A method for the determination of lysozyme based on the interaction between carboxylated carbon nanoparticles and DNA

A carbon nanoparticle, carboxylated carbon technology, applied in chemiluminescence/bioluminescence, analysis by chemical reaction of materials, etc., can solve the problems of complicated operation and low sensitivity of the method, and achieve the effect of good development prospects

Active Publication Date: 2015-09-09
凯惠睿智生物科技(上海)有限公司
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  • Description
  • Claims
  • Application Information

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Problems solved by technology

These methods have their own advantages and can meet the detection requirements of lysozyme to varying degrees, but the sensitivity of the method is not high and the operation is complicated.

Method used

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  • A method for the determination of lysozyme based on the interaction between carboxylated carbon nanoparticles and DNA
  • A method for the determination of lysozyme based on the interaction between carboxylated carbon nanoparticles and DNA
  • A method for the determination of lysozyme based on the interaction between carboxylated carbon nanoparticles and DNA

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preparation example Construction

[0023] The present invention is achieved through the following measures: a preparation method of a chemiluminescent sensor for detecting lysozyme, comprising the following steps:

[0024] (1) Preparation of ABEI-apt:

[0025] For the preparation of the ABEI-apt, preferably, the ABEI-apt is prepared by reacting an aminated DNA aptamer (apt) with ABEI using glutaraldehyde as a cross-linking agent. The specific process is as follows:

[0026] (a) 20 μL 10 -6 ~10 -4 Add the aminated DNA aptamer of M to 5% glutaraldehyde in phosphate buffer solution, and keep stirring for 2 hours;

[0027] (b) Add 1-10 mg of ABEI solution and stir overnight at room temperature to obtain the obtained product.

[0028] (2) Preparation of carboxylated carbon nanoparticles:

[0029] The preparation of described carboxylated carbon nanoparticles, preferably, the specific process is as follows:

[0030] (a) Preparation of carbon nanoparticles:

[0031] Preferably, 0.1-1.0 g of chitosan is dissolve...

Embodiment

[0048] Example: A method for the determination of lysozyme based on the interaction between carboxylated carbon nanoparticles and DNA.

[0049] 1. Experimental part

[0050] 1.1 Instruments and reagents

[0051] IFFM-E Flow Injection Chemiluminescence Analyzer (Xi'an Ruimai Analytical Instrument Co., Ltd.); Lmax II / II384 Chemiluminescence Analyzer (MD Company, USA); THZ-82A Air Bath Constant Temperature Oscillator (Quantan Medical Instrument Factory) .

[0052] Chemiluminescence reagent (ABEI) was purchased from Aladdin Reagent Company; aminated magnetic beads (preferred, 2-3 μm in diameter) were purchased from Tianjin Beisile Chromatography Technology Development Center; cocaine and codeine were purchased from China Institute for the Control of Pharmaceutical and Biological Products ; Lysozyme (Lysozyme, Lyz), thrombin, adenosine triphosphate (ATP), bovine serum albumin (BSA) and immunoglobulin G were all purchased from Sigma Company; N-hydroxysuccinimide (NHS), 1-ethyl - ...

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Abstract

The invention provides a lysozyme content detection method which is used for detecting lysozyme content in cells and biological samples and belongs to the technical field of a chemiluminiscence aptasensor. The lysozyme content detection method comprises the following steps of: modifying amination magnetic beads by using carboxylation carbon nanoparticles so as to obtain carbon nanoparticle modified magnetic beads; and modifying lysozyme aptamer by using a chemiluminiscence agent ABEI (3-aminophthalic acid cyclic hydrazide) so as to prepare a marked aptamer; combining the marked aptamer and the lysozyme in the presence of lysozyme, mixing a solution, which is subjected to the action of the lysozyme-marked aptamer, with the carbon nanoparticle modified magnetic beads, wherein liquid supernatant generates chemiluminiscence in the presence of H2O2 and Co<2+> after magnetic separation, thus realizing the detection of the lysozyme. The lysozyme content detection method is simple and low in cost.

Description

technical field [0001] The invention belongs to the field of methods for measuring lysozyme, in particular to a method for measuring lysozyme based on the interaction between carboxylated carbon nanoparticles and DNA. Background technique [0002] Lysozyme (Lysozyme, Lyz) is also known as cytoplasmic enzyme (Muramidase) or N-acetylmuramide polysaccharide hydrolase (N-axetylmuramide glycanhydrolase). Because lysozyme can hydrolyze the cell wall of Gram-positive bacteria and block the murein polysaccharide to destroy the cell wall, it has a bactericidal effect. Lysozyme is essential in the immune system of animals and humans, and its content is different in different tissues, and the content is different when the disease occurs and under normal conditions. Reduced lysozyme content in tissues can easily cause many diseases. Therefore, the detection of lysozyme content is of great significance. [0003] The current traditional method for measuring lysozyme has low sensitivity...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/76
Inventor 混旭刘芳
Owner 凯惠睿智生物科技(上海)有限公司
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