Fluorescence signal enhancement method based on quantum dot cancer detection

Abstract

The invention discloses a fluorescence signal enhancement method based on quantum dot cancer detection, which comprises the following steps: step A, modifying the surface of a quantum dot by using an azido modification method, and then entering step B; step B, modifying the surface of the biological probe by utilizing an enzymolysis method, and then entering step C; and step C, connecting the quantum dots and the biological probe by using a bifunctional connecting agent. Aiming at the problem of weak fluorescence intensity caused by low connection efficiency of a biological probe and a quantum dot in a detection process, measures of properly modifying the surfaces of the quantum dot and the biological probe, connecting the quantum dot and the biological probe by using a bifunctional connecting agent and the like are adopted, so that the affinity of the biological probe and the quantum dot is enhanced; more stable and efficient connection can be formed between the biological probe and the quantum dots, and the intensity of a fluorescence signal in the detection process is effectively improved.

Description

Technical Field

[0001] The invention relates to a fluorescence signal enhancement method based on quantum dot cancer detection, and belongs to the technical field of quantum dot fluorescence detection application. Background Art

[0002] Quantum dots are a type of semiconductor material composed of nano-scale particles with unique physical and chemical properties. Their working principle is that when energy is applied to atoms, electrons are excited and transferred to higher energy levels. When electrons return to a lower stable state, the extra energy is emitted in the form of light corresponding to a specific frequency. In cancer detection, quantum dots are connected to biological probes to form quantum dot markers. When a sample containing a target substance is combined with this marker, the quantum dot marker will specifically bind to the target substance to form a quantum dot-target substance complex, which receives and detects the reflected signal after being irradiated with light of a specific wavelength.

[0003] In the existing technology, although quantum dot cancer detection technology is mature, there is still the problem of weak fluorescence signal during the detection process. The main reason is that the connection efficiency between biological probes and quantum dots is low, resulting in a reduction in the number of biomarkers that can be bound and weak fluorescence signals, which affects the efficiency of quantum dot fluorescence detection of cancer. Summary of the invention

[0004] The problem to be solved by the present invention is: a fluorescence signal enhancement method is needed to achieve higher efficiency of quantum dot-bioprobe complex binding with biomarkers and more sensitive fluorescence detection.

[0005] In order to solve the above technical problems, the present invention adopts the following technical solutions: A method for enhancing fluorescence signals based on quantum dot cancer detection comprises the following steps: Step A. Modify the surface of quantum dots using an azido modification method, and then proceed to step B; Step B. Modify the surface of the biological probe by enzymatic hydrolysis, and then proceed to step C; Step C. Connecting quantum dots to biological probes using a bifunctional linker.

[0006] In the aforementioned method for enhancing fluorescence signals based on quantum dot cancer detection, step A comprises the following steps: Step A1: synthesizing quantum dots by an ion precursor reaction method based on a solid fiber membrane; Step A2: pre-treating the surface of the synthesized quantum dots to introduce functional groups such as carboxyl and amino groups to improve the activity of subsequent reactions; Step A3: using click chemistry reaction, the pretreated quantum dots and the compound containing an azide group are dissolved in a reaction solvent in the presence of a catalyst to react, and the reaction progress is monitored by ultraviolet-visible spectroscopy to successfully connect the azide group to the surface of the quantum dots, and then proceed to step B; The aforementioned method for enhancing fluorescence signals based on quantum dot cancer detection, step A1 further comprises the following steps: Step A1-1: preparing a sodium citrate fiber membrane by an electrospinning method, wherein the size of the solid fiber membrane is nanometer or micrometer scale; Step A1-2: placing the prepared solid fiber membrane into a cationic precursor solution, wherein the cationic precursor solution is prepared by dissolving cadmium chloride CdCl2 in ethanol; Step A1-3: Inject anionic precursor S-ODE into the cross-linked solid fiber membrane for reaction. The reaction temperature of anionic and cationic precursors at the solid-liquid interface is 120-160° C., and quantum dots are obtained after the reaction is completed.

[0007] In the aforementioned method for enhancing fluorescence signals based on quantum dot cancer detection, step A2 further comprises the following steps: Step A2-1: First, the quantum dots are uniformly dispersed in a selected solvent to form a stable dispersion, and then a mixed solution of hydrogen peroxide and an organic acid is slowly added dropwise. The reaction temperature is generally controlled between room temperature and 60° C. At the same time, the reaction system is continuously stirred to ensure that the reactants are fully contacted and the oxidation reaction proceeds uniformly; The mixing ratio of hydrogen peroxide and organic acid is determined according to the specific oxidation target and the properties of the quantum dots; Selecting a reducing agent to remove unreacted hydrogen peroxide after the oxidation reaction is completed during the pretreatment of the quantum dot surface; Step A2-2: After the oxidation reaction is completed, the insoluble by-products are treated, including: Wash the quantum dots by centrifugation multiple times, precipitate the quantum dots with ethanol, and then discard the supernatant, repeating the process multiple times until no obvious impurities are detected in the supernatant; or Separation by filtration; Step A2-3: The washed quantum dots are dried at 40°C-60°C by vacuum drying.

[0008] The aforementioned method for enhancing fluorescence signals based on quantum dot cancer detection, in step A3, comprises the following steps: Step A3-1: adding the pretreated reactants, catalyst, and ligand reagent into a reaction container in sequence; Step A3-2: continuously stirring the reaction mixture; Step A3-3: After the reaction is completed, the target product is separated and purified from the reaction mixture.

[0009] The aforementioned method for enhancing fluorescence signals based on quantum dot cancer detection, in step B, comprises the following steps: Step B1: Prepare biological samples; Step B2: Thoroughly remove blood stains and impurities on the surface; Step B3: Cut the sample into pieces and place them in a sterile culture dish, and add enzyme solution into the sterile culture dish; Step B4: constant temperature culture for 2-4 hours; Step B5: Select a sterile filter membrane with appropriate pore size for filtration and centrifugation; Step B6: Collect the precipitate, which is the desired biological probe, and then proceed to step C.

[0010] The aforementioned method for enhancing fluorescence signals based on quantum dot cancer detection comprises, in step B2: Step B2-1: using physiological saline to rinse the biological sample to be made into the biological probe; Step B2-2: Soaking the sample in a buffer solution containing an antimicrobial agent for a set time to kill or inhibit the growth of microorganisms on the surface; The aforementioned method for enhancing fluorescence signals based on quantum dot cancer detection, in step C, comprises the following steps: Step C1: Activation of linker Bis-PEG37-NHS ester; Step C2: The quantum dots react with the linking agent, so that the linking agent is combined with the carboxyl group and the amino group on the surface of the quantum dots; Step C3: connecting the biological probe to the quantum dots via a linker; Step C4: Purify the connected biological probe-quantum dot complex using dialysis.

[0011] The aforementioned method for enhancing fluorescence signals based on quantum dot cancer detection comprises, in step C1: Step C1-1: dissolving the selected bifunctional linker in phosphate buffer; Step C1-2: adding an appropriate amount of activator to the linker solution, wherein the molar ratio of the activator to the linker is 5:1, and continuously stirring the reaction mixture during the reaction to allow the activator to fully contact with the linker; Step C1-3: Add an appropriate amount of activator to the linker solution to chemically change the specific functional groups of the linker.

[0012] The aforementioned method for enhancing fluorescence signals based on quantum dot cancer detection, in step C4, comprises the following steps: Step C4-1: Cut the dialysis membrane into a suitable length, which is determined according to the size of the dialysis container and the sample volume, and soak the dialysis membrane in deionized water for 15-30 minutes; Step C4-2: Place the dialysis bag containing the sample into the dialysis container, so that the dialysis bag is completely immersed in the dialysis solution, wherein the dialysis solution is NaCl, and cover the dialysis container with a plastic wrap or a lid; Step C4-3: After dialysis is completed, remove the dialysis bag from the dialysis container, wipe the dialysate on the outside of the dialysis bag with a clean paper towel or gauze, then open the dialysis clamp at one end of the dialysis bag and transfer the dialyzed sample to a clean beaker.

[0013] Beneficial effects achieved by the present invention: A fluorescence signal enhancement method based on quantum dot cancer detection of the present invention, by modifying the surface of quantum dots with azide groups and modifying the surface of biological probes by enzymatic hydrolysis, can reduce the quenching effect of surface defects on fluorescence, expose specific recognition sites, improve the accuracy of fluorescence detection, and reduce the error of fluorescence detection to a certain extent.

[0014] In order to address the problem of weak fluorescence signals in current quantum dot cancer detection technology, the surface of quantum dots is specifically modified, the surface of biological probes is treated appropriately, and bifunctional linkers are used to build a more stable and efficient connection system between quantum dots and biological probes, thereby significantly enhancing the intensity of fluorescence signals. This greatly improves the sensitivity and accuracy of quantum dots in cancer detection applications, provides more powerful technical support for early diagnosis of cancer, disease monitoring, and related research, and promotes the development of quantum dot fluorescence detection technology in the field of cancer diagnosis and treatment in a more accurate and efficient direction, which has important scientific significance and broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 This is a flow chart of the surface modification of quantum dots by azide groups in Example 1 of the present invention; Figure 2 This is a flow chart of the enzymatic modification of the biological probe surface in Example 1 of the present invention; Figure 3 This is a flow chart of connecting quantum dots and biological probes using a bifunctional linker in Example 1 of the present invention. DETAILED DESCRIPTION

[0016] The embodiments of the present invention are described in detail above with reference to the accompanying drawings, but the present invention is not limited to the above embodiments, and various changes can be made within the knowledge scope of ordinary technicians in this field without departing from the purpose of the present invention. Example 1

[0017] This embodiment provides a method for enhancing fluorescence signals based on quantum dot cancer detection, which includes modifying the surfaces of quantum dots and biological probes and connecting the two using a bifunctional linker, including the following steps: Step A. Modify the surface of quantum dots using an azido modification method, and then proceed to step B; Step B. Modify the surface of the biological probe by enzymatic hydrolysis, and then proceed to step C; Step C. Connecting quantum dots to biological probes using a bifunctional linker.

[0018] The step A comprises the following steps: Step A1: synthesizing quantum dots by an ion precursor reaction method based on a solid fiber membrane; Step A1 further comprises the following steps: Step A1-1: preparing a sodium citrate fiber membrane by an electrospinning method, wherein the size of the solid fiber membrane is nanometer or micrometer scale; Step A1-2: placing the prepared solid fiber membrane into a cationic precursor solution, wherein the cationic precursor solution is prepared by dissolving cadmium chloride (CdCl2) in ethanol; Step A1-3: Inject the anionic precursor S-ODE (S-oleylamine, Dodecylamine, Ethanolamine) into the cross-linked solid fiber membrane for reaction. The reaction temperature of the anionic and cationic precursors at the solid-liquid interface is 120-160°C. After the reaction is completed, quantum dots are obtained.

[0019] Step A2: pre-treating the surface of the synthesized quantum dots to introduce functional groups such as carboxyl and amino groups to improve the activity of subsequent reactions; Step A2 further comprises the following steps: Step A2-1: First, the quantum dots are uniformly dispersed in a selected solvent (such as toluene) to form a stable dispersion, and then a mixed solution of hydrogen peroxide and an organic acid is slowly added dropwise. The reaction temperature is generally controlled between room temperature and 60°C, and the reaction system is continuously stirred to allow the reactants to fully contact and allow the oxidation reaction to proceed uniformly. The mixing ratio of hydrogen peroxide and the organic acid is determined according to the specific oxidation target and the properties of the quantum dots. For example, when the quantum dot type is cadmium selenide CdSe and formic acid is used as the organic acid, the volume ratio of hydrogen peroxide to formic acid is between 1:0.5 and 1:1. Selecting a reducing agent such as sodium sulfite to remove unreacted hydrogen peroxide after the oxidation reaction is completed during the pretreatment of the quantum dot surface; Step A2-2: After the oxidation reaction is completed, the insoluble by-products are treated, including: Wash the quantum dots by centrifugation multiple times, precipitate the quantum dots with ethanol, and then discard the supernatant, repeating the process multiple times until no obvious impurities are detected in the supernatant; or The separation was performed by filtration. A PTFE filter membrane with a pore size of 0.45 microns was used, and the pretreated quantum dot solution was slowly poured into a vacuum filter. The vacuum degree was maintained between -0.05 and -0.08 MPa. 20 ml of the sample was filtered through a 0.45 micron pore size filter membrane for 10-20 minutes. After the filtration was completed, the filtrate was collected; Step A2-3: The washed quantum dots are dried at 40°C-60°C by vacuum drying.

[0020] Step A3: using click chemistry reaction, the pretreated quantum dots and the compound containing an azide group are dissolved in a reaction solvent in the presence of a catalyst to react, and the reaction progress is monitored by ultraviolet-visible spectroscopy to successfully connect the azide group to the surface of the quantum dots, and then proceed to step B; The reaction solvent is generally a polar aprotic solvent, such as N,N-dimethylformamide, dimethyl sulfoxide, and the reaction temperature is carried out at room temperature; The catalyst is a complex of CuBr and PMDETA; In step A3, the following steps are included: Step A3-1: Add the pretreated reactants, catalysts, ligands and other reagents into the reaction container in sequence. During the addition process, care should be taken to avoid mutual contamination and violent reactions between the reagents; The ligand refers to a class of compounds that form a complex with the catalyst to adjust the activity and selectivity of the catalyst, and here refers to PMDETA; Step A3-2: continuously stirring the reaction mixture; Step A3-3: After the reaction is completed, the target product is separated and purified from the reaction mixture.

[0021] In step B, the following steps are included: Step B1: Prepare biological samples; Step B2: Thoroughly remove blood stains and possible impurities on the surface; In step B2, it includes: Step B2-1: using physiological saline to rinse the biological sample to be made into the biological probe; Step B2-2: Soak the sample in a buffer solution containing an appropriate amount of an antimicrobial agent for a set time, such as 10-20 minutes, to kill or inhibit the growth of microorganisms on the surface; the antimicrobial agent is a penicillin-streptomycin mixture with a mixing ratio of 1:1; Step B3: Cut the sample into pieces and place them in a sterile culture dish, and add enzyme solution into the sterile culture dish; In step B3, trypsin or collagenase is used as the enzyme solution to decompose the extracellular matrix, and 1-5 ml of the corresponding enzyme solution is added per gram of sample, while ensuring that the enzyme solution is evenly covered on the shredded sample; Step B4: constant temperature culture for 2-4 hours; Step B5: Select a sterile filter membrane with appropriate pore size for filtration and centrifugation; Step B6: Collect the precipitate, which is the desired biological probe, and then proceed to step C.

[0022] In step C, the following steps are included: Step C1: Activation of the linker Bis-PEG37-NHS ester (bis-active ester thirty-seven polyethylene glycol); In step C1, it includes: Step C1-1: dissolving the selected bifunctional linker in phosphate buffered saline (PBS) to allow the linker to be stable and active; Step C1-2: adding an appropriate amount of activator to the linker solution, wherein the molar ratio of the activator to the linker is 5:1, and continuously stirring the reaction mixture during the reaction to allow the activator to fully contact with the linker; Step C1-3: Add an appropriate amount of activator to the linker solution to chemically change the specific functional group of the linker. The nucleophilic part of the activator will attack the carbonyl carbon in the active ester group, causing the ester bond to break and form an unstable tetrahedral intermediate. Then, the intermediate will dissociate, and the oxygen atom originally connected to the carbonyl group will leave with a negative charge, while the nucleophilic part of the activator will be connected to the rest of the linker molecule through the newly formed amide bond. The active ester group is the key specific functional group in the di-active ester trisepagonal polyethylene glycol, which can more effectively enable the linker to connect biological probes and quantum dots.

[0023] Step C2: The quantum dots react with the linking agent, so that the linking agent is combined with the carboxyl group and the amino group on the surface of the quantum dots; Step C3: connecting the biological probe to the quantum dots via a linker; Step C4: Purify the connected biological probe-quantum dot complex using dialysis.

[0024] In step C4, the following steps are included: Step C4-1: Cut the dialysis membrane into a suitable length, which is determined according to the size of the dialysis container and the sample volume, and soak the dialysis membrane in deionized water for 15-30 minutes. After soaking, check whether the dialysis membrane has any damage or holes; Step C4-2: Place the dialysis bag containing the sample into the dialysis container, so that the dialysis bag is completely immersed in the dialysis solution, and the dialysis solution is selected from NaCl. Meanwhile, cover the dialysis container with a plastic wrap or a lid to prevent the evaporation of the dialysis solution and contamination by external impurities; Step C4-3: After dialysis is completed, remove the dialysis bag from the dialysis container, wipe the dialysate on the outside of the dialysis bag with a clean paper towel or gauze, then open the dialysis clamp at one end of the dialysis bag and transfer the dialyzed sample to a clean beaker to minimize the sample remaining in the dialysis bag.

[0025] The comparison scheme and experimental data of the present invention and the traditional method are listed below:

[0026] Under the same experimental conditions, a conventional quantum dot-bioprobe complex that has not been modified and connected by the method of the present invention (control group) and a quantum dot-bioprobe complex prepared by the method of the present invention (experimental group) were subjected to binding experiments with target substances (cancer-related biomarkers).

[0027] Control group: An equal amount of control group complex was added to a biomarker solution of a certain concentration. After sufficient reaction, the ratio of the amount of complex bound to the biomarker to the total amount of complex added was measured, and the binding efficiency of the control group was found to be approximately 30%.

[0028] Experimental group: Under the same conditions, after the experimental group complex reacted with the biomarker, the binding efficiency was measured to be about 60%.

[0029] Conclusion: The binding efficiency of the experimental group was significantly higher than that of the control group, indicating that the method of the present invention can make the quantum dot-bioprobe complex bind to the biomarker more efficiently.

[0030] The quantum dot-bioprobe complexes in the control group and the experimental group were used to detect a series of target substances (cancer-related biomarkers) with different concentration gradients, and the lowest concentration of the target substance that produced a detectable fluorescence signal was recorded.

[0031] Control group: During the detection process, when the concentration of the target substance dropped to a certain level, the fluorescence signal intensity was too weak to be accurately detected, and the lowest detectable concentration was 10 ng / mL.

[0032] Experimental group: After the quantum dot complex prepared by the present invention is combined with the target substance, a lower concentration of the target substance can be detected, and the minimum detectable concentration reaches 5 ng / mL.

[0033] Conclusion: The experimental group was able to detect lower concentrations of target substances, indicating that the fluorescence detection method of the present invention is more sensitive.

[0034] For target substance samples with known concentrations (including low, medium and high concentrations), repeated tests were performed using the detection methods of the control group and the experimental group (each group of samples was tested 10 times), and the percentage deviation between the test results and the true value was calculated.

[0035] Control group: When testing low concentration target substances, the deviation percentage range is ±15%; the deviation is ±10% at medium concentration; the deviation is ±8% at high concentration. The average deviation is (15%+10%+8%) / 3≈11%.

[0036] Experimental group: The detection deviation percentage range of low concentration target substance is ±8%; the deviation is ±5% at medium concentration; the deviation is ±3% at high concentration. The average deviation is (8%+5%+3%) / 3≈5.3%.

[0037] Conclusion: The average deviation of the experimental group was significantly smaller than that of the control group, indicating that the detection method of the present invention has higher accuracy.

[0038] Through the data comparison of the above theoretical analysis, it can be preliminarily demonstrated that the present invention has the advantages of higher efficiency in combining with biomarkers, more sensitive fluorescence detection, and higher detection accuracy in the field of quantum dot cancer detection. Example 2

[0039] In step A2-1, when the quantum dot type is InP and formic acid is selected as the organic acid, the volume ratio of hydrogen peroxide to formic acid can be between 1:0.8 and 2:1.

[0040] Other technical features are the same as those of Example 1.

[0041] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the technical principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A fluorescence signal enhancement method based on quantum dot cancer detection, characterized in that: The following steps are involved: Step A. Modify the surface of quantum dots using an azido modification method, and then proceed to step B; Step B. Modify the surface of the biological probe by enzymatic hydrolysis, and then proceed to step C; Step C. Connecting quantum dots to biological probes using a bifunctional linker.

2. A method for enhancing fluorescence signals based on quantum dot cancer detection according to claim 1, characterized in that: The step A comprises the following steps: Step A1: synthesizing quantum dots by an ion precursor reaction method based on a solid fiber membrane; Step A2: pre-treating the surface of the synthesized quantum dots to introduce functional groups such as carboxyl and amino groups to improve the activity of subsequent reactions; Step A3: Using click chemistry, the pretreated quantum dots and the azide-containing compound are dissolved in a reaction solvent in the presence of a catalyst for reaction, and the reaction progress is monitored by UV-visible spectroscopy to successfully connect the azide group to the surface of the quantum dots, and then proceed to step B.

3. The method for enhancing fluorescence signals based on quantum dot cancer detection according to claim 2, characterized in that: Step A1 further comprises the following steps: Step A1-1: preparing a sodium citrate fiber membrane by an electrospinning method, wherein the size of the solid fiber membrane is nanometer or micrometer scale; Step A1-2: placing the prepared solid fiber membrane into a cationic precursor solution, wherein the cationic precursor solution is prepared by dissolving cadmium chloride CdCl2 in ethanol; Step A1-3: Inject anionic precursor S-ODE into the cross-linked solid fiber membrane for reaction. The reaction temperature of anionic and cationic precursors at the solid-liquid interface is 120-160° C., and quantum dots are obtained after the reaction is completed.

4. The method for enhancing fluorescence signals based on quantum dot cancer detection according to claim 2, characterized in that: Step A2 further comprises the following steps: Step A2-1: First, the quantum dots are uniformly dispersed in a selected solvent to form a stable dispersion, and then a mixed solution of hydrogen peroxide and an organic acid is slowly added dropwise. The reaction temperature is generally controlled between room temperature and 60° C. At the same time, the reaction system is continuously stirred to ensure that the reactants are fully contacted and the oxidation reaction proceeds uniformly; The mixing ratio of hydrogen peroxide and organic acid is determined according to the specific oxidation target and the properties of the quantum dots; Selecting a reducing agent to remove unreacted hydrogen peroxide after the oxidation reaction is completed during the pretreatment of the quantum dot surface; Step A2-2: After the oxidation reaction is completed, the insoluble by-products are treated, including: Wash the quantum dots by centrifugation multiple times, precipitate the quantum dots with ethanol, and then discard the supernatant, repeating the process multiple times until no obvious impurities are detected in the supernatant; or Separation by filtration; Step A2-3: The washed quantum dots are dried at 40°C-60°C by vacuum drying.

5. The method for enhancing fluorescence signals based on quantum dot cancer detection according to claim 2, characterized in that: In step A3, the following steps are included: Step A3-1: adding the pretreated reactants, catalyst, and ligand reagent into a reaction container in sequence; Step A3-2: continuously stirring the reaction mixture; Step A3-3: After the reaction is completed, the target product is separated and purified from the reaction mixture.

6. The method for enhancing fluorescence signals based on quantum dot cancer detection according to claim 1, characterized in that: In step B, the following steps are included: Step B1: Prepare biological samples; Step B2: Thoroughly remove blood stains and impurities on the surface; Step B3: Cut the sample into pieces and place them in a sterile culture dish, and add enzyme solution into the sterile culture dish; Step B4: constant temperature culture for 2-4 hours; Step B5: Select a sterile filter membrane with appropriate pore size for filtration and centrifugation; Step B6: Collect the precipitate, which is the desired biological probe, and then proceed to step C.

7. The method for enhancing fluorescence signals based on quantum dot cancer detection according to claim 6, characterized in that: In step B2, it includes: Step B2-1: using physiological saline to rinse the biological sample to be made into the biological probe; Step B2-2: Soak the sample in a buffer solution containing an antimicrobial agent for a set time to kill or inhibit the growth of microorganisms on the surface.

8. The method for enhancing fluorescence signals based on quantum dot cancer detection according to claim 1, characterized in that: In step C, the following steps are included: Step C1: Activation of linker Bis-PEG37-NHS ester; Step C2: The quantum dots react with the linking agent, so that the linking agent is combined with the carboxyl group and the amino group on the surface of the quantum dots; Step C3: connecting the biological probe to the quantum dots via a linker; Step C4: Purify the connected biological probe-quantum dot complex using dialysis.

9. The method for enhancing fluorescence signals based on quantum dot cancer detection according to claim 8, characterized in that: In step C1, it includes: Step C1-1: dissolving the selected bifunctional linker in phosphate buffer; Step C1-2: adding an appropriate amount of activator to the linker solution, wherein the molar ratio of the activator to the linker is 5:1, and continuously stirring the reaction mixture during the reaction to allow the activator to fully contact with the linker; Step C1-3: Add an appropriate amount of activator to the linker solution to chemically change the specific functional groups of the linker.

10. The method for enhancing fluorescence signals based on quantum dot cancer detection according to claim 8, characterized in that: In step C4, the following steps are included: Step C4-1: Cut the dialysis membrane into a suitable length, which is determined according to the size of the dialysis container and the sample volume, and soak the dialysis membrane in deionized water for 15-30 minutes; Step C4-2: Place the dialysis bag containing the sample into the dialysis container, so that the dialysis bag is completely immersed in the dialysis solution, wherein the dialysis solution is NaCl, and cover the dialysis container with a plastic wrap or a lid; Step C4-3: After dialysis is completed, remove the dialysis bag from the dialysis container, wipe the dialysate on the outside of the dialysis bag with a clean paper towel or gauze, then open the dialysis clamp at one end of the dialysis bag and transfer the dialyzed sample to a clean beaker.

Images