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Kit, probe and method for rapid and sensitive detection of ofloxacin

A technology for sensitive detection of ofloxacin, applied in biochemical equipment and methods, biological testing, measuring devices, etc., can solve the problems of low detection limit, high sensitivity, shortening time, etc., and achieve large Stokes shift, Fluorescence has a long lifespan and eliminates the effect of background fluorescence

Active Publication Date: 2020-07-10
XIHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to provide a kit, probe and method for rapid and sensitive detection of ofloxacin, which solves the problems of expensive equipment, long cycle and unfavorable on-site inspection in the existing detection method, and can shorten the time and reduce the detection time. Cost, low detection limit, high sensitivity

Method used

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  • Kit, probe and method for rapid and sensitive detection of ofloxacin
  • Kit, probe and method for rapid and sensitive detection of ofloxacin
  • Kit, probe and method for rapid and sensitive detection of ofloxacin

Examples

Experimental program
Comparison scheme
Effect test

experiment example 1

[0044] Experimental Example 1 Aminated Fe 3 o 4 Synthesis of nanoparticles

[0045] Weigh 2.0g CH 3 COONa and 1.0 g FeCl 3 ·6H 2 O in a small beaker, and then add 30 mL of ethylene glycol. Add 6.5g of 1,6-hexamethylenediamine melted in a water bath at 50°C to the above mixture, stir magnetically in an oil bath at 50°C, until a wine-red transparent solution is formed, transfer the solution to a high-pressure reactor, and heat at 198°C High temperature reaction 6h.

[0046] After the reaction is completed, place the reaction kettle at room temperature to cool naturally, use a magnet to separate and collect the product, discard the upper liquid, wash the product with ultrapure water and absolute ethanol alternately for three times, and finally dry the black powder at 50°C for 10 hours , and the dried product is ground to powder for later use.

experiment example 2

[0047] Experimental Example 2 Amination of NaYF 4 : Synthesis of Ce / Tb Time-Resolved Fluorescent Nanoparticles

[0048] Synthesis of Na YF by one-step solvothermal method 4 : Ce / Tb inorganic nanomaterials, the specific process is as follows:

[0049] Weigh 1mmol of phosphoethanolamine (AEP) and 1mmol of NaCl dissolved in 30mL of ethylene glycol with constant stirring, then add 0.9mmol of yttrium nitrate hexahydrate (Y(NO 3 ) 3 ·6H 2 O), 0.05mmol cerium nitrate hexahydrate (Ce(NO 3 ) 3 ·6H 2 O), 0.05mmol terbium nitrate pentahydrate (Tb(NO 3 ) 3 ·5H 2 O), continue to stir until dissolved. With stirring in a water bath at 45°C, 4 mmol of ammonium fluoride (NH 4 F) Dissolve in 10 mL of ethylene glycol solution, and then add dropwise to the above clear solution.

[0050] The above mixed solution was vigorously stirred at room temperature for 40 min, then transferred to a high-pressure reactor, and reacted at 180° C. for 4 h.

[0051] After the reaction was completed, ...

experiment example 3

[0052] Experimental example 3NaYF 4 : Construction of Ce / Tb-cDNA Time-resolved Fluorescent Probe or Nano Magnetic Bead-Apt Probe

[0053] Get 5mg nanometer material (aminated Fe of above-mentioned preparation 3 o 4 Nanoparticles or Aminated NaYF 4 : Ce / Tb time-resolved fluorescent nanoparticles) dispersed in 5mL of PBS buffer (10mmol / L Na 2 HPO 4 , 2mmol / L KH 2 PO 4 , 2.7mmol / LKCl, 137mmol / L NaCl, pH 7.4) and added 1.25mL glutaraldehyde solution (25%, V / V), incubated at 25°C for 3h under gentle shaking, separated and obtained activated nanomaterials, and used Nanomaterials were washed with PBS buffer.

[0054] The washed activated nanomaterials were dispersed in 5 mL of 0.1 mg / mL biotin solution and incubated at 37° C. for 12 h, and then the nanomaterials were washed with PBS buffer to remove unimmobilized avidin to obtain modified avidin nanomaterials.

[0055] Disperse the modified avidin nanomaterial in 5mL PBS buffer, add 90 μL 10 μmol / L nucleic acid sequence P1 (a...

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Abstract

The invention discloses a kit, a probe and a method for rapid and sensitive detection of ofloxacin. The kit comprises a nano magnetic bead-nucleic acid aptamer probe, an RCA primer, an RCA template, aT4 DNA ligase, a Phi29 DNA polymerase and a NaYF4: Ce / Tb-cDNA time-resolved fluorescent probe. Fe3O4 nano magnetic beads are used as a rapid magnetic separation carrier, a nucleic acid aptamer specifically combined with ofloxacin is used as a recognition element, a nucleic acid signal is amplified through a rolling circle amplification technology, and time-resolved fluorescence is used as a signal source so that a novel technology for specific rapid sensitive detection of residual antibiotic ofloxacin in food is constructed. The method is short in detection period without depending on the chromatographic instrument and high in sensitivity and is as low as pmol / L in detection limit and quantification limit and is suitable for detection personnel in the market or enterprises.

Description

technical field [0001] The invention relates to a kit for ofloxacin detection, in particular to a kit, a probe and a method for rapid and sensitive detection of ofloxacin. Background technique [0002] Ofloxacin (OFL) belongs to fluoroquinolone antibiotics, which has good inhibitory effect on staphylococcus, streptococcus, Streptococcus pneumoniae, Neisseria gonorrhoeae and other pathogenic bacteria. As a commonly used drug in animal husbandry, ofloxacin is widely used in the treatment and prevention of respiratory and urinary system diseases of livestock. Ofloxacin is a synthetic antibiotic that is difficult to degrade naturally. Wastewater from abuse and animal metabolism entering the environment will cause long-term pollution in soil and water. Ofloxacin and other veterinary drug residues accumulate in target organs such as liver and kidney through the food chain, causing allergic reactions or organ damage in some people. The accumulation of antibiotics such as ofloxaci...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/53C12Q1/682C12Q1/6811C12N15/11
CPCG01N33/9446C12Q1/682C12Q1/6811C12Q2531/125C12Q2521/501C12Q2563/143C12Q2563/155C12Q2563/107C12Q2525/205Y02A50/30
Inventor 黄玉坤王冲陈祥贵杜钢王力均杨潇
Owner XIHUA UNIV
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