Preparation method and application of Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor

Abstract

The present invention provides a preparation method and application of a Co‑TBAPy / graphene oxide quantum dot photoelectrochemical biosensor; the method comprises the following steps: S1, preparation of Co‑TBAPy material: S2, preparation of Co‑TBAPy / graphene oxide quantum dot composite material: S3, preparation of Co‑TBAPy / graphene oxide quantum dot photoelectrochemical biosensor; the present invention also relates to the aforementioned Co‑TBAPy / graphene oxide quantum dot photoelectrochemical biosensor for the detection of serotonin or Aβ‑40 amyloid protein. The Co‑TBAPy of the present invention is composited with graphene oxide quantum dots to form an effective carrier transport system, thereby improving the photoelectrochemical performance. At the same time, the MOFs structure of Co‑TBAPy provides an effective anchoring site for DNA aptamers, and finally constructs an efficient and stable Co‑TBAPy / graphene oxide quantum dot photoelectrochemical biosensor. By fixing different DNA aptamer probes, ultra-sensitive and highly specific detection of serotonin and Aβ‑40 amyloid protein can be achieved respectively.

Description

Technical Field

[0001] The present invention belongs to the field of biosensor technology, and in particular relates to a preparation method and application of a Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor. Background Art

[0002] In recent years, as people pay more and more attention to life and health issues, ultra-sensitive biomarker analysis is of great significance for the early diagnosis and treatment of diseases.

[0003] Semiconductor photoelectrochemical (PEC) biosensors utilize the photoelectric properties of semiconductors to detect the concentration of the substance to be measured and the analysis of biochemical process parameters related to the photogenerated current or photogenerated voltage. They have the advantages of low background noise, fast response and high sensitivity, and have therefore attracted widespread attention. During the PEC detection process, the directional movement of photogenerated carriers generates photocurrent, and the magnitude of the photocurrent response is linearly related to the concentration of the detection target, thereby establishing a quantitative detection system. In order to improve the detection specificity of PEC sensors, it is an important challenge to stably anchor the biological probe on the surface of the semiconductor photoelectrode. An effective signal amplification strategy is another core factor to further improve the sensitivity of PEC sensors. Summary of the invention

[0004] The purpose of the present invention is to provide a preparation method and application of a Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor.

[0005] The present invention is achieved through the following technical solutions:

[0006] The present invention relates to a method for preparing a Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor, comprising the following steps:

[0007] S1, Preparation of Co-TBAPy materials:

[0008] S1-1, dissolve appropriate amounts of Co(NO3)·6H2O and H4TBAPy in dioxane, DMF and H2O, transfer the homogeneous solution to a reactor for hydrothermal growth;

[0009] S1-2, after the solution is cooled to room temperature, centrifuge and wash;

[0010] S1-3, vacuum drying to obtain Co-TBAPy powder;

[0011] S2, Preparation of Co-TBAPy / graphene oxide quantum dots composite materials:

[0012] S2-1, graphene oxide quantum dots and Co-TBAPy powder are dispersed in DMF to obtain a uniform solution, and the DMF solution of graphene oxide quantum dots is slowly added to the DMF solution of Co-TBAPy during stirring, and stirred for 30 minutes;

[0013] S2-2, transferring the solution into a reactor for hydrothermal growth;

[0014] S2-3, after the solution is cooled to room temperature, it is centrifuged for washing;

[0015] S2-4, vacuum drying to obtain a Co-TBAPy / graphene oxide quantum dot composite material;

[0016] S3, Preparation of Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor:

[0017] S3-1, 5 mg of Co-TBAPy / graphene oxide quantum dot composite material, water, ethanol and naphthol were mixed and ultrasonicated for 30 minutes to obtain a uniform dispersion;

[0018] S3-2, take 50uL of the dispersed liquid and apply it on a 0.5*0.5cm 2 The FTO conductive glass surface was dried at 60 °C to form the Co-TBAPy / graphene oxide quantum dot photoanode;

[0019] S3-3, coating the aptamer probe on the surface of the Co-TBAPy / graphene oxide quantum dot photoanode and drying;

[0020] S3-4, rinse the film surface with ultrapure water to obtain the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor.

[0021] Preferably, in step S1-1, the molar concentration ratio of Co(NO3)·6H2O to H4TBAPy is 0.1:0.2, in mM, and the volume ratio of dioxane:DMF:H2O is 10:20:10, in mL.

[0022] Preferably, in step S1-1, the reaction temperature of the hydrothermal growth is 120-160° C., and the reaction time is 24-72 hours.

[0023] Preferably, in step S1-2, the centrifugal washing is performed by DMF washing for 5 times.

[0024] Preferably, in step S2-1, the mass concentration ratio of the DMF solution of Co-TBAPy to the DMF solution of graphene oxide quantum dots is 5-10:0.05-0.1, in units of mg / mL.

[0025] Preferably, in step S2-2, the reaction temperature of the hydrothermal growth is 100-120° C., and the reaction time is 10-12 hours.

[0026] Preferably, in step S2-3, the centrifugal washing is performed by DMF washing for 5 times.

[0027] Preferably, in step S3-1, the volume ratio of water, ethanol and naphthol is 500:450:50, and the unit is uL.

[0028] Preferably, in step S3-3, the aptamer probe is a DNA aptamer probe of serotonin or Aβ-40 amyloid protein.

[0029] The present invention also relates to the application of the aforementioned Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor, and the aforementioned prepared Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor is used for the detection of serotonin or Aβ-40 amyloid protein.

[0030] The present invention has the following advantages:

[0031] (1) The Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor prepared by the method involved in the present invention combines Co-TBAPy with graphene oxide quantum dots and utilizes the synergistic advantages of the two to achieve a comprehensive improvement in the performance of the photoelectrochemical biosensor, while also improving the linear detection area, sensitivity and specificity of the semiconductor photoelectrochemical sensor.

[0032] (2) The method involved in the present invention uses Co-TBAPy as a semiconductor photoelectrode material, which has good photoelectric response ability. At the same time, its MOFs structure has a large specific surface area, which can provide more binding sites for the fixation of specific probes, thereby broadening the linear detection area of ​​the sensor.

[0033] (3) The method involved in the present invention successfully loads graphene oxide quantum dots on Co-TBAPy. The modification of quantum dots not only improves the separation efficiency of photogenerated carriers, but also serves as an efficient reaction site, greatly improving the photocurrent signal intensity of the photoanode, thereby making the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor have higher sensitivity.

[0034] (4) The Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor prepared by the method of the present invention has the advantages of high detection sensitivity and excellent specificity, and has broad application potential and expansion capabilities in the fields of early screening of depression markers and early screening of Alzheimer's disease markers. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 : SEM and TEM images of Co-TBAPy / graphene oxide quantum dot composite materials; wherein a is the SEM image of Co-TBAPy powder, b is the TEM image of Co-TBAPy powder; c is the TEM image of Co-TBAPy / graphene oxide quantum dot composite materials; d is the HRTEM image of Co-TBAPy / graphene oxide quantum dot composite materials;

[0036] Figure 2 The linear curves of the detection of different substances by immobilizing different DNA aptamer probes on the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor, where a is the serotonin curve and b is the Aβ-40 amyloid protein curve;

[0037] Figure 3 This is a diagram showing the specificity evaluation of the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor for serotonin. DETAILED DESCRIPTION

[0038] The present invention is described in detail below in conjunction with specific embodiments. It should be noted that the following embodiments are only further descriptions of the present invention, but the protection scope of the present invention is not limited to the following embodiments.

[0039] Example

[0040] This embodiment relates to a method for preparing a Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor, comprising the following steps:

[0041] S1. Preparation of Co-TBAPy Materials

[0042] ① Dissolve 0.1mM Co(NO3)·6H2O and 0.2mM H4TBAPy in a mixed solution of 10mL dioxane, 20mL DMF and 10mL H2O, transfer the homogeneous solution to a reactor, and perform hydrothermal growth at 120-160℃ for 24-72 hours;

[0043] ②After the solution cools to room temperature, wash it with DMF by centrifugation for 5 times;

[0044] ③ Vacuum drying overnight to obtain Co-TBAPy powder;

[0045] S2. Preparation of Co-TBAPy / graphene oxide quantum dots composite materials

[0046] ① Disperse the graphene oxide quantum dots and the Co-TBAPy powder prepared in step S1 in DMF to obtain a Co-TBAPy DMF solution and a graphene oxide quantum dot DMF solution, respectively. Slowly add 0.05-0.1 mg / mL of the graphene oxide quantum dot DMF solution to the 5-10 mg / mL Co-TBAPy DMF solution during stirring, and stir for 30 minutes;

[0047] ② Transfer the solution to the reactor and perform hydrothermal growth at 100-120°C for 10-12 hours;

[0048] ③After the solution cools to room temperature, wash it with DMF by centrifugation for 5 times;

[0049] ④ Vacuum drying overnight to obtain the Co-TBAPy / graphene oxide quantum dot composite material;

[0050] S3. Preparation of Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor

[0051] ① Mix 5 mg of the Co-TBAPy / graphene oxide quantum dot composite material prepared in step S2 with 500 uL of water, 450 uL of ethanol and 50 uL of naphthol, and ultrasonicate for 30 minutes to obtain a uniform dispersion;

[0052] ② Take 50uL of the dispersed liquid and apply it on a 0.5*0.5cm 2 The FTO conductive glass surface was dried at 60 °C to form the Co-TBAPy / graphene oxide quantum dot photoanode;

[0053] ③ Prepare 3.6uM serotonin and Aβ-40 amyloid protein aptamer probes respectively, apply the aptamer probes on the surface of Co-TBAPy / graphene oxide quantum dot photoanode, and dry at 37°C for 2 hours;

[0054] ④ Rinse the film surface with ultrapure water to obtain the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor.

[0055] 1. Test the products obtained during the preparation process

[0056] like Figure 1 As shown, Figure 1 a shows the SEM image of Co-TBAPy. Figure 1 b shows the TEM image of Co-TBAPy, the length and width of Co-TBAPy are 20 μm and 5 μm, respectively, which provides sites for the attachment of DNA probes. Figure 1 c is the TEM image of Co-TBAPy / graphene oxide quantum dots composite material; Figure 1d is the HRTEM image of Co-TBAPy / graphene oxide quantum dot composite material. Figure 1 c and Figure 1 In d, graphene oxide quantum dots can be seen attached to the surface of Co-TBAPy, and the 0.35 nm lattice fringes appearing in the HRTEM image correspond to the (002) crystal plane of graphene oxide, proving the successful loading of graphene oxide quantum dots.

[0057] The photoelectrode prepared from the Co-TBAPy / graphene oxide quantum dot composite material was fixed with the DNA probe to obtain the photoelectrochemical biosensor. The detected solution of different concentrations was dripped on the surface of the sensor. After incubation for 15 minutes, the electrode surface was fully cleaned and dried. The working electrode was used as the negative electrode, and the Ag / AgCl electrode was used as the reference electrode. The photoelectrochemical biosensor was tested at a light intensity of 100 mW / cm 2 Under the irradiation of simulated sunlight, the detection characteristics of the sensor are tested by the current-time curve.

[0058] Figure 2 The linear curve of the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor for the detection of serotonin and Aβ-40 amyloid protein, core markers of neurodegenerative diseases. Figure 2 a is the linear curve of 5-HT. It can be seen that the photocurrent signal increases with the increase of the target substance concentration. The detection line width is 10 -13 ~10 -10 ug / L, detection limit 0.45×10 -13 ug / L (S / N=3). By replacing the aptamer probe on the sensor surface, a sensor with specific response to Aβ-40 amyloid protein was constructed. Figure 2 b It can be concluded that the photocurrent signal decreases with the increase of the target substance concentration, and the detection line width is 10 -13 ~10 -9 ug / L, detection limit 0.38×10 -13 ug / L(S / N=3).

[0059] 2. Another important indicator of the sensor: detection specificity

[0060] The DNA aptamer of serotonin was fixed to the surface of Co-TBAPy / graphene oxide quantum dot photoelectrode to prepare a series of sensors with specific response to serotonin, and interference evaluation experiments were carried out on their specificity.

[0061] like Figure 3 As shown in FIG. 1 , the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor of the present invention has a very obvious light response signal to serotonin, while for the attached Figure 3The responses of other interference items (dopamine, adrenaline, bovine serum albumin, glucose, and NaCl) were weak, indicating that the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor of the present invention has a strong detection specificity for serotonin after fixing the serotonin aptamer.

[0062] It can be seen that the Co-TBAPy of the present invention is compounded with graphene oxide quantum dots to form an effective carrier transport system, which improves the photoelectrochemical performance. At the same time, the MOFs structure of Co-TBAPy provides an effective anchoring site for the DNA aptamer, and finally constructs an efficient and stable Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor. By fixing different DNA aptamer probes, ultra-sensitive and highly specific detection of serotonin and Aβ-40 amyloid protein can be achieved respectively.

[0063] In summary, the present invention successfully synthesizes a new type of MOFs material and prepares it into a photoelectrochemical biosensor, which can efficiently anchor DNA aptamer probes. The modification of graphene oxide quantum dots not only improves the separation efficiency of photogenerated carriers, thereby greatly improving the detection sensitivity and specificity of the sensor, but also, through the fixation and modification of different DNA aptamer probes, ultra-sensitive and highly specific detection of neurological disease markers serotonin and Aβ-40 amyloid protein can be achieved respectively. As a typical MOFs material, theoretical calculations show that Co-TBAPy has very obvious hydrophilic properties, and the porous framework structure has a good fixation effect on biological probes. The composite structure formed by graphene oxide quantum dots and Co-TBAPy has excellent carrier transport performance, which can effectively improve the separation efficiency of photogenerated carriers, thereby significantly improving the current density of the photoelectrode.

[0064] The above describes the specific embodiments of the present invention. It should be understood that the present invention is not limited to the above specific embodiments, and those skilled in the art may make various modifications or variations within the scope of the claims, which will not affect the essence of the present invention.

Claims

1. A method for preparing a Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor, characterized in that: The following steps are involved: S1, Preparation of Co-TBAPy materials: S1-1, dissolve appropriate amounts of Co(NO3)·6H2O and H4TBAPy in dioxane, DMF and H2O, transfer the homogeneous solution to a reactor for hydrothermal growth; S1-2, after the solution is cooled to room temperature, centrifuge and wash; S1-3, vacuum drying to obtain Co-TBAPy powder; S2, Preparation of Co-TBAPy / graphene oxide quantum dots composite materials: S2-1, graphene oxide quantum dots and Co-TBAPy powder are dispersed in DMF to obtain a uniform solution, and the DMF solution of graphene oxide quantum dots is slowly added to the DMF solution of Co-TBAPy during stirring, and stirred for 30 minutes; S2-2, transferring the solution into a reactor for hydrothermal growth; S2-3, after the solution is cooled to room temperature, centrifuge and wash; S2-4, vacuum drying to obtain a Co-TBAPy / graphene oxide quantum dot composite material; S3, Preparation of Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor: S3-1, 5 mg of Co-TBAPy / graphene oxide quantum dot composite material, water, ethanol and naphthol were mixed and ultrasonicated for 30 minutes to obtain a uniform dispersion; S3-2, take 50 μL of the dispersion droplet and apply it on a 0.5×0.5 cm 2 The FTO conductive glass surface was dried at 60 °C to form the Co-TBAPy / graphene oxide quantum dot photoanode; S3-3, coating the aptamer probe on the surface of the Co-TBAPy / graphene oxide quantum dot photoanode and drying; S3-4, rinse the film surface with ultrapure water to obtain the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor.

2. The method for preparing the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor according to claim 1, characterized in that: In step S1-1, the molar concentration ratio of Co(NO3)·6H2O to H4TBAPy is 0.1:0.2, in mM, and the volume ratio of dioxane:DMF:H2O is 10:20:10, in mL.

3. The method for preparing the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor according to claim 1, characterized in that: In step S1-1, the reaction temperature of the hydrothermal growth is 120-160°C, and the reaction time is 24-72 hours.

4. The method for preparing the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor according to claim 1, characterized in that: In step S1-2, the centrifugal washing is performed by DMF washing for 5 times.

5. The method for preparing the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor according to claim 1, characterized in that: In step S2-1, the mass concentration ratio of the DMF solution of Co-TBAPy to the DMF solution of graphene oxide quantum dots is 5-10:0.05-0.1, in units of mg / mL.

6. The method for preparing the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor according to claim 1, characterized in that: In step S2-2, the reaction temperature of the hydrothermal growth is 100-120°C, and the reaction time is 10-12 hours.

7. The method for preparing the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor according to claim 1, characterized in that: In step S2-3, the centrifugal washing is performed by DMF washing for 5 times.

8. The method for preparing the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor according to claim 1, characterized in that: In step S3-1, the volume ratio of water, ethanol and naphthol is 500:450:50, and the unit is μL.

9. The method for preparing the Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor according to claim 1, characterized in that: In step S3-3, the aptamer probe is a DNA aptamer probe of serotonin or Aβ-40 amyloid protein.

10. An application of a Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor, characterized in that: The Co-TBAPy / graphene oxide quantum dot photoelectrochemical biosensor prepared according to claim 1 is used for the detection of serotonin or Aβ-40 amyloid protein.

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