CN113552337A - Full-spectrum high-efficiency microplate reader system - Google Patents
Full-spectrum high-efficiency microplate reader system Download PDFInfo
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- CN113552337A CN113552337A CN202110815905.4A CN202110815905A CN113552337A CN 113552337 A CN113552337 A CN 113552337A CN 202110815905 A CN202110815905 A CN 202110815905A CN 113552337 A CN113552337 A CN 113552337A
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- 238000001228 spectrum Methods 0.000 title claims abstract description 43
- 238000002965 ELISA Methods 0.000 claims abstract description 32
- 238000002835 absorbance Methods 0.000 claims abstract description 14
- 239000013307 optical fiber Substances 0.000 claims description 10
- 230000008878 coupling Effects 0.000 claims description 4
- 238000010168 coupling process Methods 0.000 claims description 4
- 238000005859 coupling reaction Methods 0.000 claims description 4
- 238000005259 measurement Methods 0.000 abstract description 9
- 238000012360 testing method Methods 0.000 abstract description 5
- 239000000126 substance Substances 0.000 abstract description 4
- 238000004904 shortening Methods 0.000 abstract description 2
- 238000001514 detection method Methods 0.000 description 5
- 230000003287 optical effect Effects 0.000 description 5
- 102000004190 Enzymes Human genes 0.000 description 4
- 108090000790 Enzymes Proteins 0.000 description 4
- 238000006243 chemical reaction Methods 0.000 description 3
- 239000000835 fiber Substances 0.000 description 3
- 230000003595 spectral effect Effects 0.000 description 3
- 230000003321 amplification Effects 0.000 description 2
- 239000000427 antigen Substances 0.000 description 2
- 102000036639 antigens Human genes 0.000 description 2
- 108091007433 antigens Proteins 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 230000007547 defect Effects 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 238000000034 method Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000003199 nucleic acid amplification method Methods 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 238000012545 processing Methods 0.000 description 2
- UJMBCXLDXJUMFB-GLCFPVLVSA-K tartrazine Chemical compound [Na+].[Na+].[Na+].[O-]C(=O)C1=NN(C=2C=CC(=CC=2)S([O-])(=O)=O)C(=O)C1\N=N\C1=CC=C(S([O-])(=O)=O)C=C1 UJMBCXLDXJUMFB-GLCFPVLVSA-K 0.000 description 2
- 229960000943 tartrazine Drugs 0.000 description 2
- 235000012756 tartrazine Nutrition 0.000 description 2
- 239000004149 tartrazine Substances 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 235000013405 beer Nutrition 0.000 description 1
- 238000006555 catalytic reaction Methods 0.000 description 1
- 239000003593 chromogenic compound Substances 0.000 description 1
- 238000004737 colorimetric analysis Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 238000011160 research Methods 0.000 description 1
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/01—Arrangements or apparatus for facilitating the optical investigation
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/255—Details, e.g. use of specially adapted sources, lighting or optical systems
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/01—Arrangements or apparatus for facilitating the optical investigation
- G01N2021/0106—General arrangement of respective parts
- G01N2021/0112—Apparatus in one mechanical, optical or electronic block
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Abstract
The invention provides a full-spectrum high-efficiency microplate reader system, which comprises: the device comprises a full-spectrum light source, an ELISA plate, a detector for measuring the full-spectrum absorbance of a sample transilluminated by the full spectrum in a hole of the ELISA plate, and a motion control module for controlling the movement of the ELISA plate relative to the full-spectrum light source. The system can measure the absorbance data of a sample full spectrum (200nm-1100nm) at one time without adjusting the wavelength during measurement, thereby greatly shortening the measurement time and obtaining more accurate data for unstable substances. The motion and the positioning of the ELISA plate can be accurately controlled through the motion control module, and the accuracy of the test is ensured.
Description
Technical Field
The invention relates to the field of detection instruments, in particular to a full-spectrum high-efficiency microplate reader system.
Background
A microplate Reader (Micro-plate Reader) is an instrument for reading and analyzing the experimental results of enzyme-linked immunosorbent assay (EIA). The existing enzyme-linked immunosorbent assay is based on a colorimetric method and is carried out by an enzyme catalysis chromogenic substrate coupled on an antigen or an antibody, the reaction result is displayed by color, and the concentration of the antibody or the antigen to be detected in a sample is judged according to the depth of color development, namely the magnitude of absorbance value. Microplate readers are widely used in clinical examinations, biological research, agricultural science, food and environmental science, and especially in recent years, the use of microplate readers in hospitals or domestic communities has become more and more widespread due to the use of a large number of enzyme-linked immunoassay test cassettes.
The microplate reader is a phase-change photoelectric colorimeter or spectrophotometer in fact, and the basic working principle of the microplate reader is basically the same as that of a main structure and the photoelectric colorimeter. The light wave emitted by the light source lamp is changed into a beam of monochromatic light through the optical filter or the monochromator, and enters the sample to be measured in the plastic micropore pole. One part of the monochromatic light is absorbed by the specimen, the other part of the monochromatic light penetrates through the specimen and irradiates on a photoelectric detector, the photoelectric detector converts the light signals with different strengths of the specimen to be detected into corresponding electric signals, the electric signals are sent into a microprocessor for data processing and calculation after signal processing such as prepositive amplification, logarithmic amplification, analog-to-digital conversion and the like, and finally, the result is displayed by a display and a printer. The microprocessor also controls the movement of the mechanical driving mechanism in the X direction and the Y direction through the control circuit to move the microporous plate, thereby realizing the automatic sample introduction detection process. And other microplate readers adopt manual movement of a microporous plate for detection, so that a mechanical driving mechanism and a control circuit in the X and Y directions are omitted, the microplate reader is smaller and more compact, and the microplate reader is simpler in structure.
The basic functions of an enzyme-labeled analyzer as a microplate colorimeter are not limited to colorimetric determination, but are different in terms of determination wavelength range, absorbance range, optical system, detection speed, temperature control, qualitative and quantitative determination software functions, and the like. The existing middle and low-end enzyme mark analyzers in the market can obtain different measuring wavelengths by adopting different optical filters, and the enzyme mark analyzer needs to be provided with a component capable of automatically converting the optical filters. The instrument can be simultaneously provided with 6-8 optical filters, and the measuring wavelength is between 400-700 nm and comprises the two wavelengths. The high-end microplate reader adopts a grating system to filter light, and light with specific wavelength and adjustable step can be obtained. The measurement is performed by the absorbance of light by the substance to be measured.
The existing microplate reader measures the absorbance of a full spectrum (200-. The measurement principle is that the light source generates light of 200nm, 200nm data is measured and stored, the light source is adjusted to 201nm step by step, and the measurement and the storage are carried out in sequence. 3-5 minutes were required to complete one well. If the 96-well data is measured, it takes several hours. Too long time can seriously affect the experiment. For some reagents that are not characterized by long-term storage in an air environment, are susceptible to oxidation, or are susceptible to other chemical reactions, the reagents may have been denatured before the data is tested. The defect seriously influences the experimental efficiency and the experimental accuracy, and simultaneously is not convenient for rapidly analyzing the characteristics of the substance in the full spectrum.
Disclosure of Invention
The invention aims to provide a full-spectrum high-efficiency microplate reader system aiming at the defects of the prior art, the system is not used for obtaining light with a certain specific wavelength through a light filtering system according to the traditional technical route, but uses a light source with full-spectrum wavelength as incident light, and abandons the traditional photoelectric sensor for simply detecting light intensity in the aspect of detection. Instead, a miniature fiber optic spectrometer containing a CCD capable of measuring the full spectrum is used as the detector. Therefore, the invention can detect all light intensity data in the effective wavelength range of the micro spectrometer and the light source system, and has extremely short time consumption.
The technical scheme adopted by the invention is as follows:
a full spectrum high efficiency microplate reader system, comprising:
the device comprises a full-spectrum light source, an ELISA plate, a detector for measuring the full-spectrum absorbance of a sample transilluminated by the full spectrum in a hole of the ELISA plate, and a motion control module for controlling the movement of the ELISA plate relative to the full-spectrum light source.
Further, the detector is a spectrometer.
Furthermore, the motion control module comprises a stepping motor, a master control circuit, a power supply, a driver, two groups of guide rails, a sliding block, a coupling, an ELISA plate bracket for fixing the ELISA plate, an upper computer and a sensor; the two groups of guide rails and the slide blocks are respectively arranged in the X direction and the Y direction, the ELISA plate support is fixed on the slide blocks of one group of guide rails and slide blocks, and the bottoms of the group of guide rails and slide blocks are fixed on the slide blocks of the other group of guide rails and slide blocks through the shaft coupling. The sensor is arranged at the original point position in the X and Y directions and is used for locating the original point of the ELISA plate.
The main control circuit is used for receiving a motion instruction sent by the upper computer and sending the motion instruction to the driver, the driver drives the stepping motor to control the sliding block to move on the guide rail, and the power supply provides electric energy for the driver.
And the upper computer sends out a motion instruction according to the current positions and preset positions of the two groups of guide rails and the slide blocks.
Furthermore, the upper computer is also connected with the detector and used for displaying current waveform data and storing the current data.
Furthermore, the master control circuit integrates a control chip, a serial port chip, a key and a signal output circuit, the signal output circuit is connected with the driver, the control chip adopts STM32, and the serial port chip adopts CH 340.
Furthermore, the device also comprises two collimating mirrors which are respectively connected with the full-spectrum light source and the detector through optical fibers and are oppositely arranged.
The invention has the beneficial effects that:
(1) the system can measure the absorbance data of a sample full spectrum (200nm-1100nm) at one time without adjusting the wavelength during measurement, thereby greatly shortening the measurement time and obtaining more accurate data for unstable substances. Taking a 96-well plate as an example, measuring one well data may take only 10-20 seconds, and the time required to complete all 96 wells is only 1/30-1/15 of the prior art route. Greatly improves the experimental efficiency and is very beneficial to the experiment with a small time window. After the spectral data are measured, the spectral data can be instantly operated according to the Lambert beer law through a software program end, and the absorbance value is obtained. The resolution depends on the resolution of the micro spectrometer, which is an important part.
(2) The motion and the positioning of the ELISA plate can be accurately controlled through the motion control module, and the accuracy of the test is ensured.
Drawings
FIG. 1 is a schematic diagram of the system architecture;
FIG. 2 is a system block diagram;
FIG. 3 is a flowchart of a master control routine;
FIG. 4 is a graph of the test results for tartrazine solution;
in the figure: 1-full spectrum light source, 2-optical fiber, 3-spectrometer, 4-optical fiber support, 5-collimating mirror flange, 6-collimating mirror, 7-instrument shell, 8-ELISA plate support, 9-bearing and baffle, 10-ball screw, 11-guide rail, 12-base, 13-driver, 14-stepping motor, 15-main control circuit, 16-power supply, 17-bearing, 18-coupler, 19-workbench and 20-proximity switch.
Detailed Description
The invention discloses a full-spectrum high-efficiency microplate reader system, which comprises:
the device comprises a full-spectrum light source, an ELISA plate, a detector for measuring the full-spectrum absorbance of a sample transilluminated by the full spectrum in a hole of the ELISA plate, and a control device for controlling the movement of the ELISA plate relative to the full-spectrum light source.
It should be noted that the detector for measuring the full-spectrum absorbance of the sample transilluminated by the full spectrum in the hole of the microplate is a detector which optionally contains a CCD and can measure the full spectrum, such as a spectrometer and the like; the control device can be a belt drive, a sliding table and the like.
The invention is further illustrated with reference to a specific embodiment.
As shown in fig. 1-2, the system mainly comprises a full spectrum light source 1, a spectrometer 3, a high-precision ball screw two-dimensional sliding table and an elisa plate, wherein the elisa plate is a 96-porous plate. High accuracy ball two-dimensional slip table specifically includes: the device comprises a stepping motor 14, a main control circuit 15, a power supply 16, a driver 13, two groups of guide rails and sliders, a coupler 18, an ELISA plate bracket 8, an upper computer and a sensor. The two groups of guide rails and the slide blocks are respectively arranged in the X direction and the Y direction, the ELISA plate bracket 8 is fixed on the slide blocks of one group of guide rails and slide blocks, and the bottoms of the group of guide rails and slide blocks are fixed on the slide blocks of the other group of guide rails and slide blocks through the shaft coupling 18. The proximity switch 20, i.e., the zero point sensor, is disposed at the original point position in the X and Y directions, and is used for locating the original point of the microplate.
The upper computer is used for selecting a hole to be measured, setting a preset position and sending a motion instruction according to the current positions and preset positions of the two groups of guide rails and the sliding block. And simultaneously displaying the current waveform data and storing the current data. The waveform data includes spectral data and absorbance data. The data can be saved locally in the form of an array.
The main control circuit 15 is mainly used for receiving a motion instruction sent by an upper computer and sending a signal for controlling the motor to the driver 13, the power supply 16 is connected with the driver 13, and the driver 13 drives the stepping motor 14 to operate. The main control circuit 15 is integrated with a control chip, a serial port chip, a key and a signal output circuit, preferably, the control chip adopts STM32, and the serial port chip adopts CH 340. The signal output circuit is directly connected with the signal input port of the driver. The key can also be used for setting data of a plurality of holes to be scanned and stored fully automatically at a time.
Specifically, the program flow of the main control circuit 15 is as shown in fig. 3, and after the system is powered on, the current position automatically returns to zero, that is: the main control circuit 15 starts a timer, judges whether a sensor signal reaches a threshold value, stops sending a control signal to the driver 13 if the sensor signal indicates that the current position is the original position, sends the current position to the upper computer to be zero, and interrupts the timer; if not, a control signal is sent to the driver 13, and the judgment is carried out again according to the sensor signal until the threshold value is reached. And then the main control circuit 15 receives the instruction of the upper computer, judges whether a correct instruction is received, if so, sends a control signal to the driver 13, the driver 13 controls the stepping motor 14 to move, calculates the current position after the movement according to the pulse, and sends the current position to the upper computer, otherwise, receives the instruction of the upper computer again until the preset position is reached.
Further, the ELISA plate support 8 is of a middle hollow structure, the ELISA plate is fixed in the middle of the ELISA plate support 8 through the edge, light path measurement is facilitated, and meanwhile the ELISA plate can be taken and placed.
As a preferred scheme, the optical fiber support device further comprises two collimating mirrors 6 and an optical fiber support 4, the optical fiber support 4 is of an E-shaped three-layer plate structure, the upper layer and the lower layer are respectively used for fixing the two collimating mirrors 6, specifically, the full-spectrum light source 1(360nm-2000nm) is connected with the optical fiber 2 of the SMA905 joint, the other end of the optical fiber 2 is connected with the collimating mirror 6 through threads, and the collimating mirror 6 is vertically connected with the middle plate of the optical fiber support 4 through threads and a collimating mirror flange 5. The spectrometer 3 is connected to the top plate of the fiber support 4 by the same fiber 2 and collimator 6 and flange 5. The two collimating mirrors 6 are arranged opposite to each other. The 96 multi-hole plate can move between the top plate and the middle plate under the control of the ELISA plate bracket 8 and the motion control module, and the specified position is aligned with the measuring light path for measurement.
In this example, a series of solutions were prepared in a gradient for tartrazine powder, and 9 wells of solutions were added to each concentration in a 96-well plate. There were 48 wells containing pure water. Half the number of a 96-well plate is used. The data measured by the microplate reader of the invention is shown in fig. 4, and the section of test data is removed because the noise of the invisible light wave band part is larger. The magnitude of absorbance of solutions of different concentrations can be seen with reference to pure water. In the 500-700nm band range, the solution has better linearity. The whole process takes about 4 minutes and half minutes. The entire 96-well plate 8 was tested for 8 minutes. Compared with the traditional microplate reader, the speed is greatly improved.
It should be understood that the above examples are only for clarity of illustration and are not intended to limit the embodiments. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. This need not be, nor should all embodiments be exhaustive. And obvious variations or modifications of the invention may be made without departing from the scope of the invention.
Claims (6)
1. A full-spectrum high-efficiency microplate reader system, characterized by comprising:
the device comprises a full-spectrum light source, an ELISA plate, a detector for measuring the full-spectrum absorbance of a sample transilluminated by the full spectrum in a hole of the ELISA plate, and a motion control module for controlling the movement of the ELISA plate relative to the full-spectrum light source.
2. The full-spectrum high-efficiency microplate reader system of claim 1, wherein said detector is a spectrometer.
3. The full-spectrum high-efficiency microplate reader system of claim 1, wherein the motion control module comprises a stepping motor, a master control circuit, a power supply, a driver, two sets of guide rails and sliders, a coupler, an microplate holder for fixing the microplate, an upper computer and a sensor; the two groups of guide rails and the slide blocks are respectively arranged in the X direction and the Y direction, the ELISA plate support is fixed on the slide blocks of one group of guide rails and slide blocks, and the bottoms of the group of guide rails and slide blocks are fixed on the slide blocks of the other group of guide rails and slide blocks through the shaft coupling. The sensor is arranged at the original point position in the X and Y directions and is used for locating the original point of the ELISA plate.
The main control circuit is used for receiving a motion instruction sent by the upper computer and sending the motion instruction to the driver, the driver drives the stepping motor to control the sliding block to move on the guide rail, and the power supply provides electric energy for the driver.
And the upper computer sends out a motion instruction according to the current positions and preset positions of the two groups of guide rails and the slide blocks.
4. The full-spectrum high-efficiency microplate reader system of claim 3, wherein the upper computer is further connected with the detector for displaying current waveform data and storing the current data.
5. The full-spectrum high-efficiency microplate reader system according to claim 3, wherein the master control circuit is integrated with a control chip, a serial port chip, a key and a signal output circuit, the signal output circuit is connected with the driver, the control chip adopts STM32, and the serial port chip adopts CH 340.
6. The full-spectrum high-efficiency microplate reader system of claim 1, further comprising two collimating mirrors, connected to the full-spectrum light source and the detector through optical fibers, respectively, and arranged opposite to each other.
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| Application Number | Priority Date | Filing Date | Title |
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| CN202110815905.4A CN113552337A (en) | 2021-07-19 | 2021-07-19 | Full-spectrum high-efficiency microplate reader system |
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| CN202110815905.4A CN113552337A (en) | 2021-07-19 | 2021-07-19 | Full-spectrum high-efficiency microplate reader system |
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2021
- 2021-07-19 CN CN202110815905.4A patent/CN113552337A/en active Pending
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Application publication date: 20211026 |
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