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A biosensor for the detection of Salmonella

A biosensor, Salmonella technology, applied in the field of biosensors, can solve the problems of long detection cycle, low specificity and sensitivity, high cost, etc., and achieve the effect of short detection cycle, improved sensitivity, and easy operation

Active Publication Date: 2021-08-17
UNIV OF JINAN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In order to solve the problems of relatively low specificity and sensitivity, high cost and long detection period of the method for detecting Salmonella in the above prior art, the present invention provides a SERS signal-based method with high specificity and sensitivity, low cost and fast detection speed. Conductive biosensors for the detection of Salmonella

Method used

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  • A biosensor for the detection of Salmonella
  • A biosensor for the detection of Salmonella
  • A biosensor for the detection of Salmonella

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Preparation of nano gold.

[0042] (1) Add 200ml of ultrapure water into the three-neck flask;

[0043] (2) Take 500 μL of HAuCl with a concentration of 0.04g / mL 4 In the centrifuge tube, add 200ml ultrapure water, stir and heat to boil, stirring speed 450rpm;

[0044](3) Under stirring conditions, take 3ml of 1% trisodium citrate solution and quickly add it to the solution in step (2). The color of the solution changes from light yellow to wine red. After continuing to heat for 15 minutes, remove the heat source and Slowly cool to room temperature, and store at 4°C for later use.

[0045] According to the absorbance value at 530nm, the concentration of gold nanoparticles in the above solution is about 0.3nM.

Embodiment 2

[0046] Example 2 Preparation of gold nanoparticles modified with Raman dyes.

[0047] Take 2 mL of nano-gold solution in a centrifuge tube, centrifuge at 10,000 rpm for 10 min, remove the supernatant, disperse the bottom precipitate into 600 μL of sterilized water, and concentrate the nano-gold solution to 3 nM. Transfer to a 1 mL glass bottle and seal with tinfoil.

[0048] Take 12 μL of DTNB solution with a concentration of 50 μM and add it to the solution treated in step 1, and continue to shake in a mixer for 1 h.

[0049] The resulting solution was centrifuged at 10,000 rpm for 10 min, the supernatant was removed, and the bottom pellet was redispersed into 2 mL of sterilized water. This process was repeated three times to remove excess dye. After the final centrifugation, DTNB-modified gold nanoparticles were dispersed into 600 μL of PB buffer (10 mM, pH 7.4), and stored at 4 °C for future use.

Embodiment 3

[0050] Example 3 Effects of different H1 concentrations on the detection of Salmonella.

[0051] (1) Add 14 μL sterilized water, 2 μL 5×PBS, 2 μL 100 μM Apt chain and 2 μL 100 μM T chain into the pre-prepared sterilized EP tube, shake for 30s, and incubate at 95°C After 5 minutes, slowly cool down to room temperature and hybridize into probes, and store at -20°C for use.

[0052] (2) Add probe (4 μL, 10 μM), H1 (2 μL, final concentrations are 0.4 μM, 0.6 μM, 0.8 μM, 1.0 μM, 1.2 μM, 1.4 μM), H2 (2 μL, 20 μM), 5×PBS ( 3 μL) and 3 μL Salmonella suspension (10 5 cfu / mL) into a centrifuge tube, shaken for 30 s, and placed in a 37°C water bath for 30 min.

[0053] (3) Take the reacted solution out of the water bath, add nano gold solution (10 μL) of modified DTNB, cysteine ​​(2 μL) and 4 μL heme (final concentration is 1 μM), and put it in a 37°C React in a water bath for 1 h.

[0054] (4) The reaction solution in step (3) is scanned by a Laboram010 laser confocal Raman spectro...

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Abstract

The present invention provides a biosensor for detecting Salmonella, including aptamer (Apt), template (T), hairpin probe 1 (H1), hairpin probe 2 (H2), heme, potassium ion (K + ), cysteine, gold nanoparticles modified with Raman dyes and buffer, can use surface Raman enhanced detection of Salmonella in food. The present invention utilizes good specificity and high detection sensitivity; mild reaction conditions and fast reaction speed; compared with other optical detection methods, the detection method is easy to operate and has a short detection cycle; the detection process is realized in a homogeneous phase, reducing the The complexity of the operation; no need for enzymes to effectively reduce the process cost of the biosensor, and is suitable for low-cost requirements in industrialization.

Description

technical field [0001] The invention relates to a biosensor for detecting Salmonella based on SERS signal transduction, in particular to a biosensor based on hybridization chain reaction amplification (HCR) and G-quadruplex DNase inhibition plasma resonance coupling and SERS in the presence of cysteine The invention relates to a biosensor for signal transduction detection of Salmonella, belonging to the technical field of biosensors. Background technique [0002] Salmonella is a common foodborne pathogen, a Gram-negative, intracellular parasitic enteric bacterium. The bacteria widely exist in nature, not only can cause acute, chronic or latent infection of livestock, poultry and other animals, but also cause food poisoning by contaminating food, posing a great threat to human beings. According to statistics, among the types of bacterial food poisoning in various countries in the world, food poisoning caused by Salmonella often ranks first. Salmonella sickens about 40,000 A...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/682C12Q1/06G01N21/65
CPCC12Q1/682G01N21/658C12Q2525/205C12Q2525/301C12Q2563/137C12Q2565/632Y02A50/30
Inventor 刘素冷雪琪王玉黄加栋
Owner UNIV OF JINAN
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