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Home / Assay Kits / General Enzyme Assays

MicroMolar Universal Kinase Assay Kit

BHE16300163

This product includes 600 ul of 10 x assay buffer, 35 ul of 100 x ATP, 35 ul of 100 x MUK Reagent A, 35 ul of 100 x MUK Reagent B, 30 ul of 1 mM ADP and 350 ul of 10 x fluorescence dye for 100 assays in a 384-well assay format.

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Product Manual Datasheet
Protocols Description Assay Protocol
1. Reagent preparation:
(1) 1 x MUK Buffer: dilute the 10 x MUK Buffer 10-fold with water. The 1x MUK Buffer is composed of 50 mM Tris-HCl, pH 8.0, 50 mM NaCl, 3 mM MgCl2, 0.5 mM DTT, 0.2 mM EDTA, 0.003% Brij-35.
(2) 10 x MUK Reagent A: dilute the 100 x MUK Reagent A 10-fold with water
(3) 10 x MUK Reagent B: dilute the 100 x MUK Reagent B 10-fold with 1x MUK Buffer
(4) 1 x fluorescence dye: dilute the 10 x fluorescence dye 10-fold with water.
 
2. Kinase assay
The kinase reaction volume is 30 µl.
(1) In a standard black 384-well plate, mix 12 µl of water, 3 µl of 10 x reaction buffer, 3 µl of 10 x kinase substrate, 3 µl of 10 x ATP, 3 µl of freshly prepared 10 x kinase. Incubate the reaction for 2 min.
(2) Add 3 µl of 10 x MUK Reagent A, 3 µl of 10 x MUK Reagent B.
(3) Incubate the reaction mixture at 37°C for 60 min.
(4) Add 30 µl of the 1 x fluorescence dye into the 30 µl of the reaction mixture.
(5) Measure the fluorescence intensity at 535 nm with excitation at 485 nm.
 
Enzyme inhibition
For the enzyme inhibition experiment, typically 50 x stock solutions with a 2-fold serial dilution in water or DMSO are prepared. In the wells of the 384-well plate, 0.6 µl of the 50 x inhibitor is mixed with a premix composed of the buffer and enzyme for 5 min. Then the substrate is added. After incubation for 2 min, 3 µl of 10 x MUK Reagent A and 3 µl of 10 x MUK Reagent B are added. The total assay volume is 30 µl. At the end of the reaction, the dye is added and the fluorescence intensity is measured.
Combination SKU Supplier No. Size Your price Quantity
BHE16300163-1 MUK100K 100 assays $392.60
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BHE16300163-2 MUK1000K 1000 assays $2209.36
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