Preparation method and detection method of nanocomposite material, lps electrochemical aptamer sensor
A nano-composite material and electrochemical technology, which is applied in the field of preparation of nano-composite materials and LPS electrochemical aptasensors, can solve the problem that the detection results are easily affected by enzymes and impurities, operators need professional training, and complex sample matrix interferes greatly and other problems, to achieve high specificity, clear and simple steps, and reduce experimental costs
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Embodiment 1
[0041] Embodiment 1 prepares P-rGO-TNT-Ag nanocomposite material
[0042] Follow the steps below:
[0043] 1) Disperse 10mg GO in 25mL ultrapure water, ultrasonically homogenize, then drop about 20μL of ammonia water to make the pH around 8-9, then add 250mg of ascorbic acid (AA) to it, stir at room temperature for 0.5h, then place in an oil bath at 95°C Continue to stir for 1 h, centrifuge and wash, and then disperse the precipitate in 10 mL ultrapure water to obtain the rGO dispersion;
[0044] 2) Add 20uL of 20wt% polydiallyldimethylammonium chloride solution (PDDA) dropwise to 1mL of the rGO dispersion prepared in step 1), and stir for 0.5h to obtain the P-rGO dispersion;
[0045] 3) 1g TiO 2 Add the powder to 20mL 10mol / L NaOH solution, stir at room temperature for 1h, then transfer the solution to a reaction kettle at 130°C for 10h, cool, centrifuge, wash with 1M HCl solution until pH = 1, and then wash with ultrapure water to pH =7, then the precipitate is dispersed ...
Embodiment 2
[0048] Example 2 Preparation of an electrochemical DNA aptamer sensor for LPS detection
[0049] Follow the steps below: (The construction principle is as follows figure 1 shown)
[0050] 1) Treat the amino-labeled LPS-binding aptamer (LBA) with 20 mM Tris-HCl (pH=7.4) buffer solution at room temperature, and place it at 4°C for use;
[0051] 2) Wash the gold electrode with piranha washing solution (98% H 2 SO 4 / 30%H 2 o 2 =3:1, v / v) Rinse with ultra-pure water after soaking for 30 minutes;
[0052] 3) Use the electrodes obtained in step 2) with 0.3 μm and 0.05 μm Al 2 o 3 The powder was polished to a mirror surface, and then the electrodes were ultrasonically treated in the order of ultrapure water, absolute ethanol, and ultrapure water for 5 minutes each, and dried at room temperature for later use.
[0053] 4) Place the electrode obtained in step 3) in 0.5M H 2 SO 4 Electrochemical activation was carried out in the medium, and the potential scan was -0.3~1.55V un...
Embodiment 3
[0057] Example 3 Detection of LPS using an electrochemical DNA aptamer sensor
[0058] Using the electrochemical DNA aptamer sensor constructed in Example 2 to detect LPS, follow the steps below:
[0059] 1. Draw the working curve
[0060] 1) Place the modified electrodes from Step 4) to Step 7) of Example 2 in a solution containing 10mM KCl and 2mM MgCl respectively. 2 Characterized in 0.1M PBS (pH=7.0), and measured its electrochemical response signal, the results are as follows figure 2 Shown: (a) bare gold electrode; (b) dropwise addition of P-rGO-TNT-Ag nanocomposite; (c) combined with LBA; (d) blocked by BSA.
[0061] 2) Add 20 μL of target endotoxin with different concentrations to the electrode of the aptasensor prepared in Example 2, and measure the current changes respectively.
[0062] 3) According to the linear relationship between the obtained current change value and the logarithmic value of the endotoxin concentration, draw a working curve (such as image 3...
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