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Kinase-Lumi™ Plus Luminescent Kinase Assay Kit

Kinase-Lumi™ Plus Luminescent Kinase Assay Kit

€ 325,50

Beyotime’s Kinase-Lumi™ Plus Luminescent Kinase Assay Kit provides a chemiluminescent method for measuring kinase activity by quantifying the amount of ATP remaining in the solution after the kinase reaction. This kit can also be used to detect other enzymes that consume ATP, such as ATPase.

Package: 1000T

SKU: S0155s
Category: , , ,

Description

Beyotime’s Kinase-Lumi™ Plus Luminescent Kinase Assay Kit provides a chemiluminescent method for measuring kinase activity by quantifying the amount of ATP remaining in the solution after the kinase reaction. This kit can also be used to detect other enzymes that consume ATP, such as ATPase.

This kit can detect ATP up to 50μM with good linearity in the range of 0-50μM. For lower concentrations of ATP, we recommend using Beyotime’s Kinase-Lumi™ Luminescent Kinase Assay Kit with the maximum detection limit of 10μM. For higher concentrations of ATP, we recommend using Beyotime’s Kinase-Lumi™ Max Luminescent Kinase Assay Kit with the maximum detection limit of 200μM or diluting the sample appropriately before the assay.

This kit is suitable for a wide range of ATP-consuming kinases, including protein kinases with protein or peptide as substrate and non-protein kinases with sugar, lipid and alcohol as substrate. This kit is also suitable for the detection of ATP-consuming ATPases.

Please refer to Figure 1 for the reaction principle of this product. The Kinase is an enzyme that catalyzes the transfer of a phosphate group from ATP to the hydroxyl group of substrate, leading to a decreased ATP content in the kinase reaction. ATP remaining in the kinase reaction can be quantified by the chemiluminescent method catalyzed by luciferase in the presence of luciferin. The luminescent signal generated in the reaction is correlated with the amount of remaining ATP, while inversely correlated with the amount of kinase activity.


Figure 1. Reaction principle of Beyotime’s Kinase-Lumi™ Plus Luminescent Kinase Assay Kit (S0155).

The kit is easy to use. The assay can be completed in one well by simply adding an equal volume of assay reagent into the sample, making it an ideal choice for both small-scale analysis and automated high-throughput screening.

The luminescent signal produced by the assay using this kit is very stable due to the use of a thermo-stable firefly luciferase and the optimized reaction conditions. The luminescence can be produced within 10 minutes after the addition of the assay reagent, and the luminescent signal is stable with no more than 10% decrease within 30 minutes.

For 96-well plates, we recommend using 50μl of kinase reaction and 50μl of assay reagent, by which S0155S are sufficient for 200 assays, respectively. In 384-well plates with10μl of kinase reaction and 10μl of assay reagent, S0155S are sufficient for 1000 assays, respectively. Other volumes can also be used, but the ratio of kinase reaction to assay reagent must be 1:1.

Packing List：
Item Component Quantity
S0155S-1 Kinase-Lumi™ Plus Luminescent Kinase Activity Assay Reagent 10ml
S0155S-2 0.5mM ATP 0.2ml
Manual – 1 copy

 

Storage Conditions：

Store at -20℃ for up to 6 months or -80℃ for up to 1 year. The Assay Reagent should be protected from light.

Precautions：

The Assay Reagent contains luciferase and should be avoided from repeated freeze-thaws. It is better to aliquot the Assay Reagent at the first use. The Assay Reagent can tolerate repeated freeze-thaws up to 3 cycles. Small amount of precipitates in the Assay Reagent  if there are any should be dissolved at room temperature before use, or removed by centrifugation before use if the precipitates could not be dissolved completely.

The kinase or ATPase reaction should be performed at room temperature (approximately 25℃) and the sample needs to be equilibrated to room temperature prior to use.

The ATP provided in the kit is only sufficient for preparing the standard curve. ATP required for kinase reactions should be ordered separately by investigatiors.

Opaque 96-well plates or 384-well plates should be used for the assay.

This product is for R&D only. Not for drug, household, or other uses.

For your safety and health, please wear a lab coat and disposable gloves during the operation.

Additional information

Weight 1 kg
Dimensions 20 × 20 × 20 cm

Instruction

Instructions for Use：

1. Reagent preparation
a. The ATP assay can be performed immediately after the kinase reaction is completed. The kinase reaction can also be frozen at -80℃ for future analysis of ATP content, but should be equilibrated to room temperature (about 25℃) and mixed well before being assayed.
b. The  Luminescent Kinase Activity Assay Reagent should be equilibrated to room temperature and mixed well before use.
2. Kinase activity assay
The following protocol is provided for assays in 96-well plates. For assays in 384-well plates, the amount of solutions should be adjusted accordingly.
a. Kinase reaction: In 50μl of kinase reaction mixture, add appropriate amounts of reaction buffer, kinase substrate, ATP and kinase, and incubate for an appropriate period of time. The optimum reaction time is the time required for consuming 20-90% (30-70% best) of ATP added in the reaction. The reaction conditions for a particular kinase need to be optimized by investigators based on  relevant literature.
b. Preparation of ATP standard: Dilute the ATP standard (0.5mM) with the kinase reaction buffer into 0, 0.3, 0.6, 1.2, 2.5, 5, 10, 20, 40, and 50 μM, respectively. Add 50μl of each dilution to designated standard wells.
c. Add 50μl of Kinase-Lumi™ Plus Luminescence Kinase Assay Reagent per well, and mix well by pipetting.
d. Incubate at room temperature (~25℃) for 10 minutes, and then measure the chemiluminescence using a microplate reader. The detection time for each well is generally 0.25-1s or longer, depending on the sensitivity of the instrument.
e. Calculate the amount of ATP remaining in reactions from the standard curve, and subsequently the kinase activity can be calculated according to the definition of enzyme activity. There is a good linearity between luminescence and ATP content in the range of 0-50μM (Figure 2).

Figure 2. ATP standard curve obtained using Beyotime’s Kinase-Lumi™ Plus Luminescent Kinase Assay Kit (S0155). This figure is for reference only, which may vary due to different experimental conditions.
Notes:
Optimization of kinase reaction conditions
a. ATP concentration: When screening kinase inhibitors, it is recommended to use low concentrations of ATP (10μM or less) for screening of ATP-competitive inhibitors and use higher concentrations of ATP for screening of ATP non-competitive inhibitors. Once the ATP concentration is changed, the enzyme and substrate concentrations as well as the reaction temperature and incubation time should be optimized accordingly.
b. Substrate concentration: It is usually advisable to set a concentration gradient of the substrate and have the substrate in excess to ensure a maximum luminescence difference between the kinase wells and the negative control wells.
c. Dosage of kinase: The optimum amount of kinase can also be determined by setting a dose gradient of kinase. Adjust the kinase dosage and reaction time to ensure 30-70% of ATP  in the reaction is consumed.
FAQ:
1. What is the difference between a Luminometer and a Fluorescence Spectrophotometer?
Luminometer detects samples emitting light without excitation. In other words, the luminometer is an instrument that detects chemiluminescence. Fluorescence Spectrophotometer detects samples that need a specific wavelength of excitation light to produce detectable fluorescence by a fluorescence spectrophotometer. Some fluorescence spectrophotometers also possess the function of luminometer. Please make sure the fluorescence spectrophotometer you are using can be used for chemiluminescence detection.
2. Can the instruments used for fluoresceinase reporter gene detection be used for this assay?
Yes. The working mechanism of the fluoresceinase reporter gene detection is the same as that of this assay.

Product files

MSDS

https://scandicbiotech.com/wp-content/uploads/2026/04/S0155.pdf

References

Related papers：

1. Bozhi Yang, Shudong Zhou, Lijun Ou, Feng Liu, Liying Yang, Jingyuan Zheng, Wenchao Chen, Zhuqing Zhang, Sha Yang, Yanqing Ma, Xuexiao Zou
.
Mol Genet Genomics. 2020 Mar;295(2):343-356. doi: 10.1007/s00438-019-01626-z. (IF 2.797)
2. Hao Pan, Ting Song, Ziqian Wang, Yafei Guo, Hong Zhang, Tong Ji, Keke Cao, Zhichao Zhang
.
J Biochem. 2021 Dec 4;170(4):539-545. doi: 10.1093/jb/mvab073. (IF 2.476)
3. Zenan Zhou, Han Yang, Xinxin Liang, Tao Zhou, Tao Zhang, Yang Yang, Jiadong Wang, Weibin Wang
.
Cell Rep. 2023 Aug 29;42(8):112907. doi: 10.1016/j.celrep.2023.112907.
4. Shifeng Liu, Chengcheng Cai, Luopin Li, He Wen, Jie Liu, Liqin Li, Qiang Wang, Xiyao Wang
.
Hortic Res. 2023 Nov 8;10(12):uhad228. doi: 10.1093/hr/uhad228.

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