Method for detecting organophosphate flame retardant based on acetylcholin esterase-gold nanoclusters
An acetylcholinesterase and organophosphate technology, which is applied in the field of nanomaterial fluorescence detection, can solve the problems of insensitive detection, prone to interference, complex signal transmission process, etc., so as to reduce detection insensitivity and interference, and simplify the detection signal transmission process. , the effect of reducing costs
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Embodiment 1
[0031] Preparation of gold nanocluster fluorescent material: mix 10mmol / L chloroauric acid 1mL with 0.2U / ml acetylcholinesterase 100μL, preheat at 80°C for 5 minutes, shake vigorously, add 1.0mol / L tris-HCl buffer Solution (pH=7.4) and 1mol / L sodium hydroxide solution regulate pH to 12.3, and make mixed solution final reaction volume reach 2mL, this moment, acetylcholinesterase and chloroauric acid concentration ratio (mU:mmol / L) are 4: 1. React at 80°C for 18 hours, filter large particles of gold with a 0.45 μm filter head, and obtain a light white solution after ultrafiltration with a 10kDa ultrafiltration tube for 13 minutes. There is obvious blue fluorescence under the irradiation of a 365nm ultraviolet light source, and acetylcholine ester is obtained. Enzyme-gold nanocluster solution, the solution concentration is about 5mmol / L. The obtained acetylcholinesterase-gold nanocluster solution maximum excitation wavelength and emission wavelength are respectively 315nm and 39...
Embodiment 2
[0033] Establishment of a method for detecting organophosphate flame retardants based on acetylcholinesterase-gold nanoclusters: 100 μL of the acetylcholinesterase-gold nanocluster solution obtained in Example 1 was mixed with tris(2-chloropropyl) phosphate ( TClPP) mixed, then add 1.0mol / L tris-HCl buffer solution (pH=7.4) to obtain 2mL mixed solution, the final system concentration of tris (2-chloropropyl) phosphate (TClPP) is 0, 25, 50, 100, 300, 500, 700, 900, 1000 ,1500,2000,2500,3000 and 4000ng / L. Incubate at 40°C for 50 minutes, measure the fluorescence intensity of acetylcholinesterase-gold nanoclusters containing different concentrations of tris(2-chloropropyl)phosphate (TClPP) at an excitation wavelength of 315nm, and calculate it as F, at a concentration of 0ng / L The fluorescence intensity was measured as F0 (see figure 2 ), calculate the value of fluorescence intensity rate of change (F-F0) / F0, establish the standard curve of (F-F0) / F0 and tris (2-chloropropyl) pho...
Embodiment 3
[0035] Optimization of incubation conditions for detection of organophosphate flame retardants based on acetylcholinesterase-gold nanoclusters: Mix 100 μL of the acetylcholinesterase-gold nanocluster solution obtained in Example 1 with tris(2-chloropropyl)phosphate (TClPP) , and then add 1.0mol / L tris-HCl buffer solution (adjust the pH range of the mixed solution to 6.0-9.0) to obtain a 2mL mixed solution, and the final system concentration of tris(2-chloropropyl)phosphate (TClPP) is 1000ng / L. Incubate at 40°C for 50 minutes, measure the fluorescence intensity of acetylcholinesterase-gold nanoclusters containing tris(2-chloropropyl)phosphate (TClPP) at an excitation wavelength of 315nm, and calculate it as F, and measure it at the concentration of 0ng / L The fluorescence intensity of acetylcholinesterase-gold nanoclusters is calculated as F0, and the value of the rate of change of fluorescence intensity (F-F0) / F0 is calculated, such as Figure 4 As shown, after the incubation, ...
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