Enzyme immunosensor for detecting Shigella species and its preparation method and application

A Shigella and enzyme immunization technology, applied in instruments, measuring devices, scientific instruments, etc., can solve the problems of complex detection process, long detection time, human and environmental hazards, etc., and achieve simplified preparation steps, short response time, highly specific effect

Inactive Publication Date: 2009-09-16
ZHEJIANG GONGSHANG UNIVERSITY
View PDF0 Cites 11 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The national standard method is convenient to obtain materials, but the sample pretreatment is complicated, the operation process is cumbersome, the detection cycle is long, and the sensitivity is low; ELISA requires special equipment, detection steps are cumbersome, high cost, long detection time, and prone to false p

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Enzyme immunosensor for detecting Shigella species and its preparation method and application
  • Enzyme immunosensor for detecting Shigella species and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0040] Example 1: Preparation of enzyme immunosensor

[0041] Ultrasonic dispersion of 2.0 mg of multi-walled carbon nanotubes in 10 mL of chitosan sol with a mass volume percentage of 1% to obtain a uniform multi-walled carbon nanotube-chitosan composite;

[0042] Fully mix the multi-walled carbon nanotube-chitosan complex and the horseradish peroxidase-labeled anti-Shigella flexneri antibody solution in equal volumes; the mixture is placed at 4°C for 12 hours;

[0043] Take 3μL of the above-mentioned mixed solution and apply dropwise to the surface of the working electrode of the disposable screen-printed electrode, and place it at room temperature for 3 to 5 hours to dry and form a film;

[0044] Soak the coated immunoelectrode in PBS phosphate buffer solution containing 0.2% BSA (bovine serum albumin), place it at 4°C for 1 hour, and rinse with double distilled water to remove the chitosan that is not firmly bound to the electrode. Enzyme immunosensor prepared by antibody memb...

Example Embodiment

[0046] Example 2: Method for detecting Shigella

[0047] The enzyme immunosensor described in Example 1 is connected to the electrochemical workstation (CHI 1030A) through a wire, and the computer is connected to the electrochemical workstation.

[0048] Put the enzyme immunosensor into the electrolytic cell, and the detection bottom liquid of the electrolytic cell contains 0.6mmol / L H 2 O 2 And 1.0mmol / L thionine acetate buffer, pH value is 7.0. Set the CV measurement parameters of the electrochemical workstation: the potential scan range -0.1~-0.6V, the scan rate is 0.1V / s, apply a voltage to the working electrode of the enzyme immunosensor, and measure the current value Ip1 between the auxiliary electrode and the working electrode It is 1.946μA.

[0049] Anal swabs from patients with vomiting and diarrhea were collected aseptically, cultured and separated, centrifuged at 8000r / min for 5 minutes, and the lower layer of bacterial sludge was taken out and diluted with pH7.4 PBS to...

Example Embodiment

[0051] Example 3

[0052] Using the immunosensor described in Example 1 and the Shigella detection method in Example 2, different concentrations of Shigella were measured to prepare a standard current-concentration curve, specifically using the Shigella flexneri standard Sample preparation 10 10 cfu / mL PBS solution, and gradually diluted to 10 4 cfu / mL. Make a standard curve, such as figure 2 Shown. The lower limit of detection: 10 4 cfu / mL; linear interval: 10 4 cfu / mL~10 10 cfu / mL; linear correlation coefficient: R 2 = 99.54%.

[0053] Anal swabs from patients with vomiting and diarrhea were taken aseptically. The sample processing and detection methods were the same as those in Example 2. Finally, ΔI = 0.9312μA was obtained. Compare the standard current-concentration curve to calculate the Shiga Fuchs contained in the tested sample. The bacteria concentration is 3×10 7 cfu / mL.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses a sensitive, rapid, accurate, high-specificity and economical enzyme immunosensor for detecting Shigella species, comprising screen printing electrodes, a sensitive membrane containing enzyme marked Shigella species antibody- carbon nano-tube- shitosan mixture is coated on the working electrode of the screen printing electrodes. In the invention, the chemical amplification function of the multi-wall carbon nano-tube and enzyme is combined with the specificity of the immunosensor, theenzyme immunosensor has the specificity of immunoreactivity and the sensitivity of the electrochemistry analysis and can accurately detect the lower-content substance. The invention also discloses a preparation method and application of the enzyme immunosensor, the Shigella species can be directly detected, with the sensitive, rapid, high-specificity advantages, enzyme immunosensor is cheap and suitable for detecting the Shigella species in the foundation and in situ.

Description

technical field [0001] The invention relates to the technical field of rapid detection of food-borne pathogenic bacteria, in particular to an enzyme immunosensor for detecting Shigella based on an antigen-antibody immune reaction and a preparation method and application thereof. Background technique [0002] Among the hundreds of millions of food-borne disease patients in the world every year, 70% are caused by eating food and drinking water contaminated by various pathogenic microorganisms. Shigella (Shigella) is one of the main pathogens that cause acute infectious diarrhea. As few as 10 bacteria can cause infection, and the infected human body will have symptoms such as nausea, vomiting, abdominal pain, diarrhea (severe pus and blood in the stool), and even endanger the human body. life. Shigella infection kills more than one million people every year, mostly children in developing countries. [0003] The rapid detection of Shigella is one of the most critical measures ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): G01N27/333G01N33/569G01N27/48
Inventor 赵广英詹学佳
Owner ZHEJIANG GONGSHANG UNIVERSITY
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap