Non-diagnostic purpose method for quantitatively detecting Salmonella pullorum and Salmonella gallinarum

A quantitative detection method, the technology of Salmonella, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of poultry breeding industry loss, threat to food safety, etc., and achieve good sensitivity, convenient detection method, and easy operation.

Inactive Publication Date: 2018-08-03
ZHEJIANG GONGSHANG UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002] Salmonella pullorum (S.pullorum) and Salmonella gallinarum (S.gallinarum) are the pathogens of pullorum (Pullorum Disease, PD)

Method used

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  • Non-diagnostic purpose method for quantitatively detecting Salmonella pullorum and Salmonella gallinarum
  • Non-diagnostic purpose method for quantitatively detecting Salmonella pullorum and Salmonella gallinarum
  • Non-diagnostic purpose method for quantitatively detecting Salmonella pullorum and Salmonella gallinarum

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] (1) Synthesis of silica nanoparticles (SiNPs):

[0047] SiNPs were synthesized by the inverse microemulsion method. The specific synthesis process is as follows: 2mL Triton X-100 (surfactant), 2mL n-hexanol (cosurfactant), 8mL cyclohexane (organic solvent) were stirred for 20min until clear and transparent, and 480μL ultrapure water was added as the dispersed phase , stirred for 20 min to make the water droplets completely dispersed in the above oil phase to form a uniform and stable W / O microemulsion, and then added 100 μL tetraethyl orthosilicate (TEOS) as a precursor for the synthesis of SiNPs, and stirred for 30 min. Add 100 μL of ammonia water (catalyst) to the system solution to promote TEOS to form nano-microspheres through hydrolysis and condensation reaction faster. After 48 hours of magnetic stirring, 10 mL of acetone was added to demulsify the microemulsion, and the precipitate was collected after centrifugation at 10,000 rpm for 5 minutes, and then washed t...

Embodiment 2

[0060] Example 2: GOx triggers an enzymatic cascade to bind Fe 3+ -SCN - Establishment of quantitative detection method for pullorum pullorum and Salmonella gallinarum typhi by colorimetric system

[0061] (1) Optimization of the concentration of antibody-coupled nano-magnetic beads

[0062] Change the concentration of antibody-coupled nanomagnetic beads IgG-MNPs: 2.4mg / mL, 3mg / mL, 4mg / mL, 6mg / mL, 12mg / mL, 24mg / mL, 48mg / mL, other experimental conditions are the same as the implementation The same as Example 1, quantitative detection of 1mL 10 7 CFU mL -1 The samples of S.pullorum and S.gallinarum to be tested. Depend on figure 2 It can be seen that as the concentration of IgG-MNPs increases, the absorbance value gradually reaches the maximum value and then maintains a plateau period and then shows a downward trend. Therefore, the optimal concentration of IgG-MNPs for analysis and testing is 6 mg / mL.

[0063] (2) Optimization of enzyme marker concentration

[0064] Chan...

Embodiment 3

[0065] Embodiment 3: Sensitivity detection

[0066] Detect 8.4×10 under the condition of embodiment 1 0 CFU mL -1 to 8.4×10 7 CFU mL -1 Pullorum and Salmonella gallinarum (S.pullorum and S.gallinarum) were used to determine the sensitivity. The relationship between the absorbance value and the concentration of Pullorum pullorum and Salmonella gallinarum typhi is as follows: Figure 4 As shown, when the concentration of S.pullorum and S. gallinarum is 8.4×10 3 CFU mL -1 to 8.4×10 7 CFU mL -1 When changing, the absorbance value has a good linear relationship with the log value of the concentration of S.pullorum and S.gallinarum. The linear equation is y=0.1992x-0.31102, the coefficient of variation R 2 = 0.98706. The detection limit of the method for the target bacteria S.pullorum and S.gallinarum was 2.36×10 3 CFU mL -1 .

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Abstract

The invention discloses a non-diagnostic purpose method for rapidly detecting Salmonella pullorum and Salmonella gallinarum. The detection method comprises the following steps: preparing Salmonella pullorum and Salmonella gallinarum antibody labeled coupled magnetic nano-particles (IgG-MNPs) and antibody and enzyme double labeled silica nano-particles (GOx-IgG-SiNPs), forming an immune compound IgG-MNPs/S having a sandwich structure, S. Pullorum and S. Gallinarum GOx-IgG-SiNPs, when the target bacteria to be detected exist, performing an enzymatically catalyzed reaction, allowing the obtainedreaction product and a color developer to form a complex product, and detecting the absorbance value of the obtained system to obtain the concentrations of the Salmonella pullorum and Salmonella gallinarum in order to realize the quantitative detection of the Salmonella pullorum and Salmonella gallinarum. A cascade reaction triggered by glucose oxidase (GOx) is combined with an Fe<3+>-SCN<-> colordevelopment system to realize the quantitative detection of Salmonella pullorum and Salmonella typhimurium. The detection method has the advantages of convenience, fastness, easiness in operation, good sensitivity, good specificity and good stability.

Description

technical field [0001] The invention belongs to the technical field of food and food raw material safety detection, in particular to a non-diagnostic non-diagnostic non-diagnostic pullorum and salmonella gallinarum typhi quantitative detection method. Background technique [0002] Salmonella pullorum (S.pullorum) and Salmonella gallinarum (S.gallinarum) are the pathogens of pullorum (Pullorum Disease, PD) and chicken typhoid (Fowl typhoid, FT), causing huge damage to the poultry industry. losses and seriously threaten food safety. Since Salmonella pullorum and Salmonella gallinarum have the same O1, O9, O12 antigens, it is difficult to strictly distinguish the two and it is not necessary in actual testing, so the two are usually tested together. The detection of Salmonella pullorum and Salmonella gallinarum is very meaningful for the prevention and control of food safety risks. [0003] The detection methods for pathogenic bacteria include standard methods and a variety of...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N33/569
CPCG01N33/54346G01N33/56916
Inventor 窦文超赵广英朱海涛
Owner ZHEJIANG GONGSHANG UNIVERSITY
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