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Nucleic acid aptamer-based escherichia coli O157:H7 colloidal gold test strip, and detection method

A nucleic acid aptamer and colloidal gold test paper technology, which is applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve problems such as unclear aptamer recognition sites, unsatisfactory detection methods, and large changes in cell surfaces, etc., to achieve benefits Rapid screening, overcoming the effects of long preparation cycle and low antibody affinity

Active Publication Date: 2017-04-19
BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] Although most of the aptamer detection methods for Escherichia coli currently use the whole cell of Escherichia coli as the target to screen the aptamer as the target, the aptamer has poor recognition specificity for Escherichia coli, and the bacteria multiply rapidly. , the cell surface changes greatly, and the aptamer recognition site is not clear, which can easily lead to false positives, which cannot meet the requirements of the detection method

Method used

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  • Nucleic acid aptamer-based escherichia coli O157:H7 colloidal gold test strip, and detection method
  • Nucleic acid aptamer-based escherichia coli O157:H7 colloidal gold test strip, and detection method
  • Nucleic acid aptamer-based escherichia coli O157:H7 colloidal gold test strip, and detection method

Examples

Experimental program
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Embodiment 1

[0053] Example 1 Escherichia coli O157 based on nucleic acid aptamer: H7 colloidal gold test strip and preparation method thereof

[0054] This example provides a rapid detection system for E. coli O157:H7 colloidal gold test strips based on nucleic acid aptamers, which includes a high-affinity adapter that can specifically recognize the outer membrane protein of E.coli O157:H7 Aptamers and colloidal gold test strips with aptamers as recognition elements.

[0055] The Escherichia coli O157 based on the nucleic acid aptamer: H7 colloidal gold test strips, including sample pad 1, colloidal gold binding pad 2, nitrocellulose membrane 3 and absorption pad 4 immobilized with streptavidin, the described A detection line 5 and a quality control line 6 are arranged on the nitrocellulose membrane 3 fixed with streptavidin, and the sample pad 1, the colloidal gold binding pad 2, the nitrocellulose membrane 3 fixed with streptavidin and the absorbent pad 4 are pasted on the plastic line...

Embodiment 2

[0080] Embodiment 2 utilizes colloidal gold test strip to detect the method for Escherichia coli O157: H7

[0081] This embodiment provides a rapid detection method for Escherichia coli based on nucleic acid aptamers. The method uses E.coliO157: H7 outer membrane protein to specifically bind nucleic acid aptamers as the recognition molecules of colloidal gold test strips, and uses nano-gold As a medium, the principle of lateral flow chromatography is used to detect whether there is E. coli in the sample.

[0082] Utilize the colloidal gold test strip prepared in embodiment 1 to detect Escherichia coli O157: The working principle of H7 is as follows figure 2 shown. The specific method is as follows:

[0083] (1) Weigh 23.5g of Brain Heart Infusion Broth (BHI) medium, add 1L of sterile water preheated to 42±1°C, and dissolve for later use;

[0084] (2) Aseptic operation Take 25g or 25mL sample into a homogenization bag, add 225mL preheated brain heart infusion broth medium i...

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Abstract

The invention provides a nucleic acid aptamer-based escherichia coli O157:H7 colloidal gold test strip. The nucleic acid aptamer-based escherichia coli O157:H7 colloidal gold test strip comprises a sample pad, a colloidal gold combined pad, a nitrocellulose membrane immobilized with streptavidin, and an absorbent pad; the nitrocellulose membrane is provided with a detection line and a quality control line; the sample pad, the colloidal gold combined pad, the nitrocellulose membrane, and the absorbent pad are glued onto a plastic liner board successively; colloidal gold labeled nucleic acid aptamer is arranged on the colloidal gold combined pad; a capture probe is immobilized on the detection line; and a quality control probe is immobilized on the quality control line. The invention also provides a method used for detecting escherichia coli O157:H7 with the nucleic acid aptamer-based escherichia coli O157:H7 colloidal gold test strip. According to the nucleic acid aptamer-based escherichia coli O157:H7 colloidal gold test strip, the escherichia coli O157:H7 outer membrane protein high specificity aptamer is taken as a recognition element of test strip instead of antibodies, so that problems of immunochromatographic test strip containing antibodies, such as high cost, large batch difference, induced cross reaction, and low sensitivity, are solved, and high-efficiency rapid detection of escherichia coli is realized.

Description

technical field [0001] The invention relates to the technical field of detection of food-borne pathogenic microorganisms, in particular to a nucleic acid aptamer-based Escherichia coli O157:H7 colloidal gold test strip and a detection method. Background technique [0002] Food poisoning caused by foodborne pathogenic microorganisms is one of the important food safety issues of global concern. According to the national foodborne disease monitoring information, the number of people with food poisoning caused by foodborne pathogenic microorganisms accounts for about 40-60% of the total number of patients with foodborne diseases. Escherichia coli O157:H7 is an important indicator for evaluating food safety among food-borne pathogens. In recent years, the detection method for Escherichia coli O157:H7 has developed from the traditional tedious and time-consuming microbial culture to the rapid detection method using immunology, molecular biology and materials technology. This typ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569G01N33/558
CPCG01N33/558G01N33/56916
Inventor 栾云霞陆安祥王纪华田晓琴付海龙
Owner BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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