CN117169201A - Dual-path composite chromogenic sensor based on alkaline phosphatase induction and application thereof - Google Patents
Dual-path composite chromogenic sensor based on alkaline phosphatase induction and application thereof Download PDFInfo
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Classifications
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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Abstract
The invention discloses an alkaline phosphatase-induced dual-path composite chromogenic sensor and application thereof, wherein a substrate L-ascorbic acid-2 phosphate magnesium salt hydrate can be hydrolyzed to generate ascorbic acid with reducibility in the presence of alkaline phosphatase, so that Ag is further prepared + Reduction to Ag 0 Depositing and growing silver shell layers on the surfaces of the gold nanoparticles, and changing the appearance of the gold nanoparticles along with the displacement of the plasmon resonance peak of the local surface and the change of color; at the same time, ascorbic acid reacts with s-nitrosoglutathione to produce NO and with NO-specific recognition molecules, with a concomitant change in color from white to pink. The two paths are subjected to compound color development, so that the human eye recognition is enhancedThe color change is bright, simple and efficient, the color development is rapid, the sensitivity is high, the early detection of the bone metastasis of the prostate cancer can be realized, and the application prospect in the aspect of daily detection of the cancer disease is wide.
Description
Technical Field
The invention relates to the technical field of nanomaterials and colorimetry detection, relates to a dual-path composite chromogenic sensor based on alkaline phosphatase induction and application thereof, and in particular relates to a composite colorimetric sensor with adjustable color generated by double colorimetric signal response and application research thereof based on alkaline phosphatase-induced gold nanoparticle surface silver deposition growth and NO specific recognition dye molecule Rd chromogenic recognition.
Background
Prostate cancer is often atypical in terms of its own characteristics, and therefore a significant number of patients have undergone distant metastasis at the time of initial visit, with bone metastasis occurring in nearly 70% of patients. Bone metastasis can lead to bone-related events, such as spinal cord compression, pathological fractures, hypercalcemia, etc., that lead to bone metabolic disorders, not only reducing the quality of life of the patient, but also affecting the selection of their treatment regimen, thereby increasing the economic burden and mortality of the patient. Currently, bone scanning (ECT) is the most commonly used and effective method for detecting bone metastasis clinically, and can detect lesions earlier, 3-6 months or even longer than X-ray film. However, the detection means requiring a complicated precision instrument or a complicated operation procedure is not suitable for daily use, and thus there is no convenient method for realizing daily monitoring of bone metastasis. In addition, under the current large background of global health threat, medical equipment and resources are in shortage and cannot be used in a whole-scale manner, so that the portable, low-price and simple-operation detection equipment is indispensable for people living in various areas, the requirements of the portable and low-price product equipment for fields such as sensitive clinical diagnosis, on-site environment monitoring and food inspection are increased increasingly, and the detection efficiency of the fields can be greatly improved through rapid in-situ detection of targets. To do this, each device must be designed as simply as possible. In this respect, colorimetric sensors that provide naked eye detection without using any specific instrument have great potential in future development, and various portable and simple detection devices have been continuously developed, especially the use of various self-test kits, which provide a guarantee for daily monitoring of health conditions.
To date, much effort has been devoted to developing various colorimetric chemical/biological sensors, however, most colorimetric sensors have low visual analysis accuracy due to their single color development, and quantitative detection of targets still requires spectrometer assistance, which increases costs to some extent and limits portability of the detection system. Therefore, colorimetric chemical/biological sensors for semi-quantitative naked eye in situ detection are becoming a research direction and are highly desirable. It is well known that human eyes are not very sensitive to changes in optical density, but they are very sensitive to changes in color with significant shifts in spectral peaks. Therefore, if the manufactured sensor shows a lot of color variety, the visual detection accuracy will be significantly improved. For example, pH paper is the most common multicolor sensor, and can quantify hydrogen ion concentration by a distinct color change, producing a multicolor change of purple, green, yellow, orange, pink, and red, among others. These vivid colors greatly improve the accuracy of visual inspection, so that the pH can be estimated without using a pH meter. Therefore, the color compound adjustable sensor is developed, the human eye identifiable degree is improved, the color which is distinguished more quickly is provided for judging whether the index is abnormal or not, the disease diagnosis efficiency can be greatly improved, the effective treatment measures are guided, and the death rate can be greatly reduced.
Disclosure of Invention
The first object of the invention is to provide a dual-path composite chromogenic sensor based on alkaline phosphatase induction, which is based on alkaline phosphatase ALP-induced gold nanoparticle surface silver deposition growth and chromogenic recognition of NO specific recognition dye molecules Rd, is simple and efficient, quick in chromogenic, high in sensitivity and high in color recognition degree, and has great development potential in research of disease detection technology.
An alkaline phosphatase-induced dual-path composite chromogenic sensor is prepared by the following method:
step (1), rhodamine B and l-phenylenediamine are dissolved in methylene dichloride, triethylamine and 1-propyl phosphoric acid cyclic anhydride DMF solution are added for reaction, and nitric oxide recognition molecules Rd are obtained after purification;
step (2), the L-ascorbic acid-2 phosphate magnesium salt hydrate AAP and silver nitrate AgNO 3 And gold nanoparticles are mixed, and the gold nanoparticles obtained in the step (1) are added into the mixed solutionAnd obtaining Rd and GSNO, and obtaining the alkaline phosphatase-induced dual-path composite chromogenic sensor.
Preferably, in the step (1), the mass ratio of rhodamine B to l-phenylenediamine is 1: (0.5-1).
Preferably, the volume ratio of triethylamine to 1-propyl phosphoric acid cyclic anhydride DMF in the step (1) is (1-3): (2-5).
Preferably, the reaction temperature in the step (1) is 30-70 ℃ and the reaction time is 10-15 h.
Preferably, the gold nanoparticles in step (2) include, but are not limited to, gold nanobipyramids, gold nanospheres, or gold nanorods; more preferably gold nano bipyramids, and the longitudinal ultraviolet absorption peak is between 700 and 800 nm.
Preferably, the concentration of L-ascorbic acid-2 phosphate magnesium salt hydrate AAP in step (2) is 3 to 7mM.
Preferably, the concentration of s-nitrosoglutathione GSNO in step (2) is 20 to 40mM.
The principle of the invention is as follows:
in the presence of alkaline phosphatase, the substrate L-ascorbic acid-2 phosphate magnesium salt hydrate AAP can be hydrolyzed to produce ascorbic acid AA with reducibility, and then Ag is hydrolyzed + Reduction to Ag 0 Depositing and growing silver shell layers on the surfaces of the gold nanoparticles, and changing the appearance of the gold nanoparticles along with the displacement of the plasmon resonance peak of the local surface and the change of color; meanwhile, AA reacts with s-nitrosoglutathione GSNO in the presence of alkaline phosphatase to produce NO and with NO-specific recognition molecule Rd, with a concomitant change in color from white to pink.
Preferably, in the step (1), the Au NBPs solution is synthesized by classical seed-mediated synthesis, such as chloroauric acid, sodium Citrate (SC) and AgNO 3 AA, sodium borohydride, hydrochloric acid and Cetyl Trimethyl Ammonium Bromide (CTAB) are used as raw materials to prepare the catalyst, and the catalyst is specifically as follows:
(1) preparation of gold core: adding 7-10 mL deionized water into a cleaned glass bottle, and then adding 0.1-0.4 mL SC solution (5-15 mM) and 0.1-0.3 mLHAuCl under the stirring condition of rotating speed of 500-900 rpm 4 Solutions (5-15 mM) followed byNaBH prepared by rapidly adding 0.1-0.3 mL of ice water under stirring at 1200rpm 4 (5-15 mM) stirring for 2-5 min, stopping stirring, and standing at 30 ℃ for more than 2h for later use;
(2) preparing a growth solution: 300-500 mL CTAB (0.05-0.15M) and 2-6 mLAgNO are stirred at 500-900 rpm 3 (5～15mM)，15～25mLHAuCl 4 (5-15 mM), 3-5 mLAA (0.05-0.15M) and 5-10 mLHCl (0.5-1.5M) were added sequentially to a 500mL clean three-necked glass bottle. And finally, adding 3-7 mL of gold nano-cores in the step (1) into the growth solution in the step (2), stirring for 30-60 s, stopping stirring, and standing at 30 ℃ for more than 6 hours to obtain the unpurified Au NBPs material.
(3) Purifying: the unpurified 24-40 mLAuNBPs were washed with water (7000-10000 rpm, 5-15 min) and the resulting precipitate was dispersed with 18-30 mL cetyltrimethylammonium chloride (CTAC) (0.05-0.15M) solution. Then sequentially adding 5-10 mLAgNO 3 (5-10 mM) and 2.5-5 mLAA (0.05-0.15M), and placing the mixture into a baking oven at 50-100 ℃ for standing for more than 4 hours to obtain the deposition bottom of the gold cone coated silver shell composite material (Au NBPs@Ag). After removing the supernatant, the Au NBPs@Ag precipitate at the bottom is uniformly dispersed by 12-20 mL of deionized water. Subsequently, 1 to 2mL of ammonia water (content of 30%) and 0.5 to 1.5. 1.5mLH are added 2 O 2 The solution (0.5-1.5M) is uniformly mixed and kept stand for 20-40 min, so as to achieve the purpose of completely etching the silver shell layer in the Au NBPs@Ag and obtaining the single dispersion AuNBPs. Finally, the etched solution is centrifugally washed (7000-10000 rpm, 3-7 min) twice, and the obtained purified AuNBPs precipitate is dispersed with deionized water for later use.
A second object of the present invention is to provide the use of the dual-path composite chromogenic sensor for detecting ALP as described above for monitoring prostate cancer bone metastasis. ALP expression in serum of normal persons and prostate cancer non-bone metastasis patients ranges from 40 to 150U/L, and if the ALP concentration exceeds 150U/L, bone metastasis risks are possible, so that early daily monitoring of bone metastasis can be achieved by detecting the ALP concentration.
Preferably, the application process is specifically as follows:
mixing a certain amount of alkaline phosphatase-induced dual-path compound chromogenic sensor with a sample to be detected, observing the color change of the mixed solution or detecting the change of the ultraviolet visible absorption spectrum of the mixed solution after the reaction is completed, so that the qualitative and quantitative analysis of alkaline phosphatase can be realized, and the degree of prostate cancer deterioration can be estimated through an alkaline phosphatase index.
The third object of the invention is to provide the application of the alkaline phosphatase-induced dual-path composite chromogenic sensor in preparing a prostate cancer bone metastasis detection kit.
Compared with the prior art, the invention has the beneficial effects that:
the invention develops a simple, efficient, low-equipment-requirement and low-cost method for realizing early daily monitoring of prostate cancer bone metastasis, and utilizes the specific reaction of ALP and AAP to generate AA, trigger a bicolor reaction and generate compound color change. The silver deposition growth reaction on the surface of the Au NBPs is stable, the color change is clear, and the specific recognition response of Rd dye molecules to NO is quick. The compound color which is favorable for the rapid distinction of human eyes is generated, the recognition degree and the detection sensitivity of human eyes are improved, the visual detection of tumor disease markers (alkaline phosphatase) can be realized, the color development is rapid, the color resolution is high, and the method has wide application prospect in the evaluation of the deterioration degree of cancer diseases.
The method is different from the traditional colorimetric method, can realize more accurate naked eye disease index ALP monitoring, and early warning of disease risk in time. The effect is realized based on a gold nanoparticle-Rd dye molecular recognition response dual-path composite colorimetric strategy, and the easily-distinguished composite color can improve the human eye recognition degree, so that the effective distinction can be easily realized at the critical index of normal and abnormal.
Drawings
FIG. 1 is a TEM image of gold nano bipyramids prepared in example 1, with a scale of 50nm.
FIG. 2 is an ultraviolet spectrum of gold nano bipyramid prepared in example 1, with a longitudinal LSPR peak at 721 nm.
FIG. 3 is a TEM characterization of the morphology change of Au NBPs in example 1 in the presence of different concentrations of ALP, scale 50nm; wherein (A) the ALP concentration is 0U/L; (B) ALP concentration is 10U/L; (C) ALP concentration is 20U/L; the ALP concentration of (D) was 60U/L.
Fig. 4 shows the color change from white to pink before and after the reaction of the dye molecule Rd with NO.
FIG. 5 is a graph showing the optical effects of different concentrations of ALP on the silver deposition growth on the surface of Au NBPs and the recombination of dye Rd recognition responses, wherein the longitudinal LSPR peak of Au NBPs is blue-shifted and the ultraviolet visible absorption peak appears after Rd reaction at 550nm in the presence of different concentrations of ALP.
FIG. 6 shows the color change after compounding the recognition response of dye Rd, which affects the silver deposition growth on the surface of Au NBPs by ALP with different concentrations: brown-reddish brown-blackish green-cyan-dark green.
Detailed Description
The invention is further elucidated below in connection with the drawings and the examples. It is to be understood that these examples are for illustration of the invention only and are not intended to limit the scope of the invention.
Example 1
(1) Synthesis and purification of gold nano bipyramid
(1) Preparation of gold nanocores 9.625mL deionized water was added to a 20mL glass bottle followed by 0.25mL SC solution (0.01M) and 0.125mL LHAuCl with stirring at 700rpm 4 Solution (0.01M) followed by rapid addition of 0.15mL of ice water prepared NaBH with stirring at 1200rpm 4 (0.01M) solution, stirring for 2min, stopping stirring, and standing at 30deg.C for more than 2 hr.
(2) Preparing a growth solution: 400mL of CTAB (0.1M), 4mL of AgNO were stirred at 700rpm 3 (0.01M)，20mLHAuCl 4 (0.01M), 3.2mLAA (0.1M) and 8mLHCl (1M) were added sequentially to a 500mL three-necked glass bottle. And finally, adding 5mL of gold nano-cores in the step (1) into the growth solution in the step (2), stirring for 30 seconds, stopping stirring, and standing at 30 ℃ for more than 6 hours to obtain an unpurified AuNBPs material.
(3) Purifying: the 36mLAuNBPs were washed twice with centrifugation (9000 rpm,10 min) and the resulting precipitate was dispersed with 30mL of CTAC (0.1M) solution. Then sequentially adding 6mLAgNO 3 (7.5 mM) and 3mLAA (0.1M) and mixingAnd (3) uniformly placing the mixture into a 65 ℃ oven for standing for more than 4 hours to obtain the deposition bottom of the gold cone coated silver shell composite material (Au NBPs@Ag). After removal of the supernatant, the bottom Au NBPs@Ag precipitate was uniformly dispersed with 20mL deionized water. Subsequently, 1.2mL of aqueous ammonia (30% content) and 1mLH were added 2 O 2 The solution (1M) is uniformly mixed and kept stand for 30min, so that the purpose of completely etching the silver shell layer in the AuNBPs@Ag to obtain single Au NBPs is achieved. Finally, the etched solution was washed twice by centrifugation (8000 rpm,5 min), and the resulting purified Au NBPs precipitate was dispersed to 2mL with deionized water for use.
The particle size of the obtained gold nano bipyramid is 70-100 nm, a TEM image is shown in figure 1, an ultraviolet spectrum is shown in figure 2, and the longitudinal LSPR peak of the gold nano bipyramid is 721 nm.
(2) Synthesis of nitric oxide recognition molecule Rd
1g of rhodamine B and 0.86g l-phenylenediamine were dissolved in 30mL of methylene chloride. 1.5mL of triethylamine and 3mL of 1-propylphosphoric acid cyclic anhydride DMF solution (50 wt.%) were added to the above mixture, followed by reaction at 50℃for 12h. The crude product was chromatographed on silica gel (CH) 2 Cl 2 Meoh=30:1, v/v) to give NO recognition molecule Rd, structural formula:
(3) Compound colorimetric detection of ALP
Sequentially adding 108 mu LH into a 0.5mL centrifuge tube 2 O (containing 10mM CTAC), 10. Mu. LAAP (5 mM), 10. Mu. LALP (0,1,5,10,20,30,40,60,80,100U/L), 50. Mu. LAgNO 3 (10 mM) and 50. Mu.L of Au NBPs (20-fold concentrated), then stirred well on a mixer and incubated in a 37℃water bath for 5min. Subsequently, 2. Mu.L of GSNO (30 mM) and 50. Mu. LRd dye probe (1 mg/mL) were removed and added one by one, stirred well and placed in a 37℃water bath for 5min. Immediately after removal, 5. Mu.LNa was added one by one 3 VO 4 (0.1M) and uniformly mixed, and inhibiting the ALP hydrolysis termination reaction. Finally, the color of the solution is photographed and the ultraviolet spectrum is measured.
FIG. 3 is a TEM characterization of silver deposition growth on surfaces of different concentrations of ALP-induced Au NBPs, where the concentration of ALP in A is 0nM; ALP concentration in B is 10U/L; ALP concentration in C is 20U/L; the ALP concentration in D is 60U/L, and the variation of the morphology size of the Au NBPs can cause the shift of the longitudinal LSPR peak of the Au NBPs, so that the color variation is generated.
FIG. 4 shows the color change of Rd dye molecules before and after recognizing NO, from white to pink.
FIG. 5 is a graph showing the effect of longitudinal LSPR peak shift on Au NBPs in the presence of different concentrations of ALP.
Fig. 6 is a graph showing the color effect of different concentrations of ALP on the silver deposition growth on the surface of Au NBPs. As can be seen from the graph, in the presence of ALP with the concentration range of 0-100U/L, au NBPs continuously grow, the longitudinal LSPR peak gradually shifts blue and partially shifts red, the color of the solution changes to brown, reddish brown, blackish green, cyan and dark green, and the color is clear and distinguishable, so that the method can realize the visual detection of the ALP concentration index in the prostate cancer bone metastasis disease.
Example 2
(1) Preparation of gold nanosphere solution
Trisodium citrate (SC, 10mL,33 mM) was added to 140mL deionized water boiling at 1000rpm (oil bath temperature was maintained at 137 ℃) for 40min. Fresh HAuCl is then added 4 (1 mL,25 mM) was injected rapidly with no significant change in color. After 60 seconds tris (hydroxymethyl) aminomethane (TB, 5mL, 0.1M) was added and the solution color changed rapidly from colorless to pale pink, kept boiling and stirred at 1000rpm for 15min. Subsequently, the temperature is reduced to 100 ℃, HAuCl is added 4 (1 mL,25 mM) was rapidly injected, the color of the solution changed to a reddish wine at about 1min, and the temperature was maintained for 20min. Finally, HAuCl 4 (1 mL,25 mM) was rapidly injected and maintained at the temperature for 20min, and then naturally cooled to obtain a gold nanosphere solution.
(2) Synthesis of nitric oxide recognition molecule Rd
1g of rhodamine B and 0.86g l-phenylenediamine were dissolved in 30mL of methylene chloride. 1.5mL of triethylamine and 3mL of 1-propylphosphoric acid cyclic anhydride DMF solution (50 wt.%) were added to the above mixture, followed by reaction at 50℃for 12h. The crude product was chromatographed on silica gel (CH) 2 Cl 2 Meoh=30:1, v/v) purification to giveNO recognition molecule Rd.
(3) Compound colorimetric detection of ALP
Sequentially adding 108 mu LH into a 0.5mL centrifuge tube 2 O (containing 10mM CTAC), 10. Mu. LAAP (5 mM), 10. Mu. LALP (0,1,5,10,20,30,40,60,80,100U/L), 50. Mu. LAgNO 3 (10 mM) and 50. Mu.L of Au NPs (concentrated), then stirred well on a mixer and incubated in a 37℃water bath for 5min. Subsequently, 2. Mu.L of GSNO (30 mM) and 50. Mu. LRd dye probe (1 mg/mL) were removed and added one by one, stirred well and placed in a 37℃water bath for 5min. Immediately after removal, 5. Mu.LNa was added one by one 3 VO 4 (0.1M) and mixed well. Finally, the color of the solution is photographed and the ultraviolet spectrum is measured.
Example 3
(1) Preparation of gold nanorod solution
(1) Sequentially adding 0.25mL of 0.01M HAuCl 4 NaBH formulated in solution and 0.60mL of 0.01M ice water 4 Sequentially adding the solutions into 9.75mL 0.10M CTAB solution, and then vigorously stirring at 1200rpm for 2 minutes to obtain nano gold core solution, and standing at room temperature for at least 120 minutes for later use.
(2) First, 4mL of 0.01M HAuCl was sequentially added to the reactor at a rotation speed of 700rpm and a water bath temperature of 28 DEG C 4 Solution, 0.8mL of 0.01M AgNO 3 Solution, 0.64ml of 0.1m ascorbic acid solution and 1.6ml of 1m HCl solution were added to 80ml of 0.1m CTAB solution; subsequently, 70. Mu.L of the nano gold core solution prepared in (1) was added to the mixed solution, and after 60 minutes, the color of the solution was gradually changed to red, and after standing overnight in a water bath at 28℃the solution was centrifuged three times at 10000rpm and concentrated to a gold nanorod solution having a concentration of about 1.4 nM.
(2) Synthesis of nitric oxide recognition molecule Rd
1g of rhodamine B and 0.86g l-phenylenediamine were dissolved in 30mL of methylene chloride. 1.5mL of triethylamine and 3mL of 1-propylphosphoric acid cyclic anhydride DMF solution (50 wt.%) were added to the above mixture, followed by reaction at 50℃for 12h. The crude product was chromatographed on silica gel (CH) 2 Cl 2 MeOH=30:1, v/v) to give the NO recognition molecule Rd.
(3) Compound colorimetric detection of ALP
Sequentially adding 108 mu LH into a 0.5mL centrifuge tube 2 O (containing 10mM CTAC), 10. Mu. LAAP (5 mM), 10. Mu. LALP (0,1,5,10,20,30,40,60,80,100U/L), 50. Mu. LAgNO 3 (10 mM) and 50. Mu.L of Au NRs (concentrated), then stirred well on a mixer and incubated in a 37℃water bath for 5min. Subsequently, 2. Mu.L of GSNO (30 mM) and 50. Mu. LRd dye probe (1 mg/mL) were removed and added one by one, stirred well and placed in a 37℃water bath for 5min. Immediately after removal, 5. Mu.LNa was added one by one 3 VO 4 (0.1M) and mixed well. Finally, the color of the solution is photographed and the ultraviolet spectrum is measured.
Application example 1
Whole blood samples from two patients were tested: first, the whole blood sample obtained was centrifuged at 3000rpm for 5 minutes, and the supernatant was collected. The above steps were then repeated 3 times to obtain serum samples. The ALP content in serum was then measured using a composite chromogenic sensor, the results of which are shown in Table 1, and compared with the detection method of commercial kit (Shanghai Shake Co., ltd.) to verify the reliability of the sensor. The study has been approved by the first institutional ethics committee of the Ningbo city (No. 013A-01,2022);
the detection scheme is as follows: sequentially adding 108 mu LH into a 0.5mL centrifuge tube 2 O (containing 10mM CTAC), 10. Mu.L AAP (5 mM), 10. Mu.L serum test solution, 50. Mu.LAgNO 3 (10 mM) and 50. Mu. LAu NRs (concentrated), then stirred well on a mixer and incubated in a 37℃water bath for 5min. Subsequently, 2. Mu.L of GSNO (30 mM) and 50. Mu. LRd dye probe (1 mg/mL) were removed and added one by one, stirred well and placed in a 37℃water bath for 5min. Immediately after removal, 5. Mu.LNa was added one by one 3 VO 4 (0.1M) and mixed well. Finally, the color of the solution is photographed and the ultraviolet spectrum is measured.
Serum samples were tested using a commercial kit (Shanghai Shake industries, ltd.) and compared with the method of the present invention, and the test results are shown in Table 1.
Table 1 comparison of commercial kit detection method with ALP marker detection results of the inventive method
The data in the table show that the effect of the method for detecting the content of the tumor marker is similar to the detection result of a commercial kit detection method (the detection precision of two samples is 100.2% and 100.7% respectively), the feasibility and the accuracy of the method for detecting the tumor marker are proved, and the effective differentiation of diseases and the screening of patients can be realized through visual color comparison, so that the method has wide application prospect in the aspects of diagnosis and evaluation of cancer diseases.
While the foregoing embodiments have been described in detail in connection with the embodiments of the invention, it should be understood that the foregoing embodiments are merely illustrative of the invention and are not intended to limit the invention, and any modifications, additions, substitutions and the like made within the principles of the invention are intended to be included within the scope of the invention.
Claims (10)
1. The dual-path composite chromogenic sensor based on alkaline phosphatase induction is characterized in that the dual-path composite chromogenic sensor is prepared by adopting the following method:
step (1), rhodamine B and l-phenylenediamine are dissolved in methylene dichloride, triethylamine and 1-propyl phosphoric acid cyclic anhydride DMF solution are added for reaction, and nitric oxide recognition molecules Rd are obtained after purification;
step (2), the L-ascorbic acid-2 phosphate magnesium salt hydrate AAP and silver nitrate AgNO 3 And (3) mixing gold nanoparticles, and adding the nitric oxide recognition molecules Rd and s-nitrosoglutathione GSNO obtained in the step (1) into the mixed solution to obtain the alkaline phosphatase-induced dual-path composite chromogenic sensor.
2. The alkaline phosphatase-induced dual-path-based composite colorimetric sensor according to claim 1, wherein the mass ratio of rhodamine B to l-phenylenediamine in step (1) is 1: (0.5-1).
3. The alkaline phosphatase-induced dual-path-based composite colorimetric sensor according to claim 1, wherein the volume ratio of triethylamine to 1-propylphosphoric acid cyclic anhydride DMF in step (1) is (1-3): (2-5).
4. The alkaline phosphatase-induced dual-path-based composite colorimetric sensor according to claim 1, wherein the reaction temperature in the step (1) is 30 to 70 ℃ and the reaction time is 10 to 15min.
5. The alkaline phosphatase-induced dual-path-based composite colorimetric sensor according to claim 1, wherein the gold nanoparticle in step (2) is one of gold bipyramids, gold nanorods, or gold nanospheres.
6. The alkaline phosphatase-induced dual-path-based composite colorimetric sensor according to claim 1, wherein the concentration of L-ascorbic acid-2 phosphate magnesium salt hydrate AAP in step (2) is 3 to 7mM.
7. The alkaline phosphatase-induced dual-path-based composite colorimetric sensor according to claim 1, wherein the concentration of s-nitrosoglutathione GSNO in step (2) is 20 to 40mM.
8. Use of an alkaline phosphatase-induced dual-path based composite chromogenic sensor as claimed in any one of claims 1 to 7 for monitoring prostate cancer bone metastasis.
9. The use according to claim 8, characterized in that the application process is specifically as follows:
mixing a certain amount of alkaline phosphatase-induced dual-path compound chromogenic sensor with a sample to be detected, observing the color change of the mixed solution or detecting the change of the ultraviolet visible absorption spectrum of the mixed solution after the reaction is completed, so that the qualitative and quantitative analysis of alkaline phosphatase can be realized, and the degree of prostate cancer deterioration can be estimated through an alkaline phosphatase index.
10. Use of an alkaline phosphatase-induced dual-path-based composite chromogenic sensor according to any one of claims 1 to 7 for the preparation of a kit for detecting bone metastasis of prostate cancer.
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