Aptamer-based rapid kanamycin detection test paper as well as preparation method and application thereof

A nucleic acid aptamer, kanamycin technology, applied in measurement devices, instruments, scientific instruments and other directions, can solve the problems of long time, high operator requirements, high cost, easy preparation and modification, improve detection efficiency, The effect of short detection time

Active Publication Date: 2017-07-28
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the application of the above method is still partially limited, such as long time-consuming, large-scale equipment and relatively large consumption of reagents, high cost, susceptible to interference, high requirements for operators, and not suitable for field applications, etc.

Method used

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  • Aptamer-based rapid kanamycin detection test paper as well as preparation method and application thereof
  • Aptamer-based rapid kanamycin detection test paper as well as preparation method and application thereof
  • Aptamer-based rapid kanamycin detection test paper as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Example 1 A rapid detection test paper for kanamycin based on nucleic acid aptamer

[0041] Preparation and functionalization of gold nanoparticles, preparation and functionalization of silver nanoparticles, pretreatment of gold standard pads and sample pads, pretreatment of nitrocellulose membranes, assembly of test strips, and detection of color development of test strips. The specific steps are:

[0042] (1) Preparation and functionalization of gold nanoparticles:

[0043] Before preparing gold nanoparticles, all glass instruments were soaked in aqua regia for 30min, then washed with distilled water, soaked in ultrapure water for 12h, and dried for later use; 100mL of 0.01% chloroauric acid was added to a 250mL round bottom flask, stirred and heated To boiling, quickly add 3.5mL of 1% trisodium citrate under vigorous stirring, continue to heat and stir for 15 minutes, and the solution turns wine red, stop heating, continue to stir for 30 minutes, let it stand, and c...

Embodiment 2

[0056] Embodiment 2 Determination of kanamycin standard solution with test strips

[0057] Use 2mL centrifuge tubes to dilute kanamycin to different concentrations, 100 µL per tube. Insert one end of the sample pad of the test strip prepared by the above method into the standard solution of kanamycin with different concentrations, and react at room temperature for 10 min. From Figure 4 It can be seen that the color of the T line basically does not change between 0-1 nmol / L and 35-400 nmol / L of kanamycin concentration, while the color of the T-line changes with the concentration of kanamycin between 1-35 nmol / L. It becomes lighter with the increase of the element concentration, and the detection line basically has no color after 35 nmol / L. Therefore, the naked eye detection limit of the test paper for kanamycin was 35 nmol / L. Put the reacted test strip into a plastic card and use a colloidal gold quantifier to detect the color development of the T and C lines, and obtain th...

Embodiment 3

[0058] Example 3 Detection of Kanamycin Residues in Milk Samples

[0059] Here, artificially contaminated milk was prepared by adding standard concentration of kanamycin to milk to obtain 5 nmol / L, 10 nmol / L, 20 nmol / L, 30 nmol / L, 50 nmol / L, 100 nmol / L , 200 nmol / L kanamycin milk samples. Add 20% trichloroacetic acid dropwise to the milk sample, adjust the pH to 4.6, and precipitate casein in a water bath at 45°C for 10 minutes, centrifuge at 10,000 r / min for 25 minutes to remove coagulated protein and fat, and filter with a 0.22 μm filter membrane , and finally adjust the pH to neutral to obtain the pretreated milk sample. The milk samples containing different concentrations of kanamycin were detected according to the above operation steps. get Figure 6 The test strips of milk samples containing different concentrations of kanamycin are shown. exist Figure 6 It can be seen from the figure that as the concentration of kanamycin increases, the color of the T line becomes...

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Abstract

The invention discloses aptamer-based rapid kanamycin detection test paper as well as a preparation method and application thereof, and belongs to the field of analytical chemistry as well as safety detection on medicine, environment and food. The aptamer-based rapid kanamycin detection test paper comprises a sample pad, a colloidal gold pad, a nitrocellulose membrane, a water absorbing pad, a PVC bottom plate, a detection line and a quality control line. By the preparation method, by virtue of the characteristics that gold nanoparticles and silver nanoparticles are easy to prepare and modify and can dually amplify a signal as well as the conformational change of an aptamer, the strength change of an optical signal of the detection line is achieved, and thus detection on kanamycin is achieved; by using a test paper method for detecting the kanamycin, simple, convenient and fast detection is achieved, detection can be performed anytime and anywhere, a result is visualized, the detection time is short, an experimental result can be obtained within 10min by just inserting one end of the sample pad of the test paper into detection liquid, and the detection efficiency can be greatly improved; through a reading of a colloidal gold quantometer, quantitative analysis can be achieved; therefore, the aptamer-based rapid kanamycin detection test paper is of great significance in detection on kanamycin residue in animal-derived food.

Description

technical field [0001] The invention relates to a nucleic acid aptamer-based rapid detection test paper for kanamycin and its preparation method and application, belonging to the fields of analytical chemistry, medicine, environment and food safety detection. Background technique [0002] Kanamycin is a broad-spectrum aminoglycoside antibiotic with the molecular formula C 18 h 36 N 4 o 11 , with a molecular weight of 582.58, can effectively treat infections caused by Gram-positive or Gram-negative bacteria. However, excessive use of kanamycin or excessive consumption of animal-derived foods containing kanamycin can cause kanamycin to accumulate in the human body through the food chain and may cause some rather serious side effects, such as affecting the cochlear nerve, inducing Ototoxicity, leading to permanent hearing loss; at the same time, it may also have a certain impact on the neuromuscular and hematopoietic system; in addition, kanamycin can also cause damage to t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/558
CPCG01N33/558
Inventor 周楠迪刘晶曾静怡田亚平
Owner JIANGNAN UNIV
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