A composite film sensor and method for detecting vibrio parahaemolyticus

A hemolytic vibrio and composite membrane technology, applied in the direction of material excitation analysis, Raman scattering, etc., can solve the problems of high substrate requirements, unfavorable rapid detection, cumbersome operation, etc., achieve low preparation cost, shorten detection time, The effect of broad application prospects

Active Publication Date: 2020-12-29
JIANGNAN UNIV
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Problems solved by technology

[0004] CN105352933A provides a method for detecting Vibrio parahaemolyticus in food based on aptamer recognition surface-enhanced Raman spectroscopy. The method prepares a silver-coated gold core-shell substrate and uses a sandwich method for Raman spectroscopy to detect pathogenic bacteria , but because the system is a solution, the detection is limited by many conditions, and centrifugation and other steps are required, which is not conducive to rapid detection.
[0005] CN106093005A provides a latent fingerprint high-definition identification method based on lysozyme Raman spectrum imaging. The method collects fingerprints on the substrate and uses the sandwich method to perform Raman spectroscopy for fingerprint imaging. , Incubated under humidity conditions, so there are certain limitations

Method used

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  • A composite film sensor and method for detecting vibrio parahaemolyticus
  • A composite film sensor and method for detecting vibrio parahaemolyticus
  • A composite film sensor and method for detecting vibrio parahaemolyticus

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Embodiment 1

[0029] Embodiment 1: the preparation of composite film sensor

[0030] (1) Preparation of nano gold solution:

[0031] Take 0.5ml of 1% chloroauric acid aqueous solution, add 49ml of deionized water and boil for 5-10 minutes (120 rotation speed), then quickly add 0.5ml of 5% sodium citrate, and wait for the solution to react to produce wine red to obtain the gold nanometer solution. Take 1ml of prepared nano-gold solution, centrifuge at 12000RPM for 20min, remove 750μL of supernatant, resuspend to obtain concentrated nano-gold solution, and store in refrigerator at 4°C.

[0032] (2) Preparation of APT-AU-PDMS composite membrane:

[0033] Main agent in PDMS: auxiliary agent (ie polydimethylsiloxane prepolymer: curing agent) = 10:1. Put the main agent and fixative into the test tube and stir well. Stir until the size of the bubbles in the colloid is uniform, and when there are no obvious large bubbles, it means that the main agent of PDMS and the fixative have been mixed even...

Embodiment 2

[0039] Embodiment 2: establishment of Vibrio parahaemolyticus standard detection curve

[0040] Take the APT-AU-PDMS composite membrane, add 500 μL of Vibrio parahaemolyticus solution of different concentrations, place it on a shaker at 37°C and incubate at 150 rpm for 1 hour, then wash it twice with PBS buffer, and then take 500 μL of the signal probe solution Place it on a shaker at 37° C. and incubate at 150 rpm for 1 h, then wash the APT-AU-PDMS composite membrane twice with PBS buffer, and measure the Raman intensity (excitation light source 633 nm). Such as Figure 4 As shown, Vibrio parahaemolyticus in 1×10 1 -1×10 5 Within the concentration range of cfu / mL and 1590cm -1 The relative Raman intensity has a good linear relationship, the linear equation is y=588.0x-495.8 (R2=0.990), and the minimum detection limit is 12cfu / mL. Specific as Figure 4 shown.

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Abstract

The invention provides a method for detecting vibrio parahemolyticus by an AU-PDMS composite membrane on the basis of aptamer identification. A specific recognition vibrio parahemolyticus aptamer is fixed on the surface of nanogold, a Raman signal module is modified as a signal probe, the specific recognition vibrio parahemolyticus is fixed on an APT-AU-PDMS composite membrane formed by a charge effect to serve as a detection substrate, and a 'sandwich' structure detection sensor is established. The lowest detection limit of the method for the vibrio parahemolyticus can be 12cfu / mL, in addition, the method can be applied to the detection of the vibrio parahemolyticus in shrimp samples, and a result is accurate and reliable.

Description

technical field [0001] The invention relates to the field of food safety detection, in particular to a method for detecting Vibrio parahaemolyticus based on an aptamer-nano gold-PDMS composite film sensor. Background technique [0002] Vibrio parahaemolyticus is a halophilic bacterium that widely exists in seafood such as shrimps, crabs and shellfish, as well as in seawater and seabed sediments. The food poisoning incidents caused by it have surpassed the salmonella poisoning incidents. When people accidentally eat food infected by Vibrio parahaemolyticus, it will cause acute gastroenteritis, diarrhea, sepsis, etc., and cause great harm to health. The outbreak of foodborne diseases caused by Vibrio parahaemolyticus is currently the one of the major public health problems. Vibrio parahaemolyticus is widely distributed in seawater, seawater sediments and aquatic products such as fish, shrimp, crab and shellfish near the coast. In the region, there are reports of a large numb...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/65
CPCG01N21/65
Inventor 段诺沈默斐王周平吴世嘉邹颖
Owner JIANGNAN UNIV
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