CN106018796B - A kind of immunochromatography detecting system and its detection method for exciting dual frequency reception principle based on single-frequency - Google Patents
A kind of immunochromatography detecting system and its detection method for exciting dual frequency reception principle based on single-frequency Download PDFInfo
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Abstract
The immunochromatography detecting system of dual frequency reception principle is excited based on single-frequency.There is provided a kind of immunochromatography detecting system and its detection method for exciting dual frequency reception principle based on single-frequency.Including ultraviolet/visible spectrophotometer, information acquisition unit and the information process analysis unit being attached thereto, the ultraviolet/visible spectrophotometer is used for the maximum excitation wavelength for determining target detection thing；Described information collecting unit is used to carry out optical scanner to the target detection thing on test strip surface；Described information processing and analysis unit is used to carry out Treatment Analysis to described electric signal, so as to export testing result.The present invention is in use, the fluorescence that target detection thing in test strip is sent is divided into the light of two frequency ranges through two points of mirrors, the light intensity of the light of two frequency ranges is normalized again, so as to eliminate interference of the detection substrate to two wave bands, the light intensity of the fluorescence then sent by the target detection thing obtains the concentration information of target detection thing.
Description
Technical field
The invention belongs to detection technique field of quarantining, it is related to a kind of immunochromatography detecting system and method, and in particular to one
Plant the immunochromatography detecting system and its detection method that dual frequency reception principle is excited based on single-frequency.
Background technology
Fluorescence immune chromatography technology is easy to operate, quick because of its, and with very high specificity and sensitivity, will necessarily take
For colloidal gold immunochromatographimethod technology, and it is generalized to the clinical detection and antibiosis of the diseases such as CRP, HCG, myocardial damage, AIDS
The field of detection of food safety such as element, clenbuterol hydrochloride, melamine.
Fluorescent test paper strip not only has application value in clinical medicine application, will have more in civilian medical health maintenance field
Wide market, it has following several big advantages：1) cheap, test strips production cost is relatively low, is detected as routine health
Easily-consumed products, will not increase too big financial burden to user；2) it is easy to operate, easily-learned easily mastered, to the professional skill of user
Level and schooling are less demanding, are extremely suitable for commercial market；3) detect quick, detection knot can be just taken in the short time
Really, without waiting for；4) it is applicable index wide, can be applied to a variety of antigens and antibody carrier, and detection sensitivity is high；5) it can be applied to
Progress quantifies continuous monitoring and personalized health guarantee.
Due to the fast development of fluorescence immune chromatography technical need, fluoroscopic examination precision and sensitivity are proposed higher
It is required that.At present, interference source maximum in fluorescence immune chromatography detection process, exactly detects the fluorescence interference of substrate in sample, by
Also existed in interference substrate and target detection thing in detection sample, and the fluorescence main peak and target detection thing of interference substrate
Main peak approach so that detecting instrument can not distinguish differentiation interference substrate and target detection thing.At this stage grow up it is each
Modulation of source method is planted, the influence of the not homologous spectrum of background magnetic field and background can be eliminated, but some interference substrates are by homologous
Excitation source irradiates, and fluorescent modulation frequency is consistent with the frequency of detectable substance, so substrate can not be eliminated by modulation of source method
Fluorescence is disturbed.Having developed time-resolved fluorescence technology again in the recent period, (such as " one kind is characterized Chinese patent using time resolution fluorescence spectral
The method and apparatus of material magnetic effect "), it is intended to by fluorescence decay time control, distinguish long decaying phosphor cycle or short decay
Fluorescence target cycle detectable substance and substrate, so as to eliminate Substrate fluorescence influence.But, in the test event of whole blood, blood red egg
The white damped cycle for waiting Substrate fluorescence damped cycle and lanthanide series rare-earth elements is close, and is all hundreds of delicate ranks, so the time
Resolved fluorometric technology is also difficult to solve Substrate fluorescence interference, but, it is necessary to pass through during the concentration of detection target detection thing
The intensity of fluorescence obtains the concentration of target detection thing, therefore existing fluorescence immune chromatography technology can not accurately obtain target
The concentration of detectable substance.
The content of the invention
It is an object of the invention to overcome the shortcoming of above-mentioned prior art there is provided one kind to excite dual frequency reception based on single-frequency
The immunochromatography detecting system and its detection method of principle, the system and method can effectively eliminate the interference of Substrate fluorescence,
And it can accurately obtain the concentration information of target detection thing.
The technical scheme is that：
A kind of immunochromatography detecting system for exciting dual frequency reception principle based on single-frequency, including ultraviolet/visible spectrophotometric
Meter, information acquisition unit and the information process analysis unit being attached thereto,
Ultraviolet/the visible spectrophotometer is used for the maximum excitation wavelength for determining target detection thing；
Described information collecting unit is used to carry out optical scanner to the target detection thing on test strip surface, and it is to target
Detectable substance projects outgoing beam, to receive the reflected fluorescent light of test paper, and the reflected fluorescent light is converted into electric signal；
Described information processing and analysis unit is used to carry out Treatment Analysis to described electric signal, so that testing result is exported,
Described information collecting unit include excitation source (1), semi-transparent semi-reflecting lens (2), lens (3), two points of mirrors (4) and
The first sensor (5) and second sensor (6) of the light intensity of the fluorescence received for detection；
The directional light that excitation source (1) is sent is irradiated to detection examination after semi-transparent semi-reflecting lens (2) reflection and lens (3) successively
In the target detection thing on paper slip surface, the fluorescence that target detection thing is ejected is after semi-transparent semi-reflecting lens (2) transmission again through two points
Mirror (4) reflects and transmitted, wherein, the fluorescence that two points of mirrors (4) reflect is irradiated on the photosurface of first sensor (5), and two
The fluorescence that point mirror (4) is transmitted is irradiated on the photosurface of second sensor (6), the output end of first sensor (5) and the
Input of the output end of two sensors (6) with described information processing and analysis unit is connected.
The fluorescence that two points of mirrors (4) reflect is irradiated to the light of the first sensor (5) through the first optical filter (7)
On quick face.
The fluorescence that two points of mirrors (4) transmit is irradiated to the light of the second sensor (6) through the second optical filter (8)
On quick face.
The directional light that the excitation source (1) sends incides the semi-transparent semi-reflecting lens (2) back reflection to lens (3),
Angle between the incident directional light of wherein described semi-transparent semi-reflecting lens (2) and the directional light of reflection is 45~135 °.
Described information process analysis unit uses single-chip microcomputer.
A kind of detection method for the immunochromatography detecting system for exciting dual frequency reception principle based on single-frequency, including following step
Suddenly：
1) excitation wavelength of fluorescence spectrum, is determined：Target detection thing is determined 280 using ultraviolet/visible spectrophotometer
~760nm absorption curve, finds the most strong characteristic absorption peak correspondence peak position of target detection thing, and as the optimal of sample is excited
Wavelength；
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 light of sample
After spectrum is focused on through semi-transparent semi-reflecting lens (2) reflection and lens (3) successively, in lens (3), focal point is irradiated in test strip, examination
After target detection thing on paper slip is stimulated, launch the fluorescence spectrum of multiband, the spectrum is after lens (3), as parallel
Light transmission is to two points of mirrors (4) of top, and two frequency range spectrum are divided into both direction spectrum, respectively fluorescence one by two points of mirrors (4)
With fluorescence two；
4), fluorescence one converts optical signals to electric signal after the first optical filter (7) optical filtering by first sensor (5)
The detection light intensity of main peak is obtained, is output on single-chip microcomputer；
5), fluorescence two converts optical signals to electric signal after the second optical filter (8) optical filtering by second sensor (6)
The detection light intensity of secondary peak is obtained, is output on single-chip microcomputer；
6), calculating is normalized in the light intensity value for obtaining two spectral detections, obtains detecting light intensity, then substitute into and demarcate
Linear equation in calculate target detection thing concentration value, normalization algorithm expression formula is as follows：
A=α T1+ β T2
Wherein, calculated value, T1 and the measurement light intensity value that T2 is main peak and secondary peak after normalization of the A to measure light intensity, alpha+beta=
1；α and β parameters are determined according to the intensity relation of spectrogram.
The invention has the advantages that：The present invention in use, send out by the target detection thing in test strip
The fluorescence gone out is divided into the light of two frequency ranges through two points of mirrors, and first sensor and second sensor detect described two frequency ranges respectively
The light intensity of light, then the light intensity of the light of two frequency ranges is normalized, so as to obtain the fluorescence that target detection thing is sent
Light intensity, so as to eliminate interference of the detection substrate to two wave bands, the light intensity of the fluorescence then sent by the target detection thing
The concentration information of target detection thing is obtained, so as to effectively improve the accuracy and speed of the concentration information detected.
Brief description of the drawings
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 the calibration curve and equation of the first six group data in the embodiment of the present invention,
Fig. 6 is the calibration curve and equation of rear six groups of data in the embodiment of the present invention；
1 it is excitation source in figure, 2 be semi-transparent semi-reflecting lens, 3 be lens, 4 be two points of mirrors, 5 be first sensor, 6 is second
Sensor, 7 be the first optical filter, 8 be the second optical filter.
Embodiment
The present invention is described in further detail below in conjunction with the accompanying drawings.
As shown in figure 1, a kind of immunochromatography detecting system for 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,
Ultraviolet/the visible spectrophotometer is used to determine that the maximum excitation wavelength of target detection thing (to be utilized ultraviolet/visible
Spectrophotometric determination target detection thing finds the most strong characteristic absorption peak pair of target detection thing in 280-760nm absorption curve
Answer the maximum excitation wavelength of peak position, as sample)；
Described information collecting unit is used to carry out optical scanner to the target detection thing on test strip surface, and it is to target
Detectable substance projects outgoing beam, to receive the reflected fluorescent light of test paper, and the reflected fluorescent light is converted into electric signal；
Described information processing and analysis unit is used to carry out Treatment Analysis to described electric signal, so as to export testing result；
Described information collecting unit includes excitation source 1, semi-transparent semi-reflecting lens 2, lens 3, two points of mirrors 4 and received for detecting
The first sensor 5 and second sensor 6 of the light intensity of fluorescence；
The directional light that excitation source 1 is sent successively through semi-transparent semi-reflecting lens 2 reflect and lens 3 after be irradiated to test strip table
In the target detection thing in face, the fluorescence that target detection thing is ejected reflected again through two points of mirrors 4 after being transmitted through semi-transparent semi-reflecting lens 2 and
Transmission, wherein, the fluorescence that two points of mirrors 4 are reflected is irradiated on the photosurface of first sensor 5, what two points of mirrors 4 were transmitted
Fluorescence is irradiated on the photosurface of second sensor 6, the output end of first sensor 5 and the output end of second sensor 6 with
The input of described information processing and analysis unit is connected.
In order to obtain purer fluorescence, the fluorescence that two points of mirrors 4 are reflected is irradiated to described through the first optical filter 7
On the photosurface of first sensor 5；The fluorescence that two points of mirrors 4 are transmitted is irradiated to described second through the second optical filter 8 and passed
On the photosurface of sensor 6.
The directional light that the excitation source 1 is sent incides the back reflection of semi-transparent semi-reflecting lens 2 to lens 3, wherein institute
The angle stated between the incident directional light of semi-transparent semi-reflecting lens 2 and the directional light of reflection is 45~135 °, preferably 90 °.
Described information process analysis unit uses single-chip microcomputer.
Fig. 2 is a kind of detection side of immunochromatography detecting system for exciting dual frequency reception principle based on single-frequency of the present invention
Method, comprises the following steps：
1) excitation wavelength of fluorescence spectrum, is determined：Target detection thing is determined 280 using ultraviolet/visible spectrophotometer
~760nm absorption curve, finds the most strong characteristic absorption peak correspondence peak position of target detection thing, and as the optimal of sample is excited
Wavelength；
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
Successively after semi-transparent semi-reflecting lens 2 are reflected and lens 3 are focused on, it is irradiated in the focal point of lens 3 in test strip, in test strips
After target detection thing is stimulated, launch the fluorescence spectrum of multiband, the spectrum is transmitted to after lens 3 as directional light
Two frequency range spectrum are divided into both direction spectrum, respectively fluorescence one and fluorescence two by two points of mirrors 4 of side, two points of mirrors 4；
4), fluorescence one converts optical signals to electric signal by first sensor 5 and obtained after the optical filtering of the first optical filter 7
The detection light intensity of main peak, is output on single-chip microcomputer；
5), fluorescence two converts optical signals to electric signal by second sensor 6 and obtained after the optical filtering of the second optical filter 8
The detection light intensity of secondary peak, is output on single-chip microcomputer；
6), calculating is normalized in the light intensity value for obtaining two spectral detections, eliminates detection substrate to wherein some ripple
The certain interference of section, obtains detecting light intensity, then substitute into calculating target detection thing concentration value in the linear equation demarcated, normalization
Algorithm expression formula is as follows：
A=α T1+ β T2
Wherein, calculated value, T1 and the measurement light intensity value that T2 is main peak and secondary peak after normalization of the A to measure light intensity, alpha+beta=
1；α and β parameters are determined according to the intensity relation of spectrogram.
With reference to specific embodiment, the present invention is elaborated.
Fluorescent microsphere is synthesized using the metallo-chelate of lanthanide series rare-earth elements europium, as the fluorescent marker of Concentration Testing,
As shown in figure 3, the peak of the fluorescent marker excitation source is 345nm positions or so, the fluorescent marker emission band
Peak have at two, at one be 615nm peak position, another place be 590nm secondary peak position, peak position light intensity and
Secondary peak position light intensity ratio about 4：1 relation；
Assuming that the light intensity of 615nm measurements is T1, and the light intensity of 590nm measurements is T2, then calculates the light intensity value after normalization
For：A=0.8*T1+0.2*T2；Because 4：1 relation, so α is 0.8 and 0.2 with β parameter values；It can enter after obtained A values
Row piecewise linearity is demarcated, and calibration process is：
1. the metallo-chelate standard sample of concentration known europium is diluted to a series of gradients in proportion, obtained known to 11 groups
Concentration C0Standard sample (as shown in table 1)；
2. 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, the light intensity value A after being normalized0(result is as shown in table 1)；
3. by the light intensity value A after normalization0With concentration value C0Correspond, obtain standard curve and linear equation (such as Fig. 4,
5th, shown in 6, abscissa is light intensity value in Fig. 4,5,6, and ordinate is concentration value, and Fig. 5 is the first six group concentration value C and light intensity value
Function relation figure between A, now concentration value C and light intensity value A exponent function relations：C=6.406e0.004A, Fig. 6 is latter six groups
Function relation figure between concentration value C and light intensity value A, now concentration value C and light intensity value A is into logarithmic relationship：C=
137.0ln(A)-781.2；
Light intensity value after the initial concentration value of the standard sample of table 1 and normalization
Detect target detection thing：Target detection thing is added in test strip, the detecting system of the present invention, inspection is opened
Measure and corresponding light intensity value A=0.8*425+0.2*106=361.2, the value are obtained after T1=425, T2=106, normalization
It is in shown in Fig. 5 in scope, therefore light intensity value A is substituted into Fig. 5 linear equation C=6.406e0.004AIt is middle to calculate the dense of sample
Angle value, as a result about 27.17ng/mL.
Claims (6)
1. a kind of immunochromatography detecting system for exciting dual frequency reception principle based on single-frequency, including ultraviolet/visible spectrophotometer,
Information acquisition unit and the information process analysis unit being attached thereto,
Ultraviolet/the visible spectrophotometer is used for the maximum excitation wavelength for determining target detection thing；
Described information collecting unit is used to carry out optical scanner to the target detection thing on test strip surface, and it is to target detection
Thing projects outgoing beam, to receive the reflected fluorescent light of test paper, and the reflected fluorescent light is converted into electric signal；
Described information processing and analysis unit is used to carry out Treatment Analysis to described electric signal, so as to export testing result, it is special
Levy and be,
Described information collecting unit includes excitation source (1), semi-transparent semi-reflecting lens (2), lens (3), two points of mirrors (4) and is used for
Detect the first sensor (5) and second sensor (6) of the light intensity of the fluorescence received；
The directional light that excitation source (1) is sent is irradiated to test strip after semi-transparent semi-reflecting lens (2) reflection and lens (3) successively
In the target detection thing on surface, the fluorescence that target detection thing is ejected is 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 on the photosurface of first sensor (5), two points of mirrors
(4) fluorescence transmitted is irradiated on the photosurface of second sensor (6), the output end of first sensor (5) and the second biography
Input of the output end of sensor (6) with described information processing and analysis unit is connected.
2. a kind of immunochromatography detecting system for exciting dual frequency reception principle based on single-frequency according to claim 1, it is special
Levy and be, the fluorescence that two points of mirrors (4) reflect is irradiated to the light of the first sensor (5) through the first optical filter (7)
On quick face.
3. a kind of immunochromatography detecting system for exciting dual frequency reception principle based on single-frequency according to claim 1, it is special
Levy and be, the fluorescence that two points of mirrors (4) transmit is irradiated to the light of the second sensor (6) through the second optical filter (8)
On quick face.
4. a kind of immunochromatography detecting system for exciting dual frequency reception principle based on single-frequency according to claim 1, it is special
Levy and be, the directional light that the excitation source (1) sends incides the semi-transparent semi-reflecting lens (2) back reflection to lens (3), its
Described in angle between the incident directional light of semi-transparent semi-reflecting lens (2) and the directional light of reflection be 45~135 °.
5. a kind of immunochromatography detecting system for exciting dual frequency reception principle based on single-frequency according to claim 1, it is special
Levy and be, described information process analysis unit uses single-chip microcomputer.
6. a kind of detection side of immunochromatography detecting system for exciting dual frequency reception principle based on single-frequency as claimed in claim 1
Method, it is characterised in that comprise the following steps：
1) excitation wavelength of fluorescence spectrum, is determined：Using ultraviolet/visible spectrophotometer determine target detection thing 280~
760nm absorption curve, finds the optimum excitation wave of target detection thing most strong characteristic absorption peak correspondence peak position, as sample
It is long；
2), target detection thing is added in test strip；
3), according to step 1) test result, excitation source (1) send the maximum excitation wavelength directional light light beam of sample according to
It is secondary through semi-transparent semi-reflecting lens (2) reflection and lens (3) focus on after, in lens (3), focal point is irradiated in test strip, test strips
On target detection thing be stimulated after, launch the fluorescent light beam of multiband, the light beam is saturating as directional light after lens (3)
Two points of mirrors (4) of top are mapped to, two frequency range light beams are divided into both direction light beam by two points of mirrors (4), respectively fluorescence one and glimmering
Light two；
4), fluorescence one converts optical signals to electric signal by first sensor (5) and obtained after the first optical filter (7) optical filtering
The detection light intensity of main peak, is output on single-chip microcomputer；
5), fluorescence two converts optical signals to electric signal by second sensor (6) and obtained after the second optical filter (8) optical filtering
The detection light intensity of secondary peak, is output on single-chip microcomputer；
6), calculating is normalized in the light intensity value for obtaining two fluoroscopic examinations, obtains detecting light intensity, then substitute into the line demarcated
Property equation in calculate target detection thing concentration value, normalization algorithm expression formula is as follows：
A=α T1+ β T2
Wherein, A is calculated value after the normalization of measurement light intensity, and T1 and T2 is the measurement light intensity value of main peak and secondary peak, alpha+beta=1；α
With β parameters determined according to the intensity relation of spectrogram.
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| 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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| 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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