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Electrochemical aptamer sensor for ampicillin detection
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An aptamer sensor, ampicillin technology, applied in the field of biosensors, can solve the problems of high cost, low specificity and sensitivity, and achieve the effect of simple electrodes, improved sensitivity and reduced complexity
Active Publication Date: 2021-02-19
UNIV OF JINAN
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[0003] In order to solve the problem of low specificity and sensitivity and high cost of the prior art method for detecting ampicillin, the present invention provides an ampicillin electrochemical sensor with high specificity and sensitivity, low cost and fast detection speed. The chain cycle and the DNA walker on the electrode are used to increase the signal, which greatly improves the detection sensitivity and obtains a lower detection limit, which can be used to detect trace amounts of ampicillin in food
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Embodiment 1
[0038] Embodiment 1 Preparation of Apt-T hybrid chain
[0039] Add 2 μL of 100 μM aptamer Apt, 2 μL of 100 μM T-chain, 2 μL of 5×PBS buffer, 4 μL of ddH 2 O was mixed evenly, reacted in a 95 °C water bath for 5 min and gradually cooled to room temperature to obtain a 10 μM Apt-T hybrid chain solution.
[0041] (1) The gold electrode is polished in 0.3 µM and 0.05 µM aluminaslurry in turn until it becomes a mirror surface, and then it is washed with PBS buffer and ddH 2 O wash repeatedly 3-5 times respectively;
[0042](2) Add 10 μL solution of 2 μM H1 chain onto the surface of the gold electrode obtained in step (1), incubate at 37 °C for 2 h, wash with PBS buffer for 3 times, and obtain H1-modified gold electrode; then add 30 μM H2 chain 10 μL was dropped onto the surface of the H1-modified gold electrode, incubated at 37 °C for 2 h, and washed with PBS buffer 3 times to obtain the H1-H2 modified gold electrode;
[0043] (3) To 0 μL, 0.25 μL, 0.5 μL, 1 μL, 2 μL, 4 μL of the Apt-T hybrid chain solution obtained in Example 1, add 1 μL of 10 nM ampicillin, 1 μL of 20 μM H3 chain solution and 100 μM H4 chain solution 3 μL, add ddH 2 O was added to 10 μL, mixed evenly, and reacted in a 37 °C water bath for 30 mi...
Embodiment 3
[0047] Example 3 H1 chain and H2 chain molar ratio screening
[0048] (1) The gold electrode is polished in 0.3 µM and 0.05 µM aluminaslurry in turn until it becomes a mirror surface, and then it is washed with PBS buffer and ddH 2 O wash repeatedly 3-5 times respectively;
[0049] (2) Add 10 μL of 2 μM H1 chain solution onto the surface of the gold electrode obtained in step (1), incubate at 37 °C for 2 h, wash with PBS buffer three times, and obtain H1-modified gold electrodes; Add 10 μL of H2 chains of μM, 10 μM, 20 μM, 30 μM, 40 μM, 50 μM onto the surface of H1-modified gold electrode, incubate at 37 °C for 2 h, wash with PBS buffer 3 times, and obtain H1-H2 Modified gold electrodes;
[0050] (3) Add 1 μL of Apt-T hybrid chain solution obtained in Example 1, 1 μL of 10 nM ampicillin, 1 μL of 20 μM H3 chain solution and 5 μL of 100 μM H4 chain solution, add ddH 2 O was added to 10 μL, mixed evenly, and reacted in a water bath at 37 °C for 30 min to obtain a reaction sol...
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Abstract
The invention provides an electrochemical aptamer sensor for ampicillin detection. The electrochemical aptamer sensor comprises a detection electrode, heme, an H3 chain, an H4 chain, an aptamer Apt and a T chain, wherein the sequences of the H3 chain, the H4 chain, the aptamer Apt and the aptamer T chain are shown as SEQ ID NO: 3-6; the detection electrode is a gold electrode of which the surfaceis modified with an H1 chain and an H2 chain, the sequences of the H1 chain and the H2 chain are shown as SEQ ID NO: 1-2, and the 5' end of the H1 chain and the 3' end of the H2 chain are modified with sulfydryl. According to the sensor, signals are amplified through circulation of a T chain and a DNA walker on an electrode, so that the detection sensitivity is greatly improved, a lower detectionlimit is obtained, and the sensor can be used for detecting trace ampicillin in food.
Description
technical field [0001] The invention belongs to the technical field of biosensors, and relates to an electrochemical aptamer sensor used for the detection of ampicillin. Background technique [0002] Ampicillin (AMP) is a β-lactam antibiotic belonging to the penicillin class, the most commonly used class of antibiotics. Ampicillin can inhibit bacterial wall synthesis and is widely used in human and veterinary treatment to prevent and treat gastrointestinal, respiratory, skin and genitourinary bacterial infections caused by bacterial infections such as streptococcus and staphylococcus. However, excessive use of ampicillin or residues of ampicillin in food or the environment can lead to serious food safety and environmental safety problems, causing symptoms such as allergic reactions and epilepsy in humans. It is of great significance to establish efficient and reliable detection methods. Conventional analytical techniques such as high performance liquid chromatography (HPLC)...
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