许进行灵敏的比色测定，用于确定细胞增殖和细胞毒性测定中的细胞活力。 Dojindo 的高水溶性四唑盐 WST-8 被细胞中的脱氢酶活性还原，产生可溶于组织培养基的黄色甲臜染料。 由细胞中脱氢酶的活性产生的甲臜染料的量与活细胞的数量成正比。 CCK-8的检测灵敏度高于MTT、XTT、MTS或WST-1等其他四唑盐。
Cell viability detection mechanism with CCK-8
ProcedureCell Counting Kit-8 requires 3 simple steps.
The handling time of Cell Counting Kit-8 is the shortest among it’s competitors. Only 15 minutes of handling time for Cell Counting Kit-8 where as longer handling time is required for both MTS and MTT assay.
StabilityCell Counting Kit-8 is a ready-to-use solution. It is also stabe at 4ºC for 1 year. Your assay can be done anytime without thaw and freeze.
SensitivityCCK-8 (WST-8) is the highest sensitive dye for the cell based assay.
Comparison of the sensitivity using tetrazolium dye
Cytotoxicity of ReagentsOnly in CCK-8, continious culture is possible without killing cells.
Observation of Cytotoxicity after incubation with each reagent.HeLa cells were incubated with WST-8 (CCK-8), WST-1, or MTS at 24 hours from addition of those reagents.
Example Data using Cell Counting Kit-8
Data for Cell Proliferation and Cytotoxicity Assay
Absorption property of WST-8The microplate reader with a 450-490 nm filter is applicable for the measurement.
Absorption spectrum of WST-8 formazan dye
How many cells should there be in a well?
For adhesive cells, at least 1,000 cells are necessary per well (100 μl medium) when using a standard 96-well plate. For leukocytes, at least 2,500 cells are necessary per well (100 μl medium) because of low sensitivity. The recommended maximum number of cells per well for the 96-well plate is 25,000. If 24-well or 6-well plates are used for this assay, please calculate the number of cells per well accordingly, and adjust the volume of the CCK-8 solution in each well to 10% of the total volume.
Can this kit be used with a 384-well plate?
Yes, you can use this kit for a 384-well plate. Add CCK-8 solution with 1/10 volume of the culture medium in a well. If the volume of CCK-8 to be added is too small, dilute CCK-8 in half with the medium and add 1/5 of the volume of culture medium in a well.
Can this kit be used with a 24-well plate?
Yes. Add CCK-8 solution with 1/10 volume of the culture medium in a well.
Does CCK-8 stain viable cells?
No, it does not stain viable cells because a water-soluble tetrazolium salt (WST-8) is used for the CCK-8 solution. The electron mediator, 1-Methoxy PMS, receives electrons from viable cells and transfers the electrons to WST-8 in the culture medium. Since its formazan dye is also highly water soluble, CCK-8 cannot be utilized for cell staining.
Does phenol red affect the assay?
No. The absorption value of phenol red in culture medium can be removed by subtracting the absorption value of a blank solution from the absorption value of each well. Therefore, culture medium containing phenol red can be used for the CCK-8 assay.
Is there a correlation between CCK-8 and the Thymidine incorporation assay?
Yes. However, please note that since CCK-8 uses a different assay mechanism from that of the Thymidine assay, the results may differ. Comparison data are shown in the technical manual, which is available at www.dojindo.com/tm.
Is CCK-8 toxic to cells?
CCK-8 solution itself is cytotoxic because of the high concentration of 1-Methoxy PMS. CCK-8 in the culture medium, however, is not cytotoxic because of the ten times dilution with culture medium. Therefore, long incubation, such as overnight and several days, is possible. The same cell culture can be used for other cell proliferation assays, such as the crystal violet assay, neutral red assay, or DNA fluorometric assays after the CCK-8 assay is completed. Since each cell has a different tolerance to CCK-8, incubate cells with CCK-8 solution and check the cell viability if a longer incubation time is necessary.
How stable is CCK-8?
CCK-8 在室温下可稳定超过 6 个月，在 0-5 oC 下可稳定一年，并具有避光保护。 如需更长的储存时间，我们建议将溶液储存在 -20 oC ； CCK-8 在 -20oC 下可稳定超过 2 年。
我没有 450 nm 滤光片。 我还可以使用哪些其他过滤器？
您可以使用吸光度在 450 nm 和 490 nm 之间的滤光片。