Superparamagnetic three-in-one aptamer sensing platform, preparation method and application

Abstract

The invention discloses a superparamagnetic three-in-one aptamer sensing platform, a preparation method and application, and belongs to the technical field of bacterial detection. The preparation method comprises the following steps: dissolving FeCl3. 6H2O, MnCl2. 4H2O and ZnCl2 in water, and reacting with NaOH to obtain ZMF NPs; the preparation method comprises the following steps: adding HAuCl4. 3H2O and sodium citrate into water, then adding NaBH4, and stirring in a dark place to obtain Au NPs; the method comprises the following steps: modifying the surface of ZMF NPs by using PEI-DTC, mixing the modified ZMF NPs with an Au NPs solution, and carrying out ultrasonic treatment to obtain ZMF coated PEI-DTC / Au NPs; then NH2OH. HCl and ZMF (at) PEI-DTC / Au NPs are added into the growth solution, and ZMFA NPs is obtained; the method comprises the following steps: activating an MRSA specific nucleic acid aptamer through a TCEP solution, mixing with ZMFA NPs, soaking in a NaCl solution, separating to obtain ZMFA-A NPs, mixing with Blockers, soaking in the NaCl solution, and separating to obtain ZMFA-AB NPs. The sensing platform has the advantages of high sensitivity, high specificity, simplicity and convenience in operation, short detection time and the like, can be used for specific capture, separation and ultra-sensitive dual-mode detection of the MRSA, and simultaneously realizes efficient photo-thermal removal.

Description

Technical Field

[0001] The present invention belongs to the technical field of bacterial detection, and particularly relates to a superparamagnetic triple aptamer sensing platform, a preparation method and an application thereof. Background Art

[0002] In recent years, with the extensive use of antibiotics, the infections of drug-resistant bacteria have been increasing, becoming a focus of attention at home and abroad. Taking Methicillin-resistant Staphylococcus aureus (MRSA) as an example, since MRSA was first discovered by British scholar Jevons in 1961, MRSA has become the main pathogenic bacterium of hospital and community infections and has spread rapidly all over the world. At present, most MRSA shows cross-resistance to first-generation and second-generation cephalosporins, clindamycin, levofloxacin, and erythromycin, and the drug resistance rate is 70% - 90%.

[0003] There are numerous cases related to MRSA infection. Therefore, quickly and accurately detecting pathogenic bacteria is of great significance for giving precise treatment plans and reducing the occurrence of severe infections. At present, the most classic method is the bacterial culture method most commonly used in clinical microbiology laboratories. However, the culture method is time-consuming and laborious, and if drug resistance detection is required, the required operation steps are more complicated, and its disadvantages are self-evident; some other more advanced methods have also been developed, such as polymerase chain reaction (PCR), high performance liquid chromatography (HPLC), and surface enhanced Raman spectroscopy (SERS), etc., which have played a certain role in clinical microbiological identification. However, these methods require expensive analytical instruments, professional operators, and a fixed and strict experimental environment. Therefore, developing a detection method for early, rapid, convenient, and accurate identification of MRSA is crucial for reducing the occurrence of severe infections.

[0004] Molecular recognition elements are bioactive substances that can specifically recognize targets, forming a complex by recognizing and binding to the targets. Currently, molecular recognition elements based on the specific interaction between antigens and antibodies are the most widely used. However, due to the complexity of the matrix in environmental, food, and clinical samples, the stability and sensitivity of current immunoassay techniques based on antigen-antibody interaction still cannot meet the high-sensitivity detection requirements in various situations. In addition, the preparation cycle of traditional antibodies is long, a large number of experimental animals are used, the production cost is expensive, and the chemical and thermal stabilities of traditional antibodies are poor, which is not suitable for long-term storage. Compared with antibodies, nucleic acid aptamers have the characteristics of high stability, no immunogenicity, easy modification, low cost, and easy large-scale in vitro synthesis. They can recognize a wide range of target molecule types, including cells, bacteria, proteins, and various small molecule substances, and can maintain high affinity and specificity at the same time. Therefore, nucleic acid aptamers are also called "chemical antibodies" and have now been combined with different detection techniques and widely used in the fields of biomedicine, environmental analysis, and food engineering, showing broad application prospects in on-site rapid detection.

[0005] Dynamic light scattering (DLS) technology is a commonly used physical characterization method. By measuring the fluctuations of the scattered light of the sample, the average particle size and particle size distribution of the sample can be calculated. DLS has been used as an emerging immunosensing tool. Compared with the absorbance level, the light scattering intensity of the nanoprobe shows extremely high response efficiency to size changes, with excellent sensitivity and specificity. It has currently been used to detect target analytes such as proteins, nucleic acids, small molecules, ions, and microorganisms and has been widely applied in the fields of food safety, environmental monitoring, and clinical diagnosis. However, in the actual sample detection by DLS technology, the complex matrix in the sample often causes a high background scattering signal, resulting in a decrease in the signal-to-noise ratio of the detection and even leading to the failure of the detection. To solve this constraint, many studies have introduced some enrichment methods (such as magnetic enrichment separation technology) in the pretreatment process to remove the matrix interference in the reaction system. However, these methods have prolonged the operation time, added experimental procedures and uncontrollable factors. In addition, at high bacterial concentrations, the interference caused by the light scattering intensity becomes obvious. Therefore, it is particularly important to introduce another reliable detection method to ensure the accuracy of the detection.

[0006] The CRISPR / Cas-mediated detection system is a very powerful and advanced technology, characterized by high sensitivity, fast reaction time, strong lysis ability, good stability, etc., and is suitable for the detection of trace bacteria in the early stage of diseases. The CRISPR-Cas system consists of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR associated proteins, namely Cas proteins. In addition, more research teams have established a variety of new rapid and sensitive nucleic acid detection technologies based on the characteristics of these nucleases, laying the foundation for the development of "next-generation" molecular diagnostic technologies. Currently, most in vitro CRISPR detection systems are designed based on activating the nuclease activity of Cas proteins. However, due to the different sources and cleavage characteristics of the used Cas nucleases, and the different crRNA sequences, the detection performances of the established various methods also vary. It is worth noting that some research teams have found that Cas12a nucleases can specifically recognize single-stranded DNA and can cleave single-stranded DNA in cis and trans. In addition, CRISPR-based methods reduce the need for large equipment and have the possibility of on-site implementation, especially in resource-limited areas. Therefore, DLS and CRISPR / Cas methods can be integrated to prepare an aptamer sensor with a dual detection mode, enabling rapid and accurate detection of pathogenic bacteria.

[0007] It is undeniable that the design and construction of highly sensitive aptamer sensors are crucial for obtaining high analytical performance and promoting clinical translational applications. However, these sensors mainly focus on the detection and identification of pathogenic bacteria and often neglect the inactivation and elimination of pathogenic bacteria. Therefore, the detection and identification of such highly infectious and pathogenic pathogenic bacteria can only play a preventive role, and there is still a risk of secondary contamination caused by pathogenic bacteria during or after detection, leading to their spread and secondary infection. Especially after the detection of pathogenic bacteria, if not properly handled, the remaining pathogenic bacteria will form stubborn biofilms and may even be transmitted to others through the testers, resulting in large-scale spread and causing serious infectious diseases. In addition, the emergence of drug-resistant bacteria has made many antibacterial drugs difficult to exert their original efficacy, and once infected, it will increase the difficulty of treatment and lead to more serious consequences. Therefore, the real-time elimination of pathogenic bacteria during their detection and identification is expected to solve this problem. However, most of the currently constructed bacterial detection systems usually involve multiple signal probes to output amplified signals, or add a pretreatment module for separating bacteria and a post-treatment module for killing bacteria, undoubtedly increasing the complexity of constructing the detection system, the tediousness of operation steps, and the experimental cost. Therefore, simplifying the construction process of the bacterial detection system and integrating capture and separation, accurate detection, and efficient inactivation into one body is of great significance.

[0008] In view of the above background, the present invention aims to provide a simple "three-in-one" multifunctional aptamer sensing platform ((ZnMn)Fe2O4@Au-Aptamer-Blocker, named ZMFA-AB) for specific capture and ultrasensitive dual-mode detection, and subsequent photothermal clearance of MRSA. Summary of the Invention

[0009] Aiming at the deficiencies of the existing technology, the purpose of the present invention is to provide a superparamagnetic three-in-one aptamer sensing platform, a preparation method and an application, which solve the problems in the existing technology.

[0010] The purpose of the present invention can be achieved by the following technical solutions:

[0011] A preparation method of an aptamer sensing platform, comprising the following steps:

[0012] Dissolve FeCl3·6H2O, MnCl2·4H2O and ZnCl2 in water, add NaOH solution, stir and heat, and wash and dry the obtained product to obtain ZMF NPs;

[0013] Add HAuCl4·3H2O and sodium citrate to water and stir, and then add NaBH4 and stir in the dark to obtain AuNPs;

[0014] Modify the surface of ZMF NPs with PEI-DTC, mix with Au NPs solution and sonicate, wash to obtain ZMF@PEI-DTC / Au NPs; then add NH2OH·HCl and ZMF@PEI-DTC / Au NPs to the growth solution, wash to obtain ZMFA NPs;

[0015] The MRSA-specific nucleic acid aptamer is activated by TCEP solution, mixed with ZMFA NPs, then soaked in NaCl solution, separated to obtain ZMFA-A NPs, and then mixed with Blockers and soaked in NaCl solution, separated to obtain ZMFA-AB NPs.

[0016] Furthermore, the concentrations of FeCl3·6H2O, MnCl2·4H2O and ZnCl2 dissolved in water are 0.1M, 0.02M and 0.025M respectively.

[0017] Furthermore, the reaction formula for preparing ZMF NPs is:

[0018] ZnCl2+MnCl2+FeCl3+NaOH→Zn 0.4 Mn 0.6 Fe2O4.

[0019] Furthermore, the concentration of sodium citrate is 1% w / w; the concentration of NaBH4 is 0.02 M.

[0020] Furthermore, the preparation process of PEI-DTC is as follows: Mix PEI and KOH in methanol under stirring, and degas the mixed solution with argon. Then add CS2. When the color of the solution turns bright yellow, it indicates that PEI-DTC has been formed.

[0021] Furthermore, the process of surface modification of ZMF NPs with PEI-DTC is as follows: Disperse ZMF NPs in methanol, and then mix and stir with PEI-DTC.

[0022] Furthermore, the growth solution is obtained by mixing K2CO3 and HAuCl4·3H2O in water.

[0023] An aptamer sensing platform is prepared by using the above preparation method.

[0024] The application of the above aptamer sensing platform in the preparation of an MRSA detection kit.

[0025] The application of the above aptamer sensing platform in the preparation of a drug for treating MRSA infection.

[0026] Advantages of the present invention:

[0027] 1. The triple-functional aptamer sensing platform provided by the present invention is used for the separation and detection of MRSA. This method has high accuracy, simple operation, short required time, and low cost. This method provides more possibilities for the clinical detection of MRSA.

[0028] 2. The triple-functional aptamer sensing platform provided by the present invention is used for the immediate clearance after the detection of MRSA. This method is convenient to operate, has high bacterial clearance efficiency, and short required time. This method provides more possibilities for the clearance after the detection of MRSA. Description of the Drawings

[0029] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for the description of the embodiments or the prior art. Obviously, for those of ordinary skill in the art, without creative efforts, other drawings can also be obtained based on these drawings.

[0030] Figure 1 It is a schematic diagram of the construction and related application principles of ZMFA-AB of the present invention;

[0031] Figure 2 It is the standard curve of the dual detection mode of ZMFA-AB of the present invention;

[0032] Figure 3 is the sensitivity characterization diagram of ZMFA-AB of the present invention;

[0033] Figure 4 is the specificity characterization diagram of ZMFA-AB of the present invention;

[0034] Figure 5 is the stability characterization diagram of ZMFA-AB of the present invention;

[0035] Figure 6 is the schematic diagram of the clinical test results of ZMFA-AB of the present invention;

[0036] Figure 7 is the CLSM and plate images of ZMFA-AB for near-infrared photothermal elimination of MRSA of the present invention. Detailed implementation manners

[0037] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts belong to the scope of protection of the present invention.

[0038] Example 1

[0039] As Figure 1 shown in A of [], a preparation method of a superparamagnetic triple aptamer sensing platform includes the following steps:

[0040] S1, preparation of ZnMnFe2O4 nanoparticles (ZMF NPs);

[0041] Dissolve FeCl3·6H2O, MnCl2·4H2O and ZnCl2 in distilled water, and their concentrations are 0.1M, 0.02M and 0.025M respectively. Subsequently, transfer 35 mL of the solution to a stainless steel autoclave lined with Teflon. Dissolve 0.88 g of NaOH in 5.0 mL of distilled water, slowly drip it into the autoclave and stir evenly. After reacting at 180 °C for 12 h, cool it to room temperature, and wash it three times with distilled water and ethanol respectively. Finally, place it in an oven at 60 °C and dry it for 12 h to finally obtain ZnMnFe2O4 nanoparticles (ZMF NPs). The reaction formula is as follows:

[0042] ZnCl2 + MnCl2 + FeCl3 + NaOH → Zn 0.4 Mn 0.6 Fe2O4

[0043] S2, Preparation of Au NPs;

[0044] Add 500 μL of HAuCl4·3H2O to 49 mL of ultrapure water. After stirring for 10 min, add 1.5 mL of sodium citrate (1% w / w) and continue stirring. Subsequently, add 500 μL of NaBH4 (0.02 M) solution under strong magnetic stirring; stir in the dark for 12 h to finally obtain Au NPs with small particle sizes.

[0045] S3, Preparation of ZMFA NPs;

[0046] First, modify the surface of ZMF NPs with PEI-DTC: Mix 500 mg of PEI and 750 mg of KOH in 50 mL of methanol under stirring; after degassing the mixed solution with argon for ten seconds, drop 695 μL of CS2 into the solution; when the solution color turns bright yellow, it indicates that PEI-DTC has been formed; then disperse 10 mg of ZMF NPs in methanol at 0.5 mg / mL and mix and stir with the PEI-DTC solution for 1 h to obtain ZMF@PEI-DTC NPs. PEI-DTC can self-assemble on the surface of ZMF NPs; after magnetic separation, the product is washed 5 times and then redispersed in ultrapure water for use.

[0047] Then, mix 10 mg of ZMF@PEI-DTC NPs with 100 mL of Au NPs solution and sonicate for 1 h; due to the formation of bidentate ligands and two chelating sulfur groups, the well-dispersed gold colloids firmly adhere to the surface of ZMF@PEI-DTC NPs, which will serve as nucleation sites to promote the subsequent growth of the gold shell; the resulting product (ZMF@PEI-DTC / Au NPs) is washed three times to remove the excess gold colloid solution (Au NPs solution) and redispersed in ultrapure water.

[0048] Finally, add 10 mL of 0.04 M NH2OH·HCl to 2 mL of ZMF@PEI-DTC / Au NPs and 8 mL of growth solution (16.6 mg of K2CO3 and 118.2 μL of HAuCl4·3H2O mixed in 10 mL of ultrapure water). K2CO3 and HAuCl4 in the growth solution are reduced by NH2OH·HCl to form a dense Au shell layer, obtaining ZMF@Au(ZMFA) NPs; ZMF@Au(ZMFA) NPs are washed three times and stored in ultrapure water for use.

[0049] S4, Preparation of ZMFA-AB NPs

[0050] First, the MRSA-specific nucleic acid aptamer (SH-Aptamer) needs to be activated with TCEP solution: Mix 50 μL of SH-Aptamer at the corresponding concentration with 1 mM TCEP solution;

[0051] After that, after 1 h, mix the activated SH-Aptamer with 1 mg of ZMFA NPs, gently shake in a constant temperature oscillator at 37 °C, then soak in 100 mM NaCl solution for 48 h, and finally wash the product (ZMFA-A NPs) after magnetic separation three times with PBS buffer for later use;

[0052] Then, mix 10 μL of the corresponding concentration of Blockers (blockers of the MRSA-specific nucleic acid aptamer) with 1 mg of ZMFA-A NPs, gently shake in a constant temperature oscillator at 37 °C, then soak in 100 mM NaCl solution for 48 h; finally, the product after magnetic separation is the superparamagnetic triple aptamer sensing platform (i.e., ZMFA-AB NPs), which is washed three times with PBS buffer for later use.

[0053] Among them, the nucleotide sequences of the MRSA nucleic acid aptamer (SH-Aptamer) and Blockers are shown in SEQ ID NO.1 and SEQ ID NO.2 respectively, as shown in Table 1 specifically:

[0054] Table 1 Nucleotide sequences of the MRSA nucleic acid aptamer and Blockers

[0055]

[0056] Note: The underlined sequences in the table are the recognition sequences of the CRISPR-Cas12a system, and the italicized and bolded parts are the corresponding complementary sequences

[0057] Example 2

[0058] In this example, the sensitivity of the ZMFA-AB prepared in Example 1 was analyzed;

[0059] As Figure 1 shown in C, under the optimal experimental conditions, dual-mode detection was performed on MRSA sample solutions at different concentrations (0 - 10 6 CFU / mL), and three parallel concentration experiments were set for each concentration gradient. As Figure 2 shown, where Figure 2 A and C in it respectively represent the particle size and fluorescence intensity values of ZMFA-A / AB for analyzing MRSA standard solutions at different concentrations in Mode-D and Mode-F, Figure 2In it, B and D respectively represent the standard curves for detecting MRSA in Mode-D and Mode-F; in the figure, for Mode-D, the logarithm of the bacterial concentration is on the abscissa, and the D H change value (ΔD H ) is on the ordinate; for Mode-F, the logarithm of the bacterial concentration is on the abscissa, and F / F0 (where F is the fluorescence intensity of the detected bacterial sample and F0 is the fluorescence intensity of the detected blank sample) is on the ordinate to plot the standard curve; it can be seen that when detecting in Mode-D, when the MRSA concentration is between 10-10 4 CFU / mL, there is a good linear relationship between ΔD H and the logarithm of the MRSA concentration, and the linear equation is y = 28.28ln(x) - 20.13 (R 2 = 0.9748); when detecting in Mode-F, when the MRSA concentration is between 10-10 6 CFU / mL, there is a good linear relationship between F / F0 and the logarithm of the MRSA concentration, and the linear equation is y = 1.164x + 1.013 (R 2 = 0.9978). When the MRSA concentration is greater than 10 4 CFU / mL, a good linear relationship is still maintained. Therefore, Mode-F can be used as a reliable supplement to Mode-D to ensure accurate detection even at high concentrations.

[0060] Using the dual-mode of the ZMFA-AB sensing platform to detect 20 negative samples, calculate the average value plus three times the standard deviation to obtain the minimum detection limits of the dual modes respectively; the results are as Figure 3 shown, Figure 3 in which A and B respectively represent the determination of the minimum detection limits of Mode-D and Mode-F modes; in the figure, the LOD of Mode-D is 4.62 CFU / mL, while the LOD of Mode-F is 2.46 CFU / mL. Figure 3 It can be seen from that Mode-F has higher sensitivity and can be used for the detection of trace pathogenic bacteria in complex samples.

[0061] Based on the above data, it shows that the Mode-F mode of ZMFA-AB can not only complement the Mode-D mode and provide more accurate detection performance at high concentrations of MRSA, but also can be used for the detection of more trace pathogenic bacteria in complex samples.

[0062] Example 3

[0063] In this example, the specificity of the ZMFA-AB prepared in Example 1 is analyzed;

[0064] As Figure 1As shown in B of [the relevant figure], in this embodiment, the specificity of ZMFA-AB was investigated by detecting Staphylococcus aureus (CMCC 26003), Escherichia coli (ATCC 25922), Escherichia coli O157:H7 (CMCC 44828 and ATCC 43888), Shigella sonnei (ATCC 25931), Pseudomonas aeruginosa (CMCC 10104), and five Salmonella strains (ATCC 10708, ATCC 13076, ATCC 9150, ATCC 9270, and ATCC 13311). As Figure 4 shown, the concentration of all bacteria, including MRSA, was 1×10 4 CFU / mL, and sterile PBS buffer was used as a blank control; Figure 4 A and B in [the relevant figure] were respectively the specificity investigations of Mode-D and Mode-F. The results showed that both dual-detection modes of ZMFA-AB exhibited good specificity for MRSA.

[0065] Example 4

[0066] In this embodiment, the stability analysis of the ZMFA-AB prepared in Example 1 was carried out;

[0067] In this embodiment, the stability of ZMFA-AB was evaluated by measuring its particle size and fluorescence intensity in pH 7.4 PBS buffer and 10% FBS environment; among them, Figure 5 A and B in [the relevant figure] respectively showed the changes in the particle size and fluorescence intensity of ZMFA-AB in PBS and PBS buffer containing 10% FBS. The results showed that ZMFA-AB had good stability.

[0068] Example 5

[0069] In this embodiment, the ZMFA-AB prepared in Example 1 was applied to the detection of clinical samples;

[0070] The 10 clinical samples used in this embodiment (including 7 MRSA-positive and 3 MRSA-negative samples) were from the Clinical Microbiology Working Group of the Department of Laboratory Medicine, Zhongda Hospital Affiliated to Southeast University. The sample types included blood, pleural effusion, ascites, bronchoalveolar lavage fluid, and catheter drainage fluid. The 10 clinical samples were correctly identified according to the standard operating procedures of the Clinical Microbiology Working Group. Detection was carried out using both Mode-D and Mode-F, and the operating procedures were the same as described above. The results were compared with the plate count results to investigate its clinical diagnostic efficacy, as Figure 6 shown: The results showed that ZMFA-AB could quickly and successfully identify MRSA from clinical samples, and the detection results of both modes were comparable to those of the plate count method, showing excellent clinical diagnostic performance.

[0071] Example 6

[0072] In this embodiment, the ZMFA-AB prepared in Example 1 was applied to in vitro photothermal clearance;

[0073] As Figure 1 shown in D of, in this embodiment, an 808 nm laser exciter (0.75 W / cm 2 ) was used to irradiate the detected bacterial solution, and an infrared thermal imager was used to take heating-up photos and record the temperature change of the solution. Each sample was irradiated for 300 s, and the temperature was recorded and photos were taken every 30 s. The MRSA after photothermal treatment was evenly spread on a blood agar plate and placed in a 37 °C bacterial incubator for 12 h. The antibacterial efficiency of ZMFA-AB was evaluated according to the growth status of the bacteria and the number of colonies. As Figure 7 shown, the results showed that MRSA showed green fluorescence without NIR laser irradiation, indicating that all were live bacteria before NIR treatment; while when MRSA combined with ZMFA-AB was irradiated with NIR laser, MRSA showed red fluorescence, indicating that almost all MRSA were killed at this time. Similarly, a large number of bacterial colonies appeared on the agar plate for MRSA in the control group. While when MRSA combined with ZMFA-AB was irradiated with NIR laser, almost no obvious colonies were observed on the agar plate, indicating that the excellent photothermal conversion performance of ZMFA-AB inactivated and cleared MRSA efficiently, and the bactericidal efficiency against MRSA was as high as 99.9%.

[0074] In the description of this specification, the descriptions with reference to the terms "one embodiment", "example", "specific example", etc. mean that the specific features, structures, materials or characteristics described in connection with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described may be combined in any one or more embodiments or examples in a suitable manner.

[0075] The above shows and describes the basic principles, main features and advantages of the present invention. Those skilled in the art of this industry should understand that the present invention is not limited by the above embodiments. The above embodiments and the descriptions in the specification only illustrate the principles of the present invention. Without departing from the spirit and scope of the present invention, the present invention will have various changes and improvements, and these changes and improvements fall within the scope of the present invention claimed.

Claims

1. A preparation method of an aptamer sensing platform, characterized in that, It includes the following steps: Dissolve FeCl3·6H2O, MnCl2·4H2O and ZnCl2 in water, add NaOH solution, stir and heat. The product obtained from the reaction is washed and dried to obtain ZMF NPs; Add HAuCl4·3H2O and sodium citrate to water and stir. Then add NaBH4 and stir in the dark to obtain Au NPs; Modify the surface of ZMF NPs with PEI-DTC, mix with the Au NPs solution and perform ultrasonic treatment. After washing, ZMF@PEI-DTC / Au NPs are obtained; then add NH2OH·HCl and ZMF@PEI-DTC / Au NPs to the growth solution. After washing, ZMFA NPs are obtained; After the MRSA-specific nucleic acid aptamer is activated by TCEP solution, it is mixed with ZMFA NPs, then soaked in NaCl solution, and ZMFA-A NPs are separated. Then it is mixed with Blockers and soaked in NaCl solution, and ZMFA-AB NPs are separated.

2. The preparation method of an aptamer sensing platform according to claim 1, characterized in that, The concentrations of FeCl3·6H2O, MnCl2·4H2O and ZnCl2 dissolved in water are 0.1M, 0.02M and 0.025M respectively.

3. The preparation method of an aptamer sensing platform according to claim 1 or 2, characterized in that, The reaction formula for preparing ZMF NPs is: ZnCl2 + MnCl2 + FeCl3 + NaOH → Zn 0.4 Mn 0.6 Fe2O4。 4. The preparation method of an aptamer sensing platform according to claim 1, characterized in that, The concentration of the sodium citrate is 1% w / w; the concentration of the NaBH4 is 0.02M.

5. The preparation method of an aptamer sensing platform according to claim 1, wherein The preparation process of the PEI-DTC is: Mix PEI and KOH in methanol under stirring, and degas the mixed solution with argon. Then add CS2. When the color of the solution turns bright yellow, it indicates that PEI-DTC has been formed.

6. The preparation method of an aptamer sensing platform according to claim 1, wherein, The process of modifying the surface of ZMF NPs with PEI-DTC is: Disperse ZMF NPs in methanol, and then mix and stir with PEI-DTC.

7. The preparation method of an aptamer sensing platform according to claim 1, characterized in that, The growth solution is obtained by mixing K2CO3 and HAuCl4·3H2O in water.

8. An aptamer sensing platform, characterized in that It is prepared by using the preparation method described in any one of claims 1-7.

9. Application of the aptamer sensing platform described in claim 8 in the preparation of an MRSA detection kit.

10. Application of the aptamer sensing platform described in claim 8 in the preparation of a drug for treating MRSA infection.

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