CN106018796A - Immunochromatographic detection system based on single-frequency excitation and double-frequency reception principle and detection method thereof - Google Patents
Immunochromatographic detection system based on single-frequency excitation and double-frequency reception principle and detection method thereof Download PDFInfo
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Abstract
The invention provides an immunochromatographic detection system based on the single-frequency excitation and double-frequency reception principle and a detection method thereof. The detection system comprises an ultraviolet/visible spectrophotometer, an information acquisition unit and an information processing and analyzing unit connected with the information acquisition unit, wherein the ultraviolet/visible spectrophotometer is used for determining the optimal excitation wavelength of a target detection object; the information acquisition unit is used for conducting optical scanning on the target detection object on the surface of a test strip; the information processing and analyzing unit is used for processing and analyzing electric signals so as to output a detection result. In the using process, fluorescent light emitted by the target detection object on the test strip is divided into light in two frequency bands through a dichotomy mirror, and then the light intensity of the light in the two frequency bands is normalized, so that disturbance of a detection substrate to the two frequency bands is eliminated, and then concentration information of the target detection object is obtained through the light intensity of the fluorescent light emitted by the target detection object.
Description
Technical field
The invention belongs to detection technique field of quarantining, relate to a kind of immunochromatography detecting system and method, be specifically related to one
Immunochromatography detecting system and the detection method thereof of dual frequency reception principle is excited based on single-frequency.
Background technology
Fluorescence immune chromatography technology is easy and simple to handle, quick because of it, and has the highest specificity and susceptiveness, necessarily will take
For colloidal gold immunochromatographimethod technology, and be generalized to the diseases such as CRP, HCG, myocardial damage, acquired immune deficiency syndrome (AIDS) Clinical detection and
The field of detection of food safety such as antibiotic, clenbuterol hydrochloride, tripolycyanamide.
Fluorescent test paper strip not only has using value in clinical medicine application, will have the widest in civilian medical health maintenance field
Wealthy market, it has following several big advantage: 1) cheap, test strips production cost is relatively low, detects as routine health
Easily-consumed products, will not be to the biggest financial burden of user increase；2) easy and simple to handle, easily-learned easily mastered, the professional skill to user
Energy level and schooling are less demanding, are extremely suitable for commercial market；3) detection is quickly, just can take inspection in the short time
Survey result, it is not necessary to wait；4) it is suitable for index wide, can be applicable to multiple antigen and antibody carrier, and detection sensitivity is high；5)
Can be applicable to carry out quantifying to monitor continuously and personalized health guarantee.
Due to the fast development of fluorescence immune chromatography technical need, fluoroscopic examination precision and sensitivity are proposed higher wanting
Ask.At present, maximum during fluorescence immune chromatography detection interference source, it is simply that the fluorescence interference of substrate in detection sample, due to
Interference substrate and target detection thing also exist in detection sample, and disturb the fluorescence main peak of substrate and the master of target detection thing
Peak is close, thus detecting instrument cannot distinguish differentiation interference substrate and target detection thing.The various light sources grown up of present stage
Modulator approach, can eliminate background magnetic field and the impact of background not homology spectrum, but some interference substrates are by homology excitation source
Irradiating, fluorescent modulation frequency is consistent with the frequency of detectable substance, disturbs so Substrate fluorescence cannot be eliminated by modulation of source method.
(such as Chinese patent, " one utilizes time resolution fluorescence spectral to characterize material magnetic effect to have developed again time-resolved fluorescence technology in the recent period
Method and apparatus "), it is intended to by fluorescence decay time control, distinguish long decaying phosphor cycle or short decaying phosphor target cycle
Detectable substance and substrate, thus eliminate Substrate fluorescence impact.But, in the test event of whole blood, the Substrate fluorescence such as hemoglobin
Damped cycle is close with the damped cycle of lanthanide series rare-earth elements, is all hundreds of delicate rank, so time-resolved fluorescence technology is also
It is difficult to solve Substrate fluorescence interference, but during the concentration of detection target detection thing, needs the intensity by fluorescence to obtain
The concentration of target detection thing, the most existing fluorescence immune chromatography technology can accurately not obtain the concentration of target detection thing.
Summary of the invention
It is an object of the invention to the shortcoming overcoming above-mentioned prior art, it is provided that one excites dual frequency reception former based on single-frequency
The immunochromatography detecting system of reason and detection method thereof, this system and method can effectively eliminate the interference of Substrate fluorescence, and
Can accurately obtain the concentration information of target detection thing.
The technical scheme is that
A kind of immunochromatography detecting system exciting dual frequency reception principle based on single-frequency, adopts including ultraviolet/visible spectrophotometer, information
Collection unit and the information process analysis unit being attached thereto,
Described ultraviolet/visible spectrophotometer is for determining the maximum excitation wavelength of target detection thing；
Described information acquisition unit is for carrying out optical scanning to the target detection thing on test strip surface, and it projects to target detection thing
Outgoing beam, to receive the reflected fluorescent light of reagent paper, and is converted to the signal of telecommunication by described reflected fluorescent light；
Described information process analysis unit for carrying out Treatment Analysis to the described signal of telecommunication, thus output detections result,
Described information acquisition unit includes excitation source (1), semi-transparent semi-reflecting lens (2), lens (3), two points of mirrors (4) and is used for
The first sensor (5) of the light intensity of the fluorescence that detection receives and the second sensor (6)；
The directional light that excitation source (1) sends is irradiated to test strip successively after semi-transparent semi-reflecting lens (2) reflection and lens (3)
On the target detection thing on surface, the fluorescence that target detection thing ejects after semi-transparent semi-reflecting lens (2) transmission again through two points of mirrors
(4) reflection and transmission, wherein, the fluorescence that two points of mirrors (4) reflect is irradiated to the photosurface of first sensor (5)
On, the fluorescence that two points of mirrors (4) transmit is irradiated on the photosurface of the second sensor (6), first sensor (5)
The outfan of outfan and the second sensor (6) all inputs with described information process analysis unit are connected.
The fluorescence that described two points of mirrors (4) reflect is irradiated to described first sensor (5) through the first optical filter (7)
On photosurface.
The fluorescence that described two points of mirrors (4) transmit is irradiated to described second sensor (6) through the second optical filter (8)
On photosurface.
The directional light that described excitation source (1) sends incides described semi-transparent semi-reflecting lens (2) back reflection to lens (3)
On, the angle between directional light and the directional light of reflection that wherein said semi-transparent semi-reflecting lens (2) is incident is 45～135 °.
Described information process analysis unit uses single-chip microcomputer.
The detection method of a kind of immunochromatography detecting system exciting dual frequency reception principle based on single-frequency, comprises the following steps:
1) excitation wavelength of fluorescence spectrum, is determined: utilize ultraviolet/visible spectrophotometer to measure target detection thing 280～760nm
Absorption curve, finds target detection thing the strongest characteristic absorption peak correspondence peak position, is the maximum excitation wavelength of sample；
2), target detection thing is added in test strip；
3), according to step 1) test result, excitation source (1) sends the maximum excitation wavelength directional light spectrum of sample and depends on
Secondary through semi-transparent semi-reflecting lens (2) reflection and lens (3) focus on after, be irradiated in test strip at lens (3) focal point,
After target detection thing in test strips is stimulated, launching the fluorescence spectrum of multiband, this spectrum, after lens (3), becomes
Directional light is transmitted to two points of mirrors (4) of top, and two frequency range spectrum are divided into both direction spectrum by two points of mirrors (4), is respectively
Fluorescence one and fluorescence two；
4), fluorescence one through the first optical filter (7) filter after, first sensor (5) convert optical signals to the signal of telecommunication and led
The detection light intensity at peak, exports on single-chip microcomputer；
5), fluorescence two after the second optical filter (8) filters, the second sensor (6) convert optical signals to the signal of telecommunication and obtain time
The detection light intensity at peak, exports on single-chip microcomputer；
6), the light intensity value that two spectral detection obtain is normalized calculating, obtains detecting light intensity, then substitute into the linear side demarcated
Calculating target detection concentration value in journey, normalization algorithm expression formula is as follows:
A=α T1+ β T2
Wherein, A be measure light intensity normalization after value of calculation, T1 Yu T2 is the measurement light intensity value of main peak and secondary peak, alpha+beta=1；
α Yu β parameter is that the intensity relation according to spectrogram determines.
The method have the advantages that the present invention in use, the target detection thing in test strip sends
Fluorescence be divided into the light of two frequency ranges through two points of mirrors, first sensor and the second sensor detect the light of said two frequency range respectively
Light intensity, then the light intensity of the light of two frequency ranges is normalized, thus obtain the light intensity of the fluorescence that target detection thing sends,
Thus eliminating the detection substrate interference to two wave bands, the light intensity of the fluorescence then sent by described target detection thing obtains target
The concentration information of detectable substance, thus effectively improve the accuracy and speed of the concentration information detected.
Accompanying drawing explanation
Fig. 1 is the principle assumption diagram of the present invention,
Fig. 2 is the workflow of the present invention,
Fig. 3 is the spectrogram in the embodiment of the present invention,
Fig. 4 is calibration map in the embodiment of the present invention,
Fig. 5 is calibration curve and the equation of the first six group data in the embodiment of the present invention,
Fig. 6 is calibration curve and the equation of rear six groups of data in the embodiment of the present invention；
Figure 1 is excitation source, 2 is semi-transparent semi-reflecting lens, 3 is lens, 4 is two points of mirrors, 5 is first sensor, 6 is the second biography
Sensor, 7 be the first optical filter, 8 be the second optical filter.
Detailed description of the invention
Below in conjunction with the accompanying drawings the present invention is described in further detail.
As it is shown in figure 1, a kind of immunochromatography detecting system exciting dual frequency reception principle based on single-frequency of the present invention, including
Ultraviolet/visible spectrophotometer, information acquisition unit and the information process analysis unit being attached thereto,
Described ultraviolet/visible spectrophotometer is for determining that the maximum excitation wavelength of target detection thing (utilizes ultraviolet/visible spectrophotometer
Mensuration target detection thing, at the absorption curve of 280-760nm, finds target detection thing the strongest characteristic absorption peak correspondence peak position, i.e.
Maximum excitation wavelength for sample)；
Described information acquisition unit is for carrying out optical scanning to the target detection thing on test strip surface, and it projects to target detection thing
Outgoing beam, to receive the reflected fluorescent light of reagent paper, and is converted to the signal of telecommunication by described reflected fluorescent light；
Described information process analysis unit for carrying out Treatment Analysis to the described signal of telecommunication, thus output detections result；Described information is adopted
Collection unit includes excitation source 1, semi-transparent semi-reflecting lens 2,3, two points of mirrors 4 of lens and the light of fluorescence received for detection
Strong first sensor 5 and the second sensor 6；
The directional light that excitation source 1 sends reflects through semi-transparent semi-reflecting lens 2 successively and is irradiated to the target on test strip surface after lens 3
In detectable substance, the fluorescence that target detection thing ejects reflects and transmission through two points of mirrors 4 after semi-transparent semi-reflecting lens 2 transmission again, its
In, the fluorescence that two points of mirrors 4 reflect is irradiated on the photosurface of first sensor 5, and the fluorescence that two points of mirrors 4 transmit shines
Be mapped on the photosurface of the second sensor 6, the outfan of first sensor 5 and the outfan of the second sensor 6 all with described letter
The input of breath processing and analysis unit is connected.
In order to obtain purer fluorescence, the fluorescence that described two points of mirrors 4 reflect is irradiated to described through the first optical filter 7
On the photosurface of first sensor 5；The fluorescence that described two points of mirrors 4 transmit is irradiated to described second through the second optical filter 8 and passes
On the photosurface of sensor 6.
The directional light that described excitation source 1 sends incides described semi-transparent semi-reflecting lens 2 back reflection to lens 3, Qi Zhongsuo
Stating the angle between directional light and the directional light of reflection of semi-transparent semi-reflecting lens 2 incidence is 45～135 °, preferably 90 °.
Described information process analysis unit uses single-chip microcomputer.
Fig. 2 is the detection method of a kind of immunochromatography detecting system exciting dual frequency reception principle based on single-frequency of the present invention,
Comprise the following steps:
1) excitation wavelength of fluorescence spectrum, is determined: utilize ultraviolet/visible spectrophotometer to measure target detection thing 280～760nm
Absorption curve, finds target detection thing the strongest characteristic absorption peak correspondence peak position, is the maximum excitation wavelength of sample；
2), target detection thing is added in test strip；
3), according to step 1) test result, excitation source 1 sends the maximum excitation wavelength directional light spectrum warp successively of sample
Semi-transparent semi-reflecting lens 2 reflects and after lens 3 focusing, is irradiated in test strip at lens 3 focal point, the target in test strips
After detectable substance is stimulated, launching the fluorescence spectrum of multiband, this spectrum, after lens 3, becomes directional light and is transmitted to top
Two points of mirrors 4, two frequency range spectrum are divided into both direction spectrum, respectively fluorescence one and fluorescence two by two points of mirrors 4；
4), fluorescence one through first optical filter 7 filter after, first sensor 5 convert optical signals to the signal of telecommunication and obtain the inspection of main peak
Light-metering is strong, exports on single-chip microcomputer；
5), fluorescence two through second optical filter 8 filter after, the second sensor 6 convert optical signals to the signal of telecommunication and obtain the inspection of secondary peak
Light-metering is strong, exports on single-chip microcomputer；
6), by the light intensity value that two spectral detection obtain it is normalized calculating, eliminates detection substrate and the specific of wherein certain wave band is done
Disturb, obtain detecting light intensity, then substitute into calculating target detection concentration value, normalization algorithm expression formula in the linear equation demarcated
As follows:
A=α T1+ β T2
Wherein, A be measure light intensity normalization after value of calculation, T1 Yu T2 is the measurement light intensity value of main peak and secondary peak, alpha+beta=1；
α Yu β parameter is that the intensity relation according to spectrogram determines.
Below in conjunction with specific embodiment, the present invention is elaborated.
Use the metallo-chelate synthesis fluorescent microsphere of lanthanide series rare-earth elements europium, as the fluorescent marker of Concentration Testing, as
Shown in Fig. 3, the peak of this fluorescent marker excitation source is about 345nm position, this fluorescent marker emission band
Peak have at two, be the peak position of 615nm at one, be the secondary peak position of 590nm at another, peak position light
By force with secondary peak position light intensity than about 4:1 relation；
Assume that the light intensity that 615nm measures is T1, and the light intensity that 590nm measures is T2, then the light intensity value after calculating normalization is:
A=0.8*T1+0.2*T2；Because 4:1 relation, so α Yu β parameter value is 0.8 and 0.2；After the A value obtained
Can carry out piecewise linearity demarcation, calibration process is:
1. the metallo-chelate standard sample of concentration known europium is diluted to a series of gradient in proportion, obtains 11 groups of concentration known C0's
Standard sample (as shown in table 1)；
The most each standard sample is detected by the detecting system of the present invention, obtains their T10And T20Value, and substitute into formula
A=α T1+ β T2, obtains the light intensity value A after normalization0(result is as shown in table 1)；
3. by the light intensity value A after normalization0With concentration value C0One_to_one corresponding, obtains standard curve and linear equation (such as Fig. 4,5,6
Shown in, in Fig. 4,5,6, abscissa is light intensity value, and vertical coordinate is concentration value, and Fig. 5 is the first six group concentration value C and light intensity
Function relation figure between angle value A, now concentration value C and light intensity value A exponent function relation: C=6.406e0.004A, Fig. 6
Being rear function relation figure between six groups of concentration value C and light intensity value A, now concentration value C becomes logarithm to close with light intensity value A
System: C=137.0ln (A)-781.2；
Light intensity value after the initial concentration value of table 1 standard sample and normalization
Detection target detection thing: add in test strip by target detection thing, opens the detecting system of the present invention, and detection obtains
T1=425, T2=106, obtain the light intensity value A=0.8*425+0.2*106=361.2 of correspondence after normalization, this value is in Fig. 5 institute
In the range of showing, therefore this light intensity value A is substituted into the linear equation C=6.406e of Fig. 50.004AThe concentration value of middle calculating sample, knot
Fruit is about 27.17ng/mL.
Claims (6)
1. excite an immunochromatography detecting system for dual frequency reception principle based on single-frequency, including ultraviolet/vis spectroscopy light
Degree meter, information acquisition unit and the information process analysis unit being attached thereto,
Described ultraviolet/visible spectrophotometer is for determining the maximum excitation wavelength of target detection thing；
Described information acquisition unit is for carrying out optical scanning to the target detection thing on test strip surface, and it is to target
Detectable substance projects irradiating light beam, to receive the reflected fluorescent light of reagent paper, and described reflected fluorescent light is converted to the signal of telecommunication；
Described information process analysis unit for carrying out Treatment Analysis to the described signal of telecommunication, thus output detections result,
It is characterized in that,
Described information acquisition unit includes excitation source (1), semi-transparent semi-reflecting lens (2), lens (3), two points of mirrors
(4) first sensor (5) of the light intensity of fluorescence and for detection received and the second sensor (6)；
The directional light that excitation source (1) sends is irradiated to successively after semi-transparent semi-reflecting lens (2) reflection and lens (3)
On the target detection thing on test strip surface, the fluorescence that target detection thing ejects is through semi-transparent semi-reflecting lens (2)
Again through the reflection of two points of mirrors (4) and transmission after transmission, wherein, the fluorescence that two points of mirrors (4) reflect is irradiated to
On the photosurface of first sensor (5), the fluorescence that two points of mirrors (4) transmit is irradiated to the second sensor (6)
Photosurface on, the outfan of first sensor (5) and the outfan of the second sensor (6) all with described letter
The input of breath processing and analysis unit is connected.
The most according to claim 1 a kind of based on single-frequency excite dual frequency reception principle immunochromatography detection system
System, it is characterised in that the fluorescence that described two points of mirrors (4) reflect is irradiated to institute through the first optical filter (7)
State on the photosurface of first sensor (5).
The most according to claim 1 a kind of based on single-frequency excite dual frequency reception principle immunochromatography detection system
System, it is characterised in that the fluorescence that described two points of mirrors (4) transmit is irradiated to institute through the second optical filter (8)
State on the photosurface of the second sensor (6).
The most according to claim 1 a kind of based on single-frequency excite dual frequency reception principle immunochromatography detection system
System, it is characterised in that the directional light that described excitation source (1) sends incides described semi-transparent semi-reflecting lens (2)
Back reflection on lens (3), the directional light of the incident directional light of wherein said semi-transparent semi-reflecting lens (2) and reflection
Between angle be 45～135 °.
The most according to claim 1 a kind of based on single-frequency excite dual frequency reception principle immunochromatography detection system
System, it is characterised in that described information process analysis unit uses single-chip microcomputer.
A kind of immunochromatography detecting system exciting dual frequency reception principle based on single-frequency
Detection method, it is characterised in that comprise the following steps:
1) excitation wavelength of fluorescence spectrum, is determined: utilize ultraviolet/visible spectrophotometer to measure target detection thing and exist
The absorption curve of 280～760nm, finds target detection thing the strongest characteristic absorption peak correspondence peak position, is detected sample
The maximum excitation wavelength of product；
2), target detection thing is added in test strip；
3), according to step 1) test result, excitation source (1) send sample maximum excitation wavelength put down
Row light spectrum is successively after semi-transparent semi-reflecting lens (2) reflection and lens (3) focus on, at lens (3) focal point
It is irradiated in test strip, after the target detection thing in test strips is stimulated, launches the fluorescence spectrum of multiband,
This spectrum, after lens (3), becomes directional light and is transmitted to two points of mirrors (4) of top, two points of mirrors (4)
Two frequency range spectrum are divided into both direction spectrum, respectively fluorescence one and fluorescence two；
4), fluorescence one through the first optical filter (7) filter after, first sensor (5) convert optical signals to
The signal of telecommunication obtains the detection light intensity of main peak, exports on single-chip microcomputer；
5), fluorescence two through the second optical filter (8) filter after, the second sensor (6) convert optical signals to
The signal of telecommunication obtains the detection light intensity of secondary peak, exports on single-chip microcomputer；
6), the light intensity value that two spectral detection obtain is normalized calculating, obtains detecting light intensity, then substitute into mark
Calculating target detection concentration value in the linear equation reserved, normalization algorithm expression formula is as follows:
A=α T1+ β T2
Wherein, A be measure light intensity normalization after value of calculation, T1 Yu T2 is the measurement light intensity value of main peak and secondary peak, α+
β=1；α Yu β parameter is that the intensity relation according to spectrogram determines.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108982448A (en) * | 2018-07-20 | 2018-12-11 | 郑州迈迪迅医疗科技有限公司 | A kind of fluorescence immunity analyzer and its detection method |
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| US20030008314A1 (en) * | 2001-06-28 | 2003-01-09 | Priuska Eric M. | Fiber-optic sensor array |
| CN201917521U (en) * | 2010-12-17 | 2011-08-03 | 北京柏奥达思科技有限公司 | Confocal fluorescent detection device and immunity bedside detector adopting same |
| CN204008471U (en) * | 2014-08-22 | 2014-12-10 | 苏州百慧华业精密仪器有限公司 | A kind of quantitative testing device of test strips |
| CN205786658U (en) * | 2016-07-05 | 2016-12-07 | 扬州千代科技有限公司 | The immunochromatography detecting system of dual frequency reception principle is excited based on single-frequency |
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2016
- 2016-07-05 CN CN201610526097.9A patent/CN106018796B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030008314A1 (en) * | 2001-06-28 | 2003-01-09 | Priuska Eric M. | Fiber-optic sensor array |
| CN201917521U (en) * | 2010-12-17 | 2011-08-03 | 北京柏奥达思科技有限公司 | Confocal fluorescent detection device and immunity bedside detector adopting same |
| CN204008471U (en) * | 2014-08-22 | 2014-12-10 | 苏州百慧华业精密仪器有限公司 | A kind of quantitative testing device of test strips |
| CN205786658U (en) * | 2016-07-05 | 2016-12-07 | 扬州千代科技有限公司 | The immunochromatography detecting system of dual frequency reception principle is excited based on single-frequency |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108982448A (en) * | 2018-07-20 | 2018-12-11 | 郑州迈迪迅医疗科技有限公司 | A kind of fluorescence immunity analyzer and its detection method |
| CN108982448B (en) * | 2018-07-20 | 2021-11-02 | 郑州迈迪迅医疗科技有限公司 | Fluorescence immunoassay instrument and detection method thereof |
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