An analysis method for detecting lipase activity based on gold nanocluster fluorescence enhancement

A technology of gold nano-clusters and analysis method, which is applied in the field of analysis of lipase activity based on fluorescence-enhanced detection of gold nano-clusters, and achieves the effects of simple and fast operation, low cost and simple method

Inactive Publication Date: 2018-11-30
NANJING UNIV OF TECH
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  • Description
  • Claims
  • Application Information

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Problems solved by technology

Currently, the detection of Cu based on AuNCs 2+ , Hg 2+ , trypsin, melamine and phosphate-containing metabolites have been reported, but there is no report on the detec

Method used

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  • An analysis method for detecting lipase activity based on gold nanocluster fluorescence enhancement
  • An analysis method for detecting lipase activity based on gold nanocluster fluorescence enhancement

Examples

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Effect test

Embodiment 1

[0019] Example 1 Mechanism Verification

[0020] Add 800 μL of gold nanoclusters and 100 μL of PB buffer solution to 2 mL centrifuge tubes a, b, c, and d respectively, then add 100 μL of methyl thioglycolate to tube b, and add 100 μL of Methyl thioglycolate and 5 μg lipase, add 100 μL of methyl thioglycolate and 30 μg lipase to tube d, and make the other volume up to 1000 μL with ultrapure water, bathe in 35°C water for 8 minutes, and detect the fluorescence intensity (such as figure 1 shown).

[0021] figure 1 It is a mechanism verification diagram. It can be seen from curve a and curve b that the addition of the substrate methyl thioglycolate does not change the fluorescence intensity of gold nanoclusters. When 5 μg / mL lipase is added, the curve c, The fluorescence of gold nanoclusters is enhanced, and when 30 μg / mL lipase is added, the fluorescence intensity is further enhanced, indicating that the higher the concentration of lipase, the more the fluorescence intensity of...

Embodiment 2

[0022] The drafting of the standard curve of embodiment 2 lipase activity detection

[0023] Add 800 μL of gold nanoclusters, 100 μL of PB buffer solution, and 100 μL of methyl thioglycolate into a 2 mL centrifuge tube, bathe in water at 35°C for 8 min, and then add different concentrations of lipase, the concentration of lipase is as follows: 0 μmol / L , 5 μmol / L, 10 μmol / L, 15 μmol / L, 20 μmol / L, 25 μmol / L, 30 μmol / L, dilute to 1000 μL with ultrapure water, measure its fluorescence intensity after 35 ° C water bath for 8 min; the concentration of lipase is On the abscissa, the increased value of the fluorescence intensity is plotted on the ordinate, and the standard curve of lipase can be drawn (such as figure 2 shown).

[0024] figure 2 It shows that with the increase of the lipase concentration, the fluorescence intensity of the solution is gradually enhanced and the lipase concentration has a good linear relationship between 5-30μmol / L, and the detection limit is 0.37μg...

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Abstract

An analysis method for detecting lipase activity based on gold nanocluster fluorescence enhancement is disclosed. According to a detection mechanism, thioglycollic acid generated by a lipase hydrolysis substrate that is methyl thioglycolate can stabilize gold nanocluster and can make up surface defects of the gold nanocluster, thus enhancing fluorescence. Lipase activity is detected through fluorescence intensity increase of the god nanocluster at 570 nm. The method is simple, rapid and sensitive, and the lowest limit of detection is 0.37 [mu]g/mL. Compared with the prior art, the method is advantageous in that the method is simple and rapid to operate and low in cost, and does not require special instruments or devices.

Description

technical field [0001] The invention belongs to the field of nano-biosensors, and in particular relates to an analysis method for detecting lipase activity based on gold nano-cluster fluorescence enhancement. Background technique [0002] Enzymes are catalytically active proteins that are produced by living organisms through natural evolution. Enzymes can increase the rate of chemical reactions in different organic or inorganic substrates, and they all have specificity. Due to this specificity, enzymes are widely used in analysis and detection as recognition elements and signal amplification elements. Enzyme activity depends on substrate concentration, temperature, pH, and ionic strength. Enzyme analysis initially uses colorimetry and nuclear magnetic resonance (NMR) methods, but these enzyme activity detection methods are often affected by low sensitivity and long reaction time. Nowadays, the development of inorganic nanoscience has provided a variety of methods, and these...

Claims

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Application Information

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IPC IPC(8): G01N21/64
CPCG01N21/6428
Inventor 田丹碧朱杰张凌华
Owner NANJING UNIV OF TECH
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