CN112213290A - Novel coronavirus detection method and system based on luminescent quantum dot material - Google Patents
Novel coronavirus detection method and system based on luminescent quantum dot material Download PDFInfo
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Abstract
The invention discloses a novel coronavirus detection method and a novel coronavirus detection system based on a luminescent quantum dot material, wherein the method comprises the following steps of: adding CdTe quantum dots into a PBS buffer solution, and uniformly mixing to obtain a first mixed solution; adding a spike protein specific antibody into the first mixed solution, uniformly mixing to obtain a second mixed solution, and testing the PL spectral intensity of the second mixed solution; dripping a sample to be tested into the second mixed solution, uniformly mixing to obtain a third mixed solution, and testing the PL spectral intensity of the third mixed solution; and if the PL spectral intensity of the third mixed solution is smaller than the PL spectral intensity of the second mixed solution, judging that the sample to be detected contains the novel coronavirus. The response time of the detection method provided by the invention is less than 2 minutes, and the response of a sample to be detected can be realized relatively quickly; the requirement on a detection instrument is low; the detection limit of the novel coronavirus reaches 1 x 10‑16mg/ml。
Description
Technical Field
The invention relates to the field of virus detection, in particular to a novel coronavirus detection method and system based on a luminescent quantum dot material.
Background
Rapid and accurate identification or identification of potential pathogens is critical for disease control, patient treatment and prevention of the prevalence of emerging infectious diseases (e.g., severe respiratory syndrome coronavirus, SARS-COV).
Currently, the technologies for clinical virus detection are mainly reverse transcription polymerase chain reaction (RT-PCR), POC technology based on antibody detection and breast Computed Tomography (CT). However, although these technical means can realize relatively accurate detection of viruses, they have high requirements on professional instruments and equipment, and meanwhile, the detection time of these technical means is long, and it is difficult to realize rapid detection of a sample to be detected.
Accordingly, the prior art is yet to be improved and developed.
Disclosure of Invention
In view of the above-mentioned shortcomings of the prior art, the present invention aims to provide a method and a system for detecting a novel coronavirus based on a luminescent quantum dot material, which aims to solve the problems of long detection time, high cost and poor detection sensitivity of the prior art for the novel coronavirus.
The technical scheme of the invention is as follows:
a novel coronavirus detection method based on a luminescent quantum dot material comprises the following steps:
adding CdTe quantum dots into a PBS buffer solution, and uniformly mixing to obtain a first mixed solution;
adding a spike protein specific antibody into the first mixed solution, uniformly mixing to obtain a second mixed solution, and testing the PL spectral intensity of the second mixed solution;
dripping a sample to be tested into the second mixed solution, uniformly mixing to obtain a third mixed solution, and testing the PL spectral intensity of the third mixed solution;
and if the PL spectral intensity of the third mixed solution is smaller than the PL spectral intensity of the second mixed solution, judging that the sample to be detected contains the novel coronavirus.
The novel coronavirus detection method based on the luminescent quantum dot material further comprises the following steps of:
adding the target spike proteins with different calibrated gradient concentrations into the second mixed solution, uniformly mixing to obtain a fourth mixed solution, and testing the PL spectral intensity of the fourth mixed solution to form a mapping table corresponding to the target spike proteins with different calibrated gradient concentrations and the PL spectral intensity;
and calculating to obtain the novel coronavirus concentration of the sample to be detected according to the mapping table and the PL spectral intensity of the third mixed solution.
The novel coronavirus detection method based on the luminescent quantum dot material is characterized in that the calibrated different gradient concentrations of the target spike protein are 1.0 x 10-4-1.0*10-16mg/ml。
The novel coronavirus detection method based on the luminescent quantum dot material comprises the steps of adding a spike protein specific antibody into the first mixed solution, uniformly mixing to obtain a second mixed solution, and testing the PL spectral intensity of the second mixed solution, wherein the steps comprise:
adding a spike protein specific antibody into the first mixed solution, oscillating for 15-30min, and uniformly mixing to obtain a second mixed solution;
the PL spectral intensity of the second mixture was measured and recorded using a PL spectrometer at an excitation wavelength of 360 nm.
The novel coronavirus detection method based on the luminescent quantum dot material comprises the following steps of dropwise adding a sample to be detected into the second mixed solution, uniformly mixing to obtain a third mixed solution, and testing the PL spectral intensity of the third mixed solution, wherein the steps comprise:
adding a sample to be detected into the second mixed solution, oscillating for 15-30min, and uniformly mixing to obtain a third mixed solution;
the PL spectral intensity of the third mixture was measured using a PL spectrometer at an excitation wavelength of 360nm and recorded.
A novel coronavirus detection system based on a luminescent quantum dot material comprises a fluorescence immunosensor and a PL spectrometer, wherein the fluorescence immunosensor comprises CdTe quantum dots and a specific antibody CSab bound to the surfaces of the CdTe quantum dots, and the specific antibody CSab is used for specifically binding with spike proteins on the surfaces of novel coronaviruses.
Has the advantages that: the invention provides a novel coronavirus detection method and system based on luminescent quantum dot materials, the response time of the detection method is less than 2 minutes, the response of a sample to be detected can be realized relatively quickly, and the method has the advantages of rapidness, simplicity and convenience compared with technologies such as PCR (polymerase chain reaction) which need longer response time and complicated sample processing steps; meanwhile, the detection method has lower requirements on detection instruments, can realize the detection of the sample to be detected by a common fluorescence spectrometer, is simpler, can be stably dispersed in a liquid phase, saves the complicated pretreatment step of the sample to be detected, and further provides convenience for the detection and storage of the sample; the detection method has a limit of detection (LOD) of 1 × 10 for the novel coronavirus-16mg/ml。
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FIG. 1 is a flow chart of a preferred embodiment of the method for detecting coronavirus based on luminescent quantum dot material.
FIG. 2 is a schematic diagram of a novel coronavirus detection method based on a luminescent quantum dot material.
FIG. 3 is a flow chart of another method for detecting coronavirus based on luminescent quantum dot material according to a preferred embodiment of the present invention.
FIG. 4 is a PL spectrum measured after addition of different gradient concentrations of the target spike protein.
Detailed Description
The invention provides a novel coronavirus detection method and system based on a luminescent quantum dot material, and the invention is further described in detail below in order to make the purpose, technical scheme and effect of the invention clearer and clearer. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.
Referring to fig. 1, fig. 1 is a diagram illustrating a novel coronavirus detection method based on a luminescent quantum dot material according to the present invention, which includes the following steps:
s10, adding CdTe quantum dots into PBS buffer solution, and uniformly mixing to obtain a first mixed solution;
s20, adding a spike protein specific antibody into the first mixed solution, uniformly mixing to obtain a second mixed solution, and testing the PL spectral intensity of the second mixed solution;
s30, dripping a sample to be tested into the second mixed solution, uniformly mixing to obtain a third mixed solution, and testing the PL spectral intensity of the third mixed solution;
and S40, if the PL spectral intensity of the third mixed solution is smaller than that of the second mixed solution, judging that the sample to be detected contains the novel coronavirus.
As shown in fig. 2, this embodiment provides a novel coronavirus detection method using CdTe quantum dots as fluorescent labels, where the CdTe quantum dots have a large number of negatively charged carboxyl groups on their surfaces, and after a spike protein specific antibody (CSab) is added to a first mixed solution containing CdTe quantum dots, the specific antibody of the spike protein has a positive surface electrical property, so that the CdTe quantum dots and the spike protein specific antibody can be combined together through an electrostatic acting force to form a fluorescent immunosensor for specifically identifying the spike protein on the surface of the novel coronavirus; the fluorescence immunosensor can emit light under the irradiation of laser with specific wavelength due to the existence of the CdTe quantum dot, the light-emitting signal can be detected and recorded through a PL spectrometer, after a sample to be detected is added into a second mixed solution, if the sample to be detected contains spike protein, a specific antibody CSab in the fluorescence immunosensor can be specifically combined with the spike protein to form a CdTe quantum dot-CSab-spike protein system, so that the fluorescence intensity of the CdTe quantum dot is influenced, the CdTe quantum dot is quenched, the intensity is gradually reduced along with the concentration of the added spike protein, and data shows that the PL spectral intensity is linearly related to the concentration of the added spike protein in the quenching process, and when the concentration of the added spike protein is high enough, the color of the third mixed solution can be obviously changed by naked eyes under 360nm ultraviolet light. Therefore, the present embodiment can determine whether spike protein exists in the sample to be tested according to the change of the fluorescence signal intensity of the solution after the sample to be tested is added, i.e. determine whether the novel coronavirus exists in the sample to be tested.
The response time of the novel coronavirus detection method based on the luminescent quantum dot material provided by the embodiment is less than 2 minutes, the response of a sample to be detected can be rapidly realized, and the method has the advantages of rapidness, simplicity and convenience compared with technologies such as PCR (polymerase chain reaction) which need a longer response time and complicated sample processing steps; meanwhile, the detection method has lower requirements on detection instruments, can realize the detection of the sample to be detected by a common fluorescence spectrometer, is simpler, can be stably dispersed in a liquid phase, saves the complicated pretreatment step of the sample to be detected, and further provides convenience for the detection and storage of the sample; the detection method has a limit of detection (LOD) of 1 × 10 for the novel coronavirus-16mg/ml。
In some embodiments, the present invention further provides a method for detecting a novel coronavirus based on a luminescent quantum dot material, which can detect a concentration of the novel coronavirus in a sample to be detected, as shown in fig. 3, and specifically includes the following steps:
s100, adding CdTe quantum dots into a PBS buffer solution, and uniformly mixing to obtain a first mixed solution;
s200, adding a spike protein specific antibody into the first mixed solution, uniformly mixing to obtain a second mixed solution, and testing the PL spectral intensity of the second mixed solution;
s300, dripping a sample to be tested into the second mixed solution, uniformly mixing to obtain a third mixed solution, and testing the PL spectral intensity of the third mixed solution;
s400, adding the target spike proteins with different calibrated gradient concentrations into the second mixed solution, uniformly mixing to obtain a fourth mixed solution, and testing the PL spectral intensity of the fourth mixed solution to form a mapping table corresponding to the target spike proteins with different calibrated gradient concentrations and the PL spectral intensity;
s500, calculating to obtain the novel coronavirus concentration of the sample to be detected according to the mapping table and the PL spectral intensity of the third mixed solution.
In this embodiment, target spike proteins with different calibrated gradient concentrations are added into the second mixed solution in advance, so as to obtain PL spectrum intensities corresponding to the target spike proteins with different gradient concentrations, and form a corresponding mapping table; and finally, searching the target spike protein concentration corresponding to the PL spectral intensity of the third mixed solution from the mapping table according to the PL spectral intensity of the third mixed solution, namely the novel coronavirus concentration of the sample to be detected.
In a specific embodiment, 100ul of CdTe quantum dots with emission wavelength of 520nm are added into 1ml of PBS buffer solution, and after uniform mixing, PL spectrum is tested and recorded under excitation wavelength of 360 nm; adding 20ul of spike protein specific antibody CSab into the solution, slowly shaking for 15-30min to fully mix and react, testing the PL spectrum of the obtained solution under the same excitation condition, recording, and preparing the target spike protein into 1.0 x 10-4-1.0*10-16mg/ml, and the solutions are added dropwise from low to high in sequence, each gradient is 10ul, the PL spectrum after each gradient concentration is tested and is shown in figure 4, the PL spectrum intensity of the solution is gradually reduced along with the increase of the concentration of the target spike protein, and the PL spectrum in figure 4 is added dropwise from 1.0 x 10 from low to high in sequence according to the height of the wave peak-4mg/ml、1.0*10-5mg/ml、1.0*10-6mg/ml、 1.0*10-7mg/ml、1.0*10-8mg/ml、1.0*10-9mg/ml、1.0*10- 10mg/ml、 1.0*10-11mg/ml、1.0*1012mg/ml、1.0*10-13mg/ml、1.0*10-14mg/ml、 1.0*10-15mg/ml、1.0*10-16PL spectra measured after mg/ml, 0mg/ml target spike protein. The experimental results show that the novel coronavirus detection method based on the luminescent quantum dot material can rapidly and accurately detect spike protein, and the detection Limit (LOD) can reach 1 × 10-16mg/ml, with a linear range of 1 x 10-4—1*10-15。
In some embodiments, there is also provided a novel coronavirus detection system based on luminescent quantum dot materials, comprising a fluorescence immunosensor and a PL spectrometer, wherein the fluorescence immunosensor comprises CdTe quantum dots and specific antibody CSab bound to the surfaces of the CdTe quantum dots, and the specific antibody CSab is used for specifically binding with spike proteins of novel coronavirus surfaces.
In this embodiment, the CdTe quantum dots have a large number of negatively charged carboxyl groups on their surfaces, and after adding a spike protein specific antibody (CSab) to a first mixed solution containing CdTe quantum dots, the specific antibody of the spike protein has a positive surface electrical property, so that the CdTe quantum dots and the spike protein specific antibody can be combined together by electrostatic force to form a fluorescence immunosensor for specific recognition of the spike protein on the surface of the novel coronavirus; the fluorescence immunosensor can emit light under the irradiation of laser with specific wavelength due to the existence of the CdTe quantum dot, the light-emitting signal can be detected and recorded through a PL spectrometer, after a sample to be detected is added into a second mixed solution, if the sample to be detected contains spike protein, a specific antibody CSab in the fluorescence immunosensor can be specifically combined with the spike protein to form a CdTe quantum dot-CSab-spike protein system, so that the fluorescence intensity of the CdTe quantum dot is influenced, the CdTe quantum dot is quenched, the intensity is gradually reduced along with the concentration of the added spike protein, and data shows that the PL spectral intensity is linearly related to the concentration of the added spike protein in the quenching process, and when the concentration of the added spike protein is high enough, the color of the third mixed solution can be obviously changed by naked eyes under 360nm ultraviolet light. Therefore, the present embodiment can determine whether spike protein exists in the sample to be tested according to the change of the fluorescence signal intensity of the solution after the sample to be tested is added, i.e. determine whether the novel coronavirus exists in the sample to be tested.
In summary, the invention provides a novel coronavirus detection method and system based on luminescent quantum dot material, the response time of the detection method is less than 2 minutes, the response of a sample to be detected can be realized relatively quickly, and the detection method needs longer response time and complex sample treatment compared with PCR (polymerase chain reaction) and the likeThe step technology has the advantages of rapidness, simplicity and convenience; meanwhile, the detection method has lower requirements on detection instruments, can realize the detection of the sample to be detected by a common fluorescence spectrometer, is simpler, can be stably dispersed in a liquid phase, saves the complicated pretreatment step of the sample to be detected, and further provides convenience for the detection and storage of the sample; the detection method has a limit of detection (LOD) of 1 × 10 for the novel coronavirus-16mg/ml。
It is to be understood that the invention is not limited to the examples described above, but that modifications and variations may be effected thereto by those of ordinary skill in the art in light of the foregoing description, and that all such modifications and variations are intended to be within the scope of the invention as defined by the appended claims.
Claims (6)
1. A novel coronavirus detection method based on a luminescent quantum dot material is characterized by comprising the following steps:
adding CdTe quantum dots into a PBS buffer solution, and uniformly mixing to obtain a first mixed solution;
adding a spike protein specific antibody into the first mixed solution, uniformly mixing to obtain a second mixed solution, and testing the PL spectral intensity of the second mixed solution;
dripping a sample to be tested into the second mixed solution, uniformly mixing to obtain a third mixed solution, and testing the PL spectral intensity of the third mixed solution;
and if the PL spectral intensity of the third mixed solution is smaller than the PL spectral intensity of the second mixed solution, judging that the sample to be detected contains the novel coronavirus.
2. The method for detecting the novel coronavirus based on the luminescent quantum dot material as claimed in claim 1, further comprising the following steps:
adding the target spike proteins with different calibrated gradient concentrations into the second mixed solution, uniformly mixing to obtain a fourth mixed solution, and testing the PL spectral intensity of the fourth mixed solution to form a mapping table corresponding to the target spike proteins with different calibrated gradient concentrations and the PL spectral intensity;
and calculating to obtain the novel coronavirus concentration of the sample to be detected according to the mapping table and the PL spectral intensity of the third mixed solution.
3. The method for detecting coronavirus based on luminescent quantum dot material as claimed in claim 2, wherein the calibrated different gradient concentration of the target spike protein is 1.0 x 10-4-1.0*10-16mg/ml。
4. The method for detecting the novel coronavirus based on the luminescent quantum dot material as claimed in claim 1, wherein the steps of adding a spike protein specific antibody into the first mixed solution, uniformly mixing to obtain a second mixed solution, and testing the PL spectral intensity of the second mixed solution comprise:
adding a spike protein specific antibody into the first mixed solution, oscillating for 15-30min, and uniformly mixing to obtain a second mixed solution;
the PL spectral intensity of the second mixture was measured and recorded using a PL spectrometer at an excitation wavelength of 360 nm.
5. The method for detecting the novel coronavirus based on the luminescent quantum dot material, according to claim 1, wherein the step of dripping a sample to be detected into the second mixed solution, uniformly mixing to obtain a third mixed solution, and testing the PL spectral intensity of the third mixed solution comprises the following steps:
adding a sample to be detected into the second mixed solution, oscillating for 15-30min, and uniformly mixing to obtain a third mixed solution;
the PL spectral intensity of the third mixture was measured using a PL spectrometer at an excitation wavelength of 360nm and recorded.
6. A novel coronavirus detection system based on a luminescent quantum dot material is characterized by comprising a fluorescence immunosensor and a PL spectrometer, wherein the fluorescence immunosensor comprises CdTe quantum dots and a specific antibody CSab bound to the surfaces of the CdTe quantum dots, and the specific antibody CSab is used for specifically binding with spike proteins on the surfaces of novel coronaviruses.
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Application publication date: 20210112 |