Black phosphorus mediated copper ion aggregation electrochemical signal conversion detection method for activity of phosphatase
A technology for phosphatase activity and signal conversion, applied in the field of phosphatase activity detection, can solve the problems of low sensitivity, time-consuming, laborious, etc., and achieve the effect of high sensitivity and simple operation
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Embodiment 1
[0025] Preparation of glassy carbon electrode (GCE) drop-coated with black phosphorus nanomaterials: put the black phosphorus nanosheets in an ultrasonic instrument for 2 h, then apply a voltage of 10 V, electrolyze the solution of black phosphorus nanosheets for 30 min, and then place them in an ultrasonic Ultrasonic in the instrument for 2 h, the purpose is to peel off the black phosphorus nanosheets. Then, the black phosphorus solution (6 μg) was dropped on the surface of the GCE and placed in a dry box to obtain a glassy carbon electrode drop-coated with black phosphorus nanomaterials.
Embodiment 2
[0027] (1) Preparation of reaction solutions containing different types of enzymes:
[0028] 10 μL 2 mM PPi and 50 μL PPase (1.5 μL 1 mU / μL PPase and 44.5 μL water), 86 μL HEPESbuffer, react in a water bath at 25 °C for 55 min.
[0029] Keeping other conditions unchanged, 50 μL PPase in the above process was changed to 50 μL (11.55 μL 1299mU / mL GOx and 38.45 μL water) glucose oxidase (GOx) respectively, so that its final concentration was 100 mU / mL and 50 μL ( 3.75 μL 4 mU / μL CpG MTase, 15 μL 10×NEBuffer, 0.75 μL 200×S-adenosylmethionine, 30.5 μL water) CpG methyltransferase (CpG Methyltransferase), so that its final concentration is 100 mU / mL. The measured data correspond to figure 2 GOx and CpG MTase histograms in .
[0030] Keep other conditions unchanged, change 50 μL PPase to 1.5 μL 1mU / mL PPase, 11.55 μL 1299 mU / mL GOx, 3.75 μL 4 mU / μL CpG MTase, 15 μL 10×NEBuffer, 0.75 μL 200×S -adenosylmethionine, a mixture of 17.45 μL of water, so that the concentrations of PPase,...
Embodiment 3
[0036] (1) 10 μL 2 mM PPi and 50 μL PPase (enzyme solution with different concentrations), then add 86 μL HEPES buffer (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid), at 25 ℃ React in a water bath for 55 min.
[0037] (2) Then add 8 μL of 2 mM CuSO to the above solution 4 , 25 ℃ water bath reaction for 10 min. 30 μL of the reaction solution was dropped on the surface of the glassy carbon electrode coated with black phosphorus nanomaterial obtained in Example 1, and reacted for 5 h. The reaction solution was washed with deionized water and blown dry with gentle nitrogen gas.
[0038] (3) Place the above electrode in pH7.2 HEPES buffer (containing 50 mM KNO 3 ) in the solution of square wave voltammetry, the potential window is selected as -0.3 V-0.2 V, the potential jump is 4 mV, the amplitude is 25 mV, the frequency is 25 Hz, and the characteristic peak appears around 56 mV.
[0039] The measured SWAV curve changes with different PPase concentrations, such as imag...
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