A visual detection sensor based on ferric oxide nanocluster catalytic signal amplification and its construction and application
A detection sensor, ferric oxide technology, applied in the direction of chemical reaction analysis of materials, material analysis by observation of the influence of chemical indicators, preparation of test samples, etc., can solve the problems of low sensitivity, Achieve the effect of improving detection sensitivity, good stability, and simple and convenient construction
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Embodiment 1
[0095] Embodiment 1: synthesizing functionalized Fe with catalytic activity 3 o 4 NPC probe, specifically includes the following steps:
[0096]1. Take 20mL of ethylene glycol as the base solution, mix and add 0.6g of ferric chloride hexahydrate, stir to form a clear solution, add 1.5g of sodium acetate to the clear solution, stir the mixed solution vigorously for 30 minutes, add the evenly stirred mixed solution After reacting in a polytetrafluoroethylene autoclave at 200°C for 16 hours, cool the autoclave at room temperature, wash the product with ultrapure water and ethanol, remove excess liquid with a magnetic separator, and dry the product in vacuum for 12 hours to obtain a surface Carboxylated Fe 3 o 4 NP;
[0097] 2. Dissolve 1 mg EDC in 1×PBS buffer by vortexing, and 1 mg Fe 3 o 4 Add NP to it, incubate at room temperature for 30 minutes to activate, add 12.5 μg PLL, and incubate overnight at 4°C. Remove excess unreacted PLL and EDC with a magnetic separator, Fe...
Embodiment 2
[0101] Embodiment 2: Construct based on above-mentioned Fe 3 o 4 Visual detection sensor for NPC probe catalytic signal amplification:
[0102] 1. Activate 10 mg Van with 300 μL EDC (20 mg / mL) and 300 μL NHS (1.0 mg / mL), add 250 μL BSA (1.0 mg / mL) to the activated Van solution, and incubate overnight at 4°C. The reaction was terminated by adding 40 μL of Tris buffer (50 mM, pH 7.2) to the BSA-Van mixture. Use a 10kDa ultrafiltration tube to remove excess unreacted Van and EDC by centrifugation at 5000rpm.
[0103] 2. Add BSA-Van to the coating solution (0.01M, Tris-HCl buffer solution with pH 8.5), coat the 96-well plate with 50 μg / mL BSA-Van, 100 μL per well, and shake on a shaker at 600-700 rpm overnight at 4°C. The coating solution was removed, and the modified wells were washed three times with a washing solution (1×PBS buffer solution containing 0.05% Tween-20), 300 μL per well. Then block the microplate with blocking solution (PBS buffer containing 3% BSA and 0.05% ...
Embodiment 3
[0104] Example 3 is based on Fe 3 o 4 The NPC probe catalytic signal amplification visual detection sensor can quickly and visually detect Listeria monocytogenes. The specific steps are as follows:
[0105] 1. Detect bacterial sample preparation, the bacterium detected in the laboratory is Listeria monocytogenes (Listeria monocytogenes) (bacterial strain number is CMCC54007, purchased from Guangzhou Institute of Microbiology) in 10mL LB medium (the formula of LB medium is as follows: 1.0g tryptone, 0.5g of yeast extract, and 1.0g of NaCl were added to 100mL of water, and the pH was adjusted to 7.0 with NaOH solution), and the overnight cultured bacteria had an OD600 of 1 and a concentration of about 1×10 8 cfu / mL, take 1mL of the above bacterial solution, centrifuge at 5000rpm for 5 minutes, remove the medium, wash with PBS buffer three times, and redisperse the bacterial cells in 0.1M PBS buffer, and carry out gradient dilution to obtain the tested bacteria at each concentra...
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