CN112011455A - Temperature field and light path double-channel detection device for real-time fluorescence quantitative PCR instrument - Google Patents
Temperature field and light path double-channel detection device for real-time fluorescence quantitative PCR instrument Download PDFInfo
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- CN112011455A CN112011455A CN202010966729.XA CN202010966729A CN112011455A CN 112011455 A CN112011455 A CN 112011455A CN 202010966729 A CN202010966729 A CN 202010966729A CN 112011455 A CN112011455 A CN 112011455A
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L7/00—Heating or cooling apparatus; Heat insulating devices
- B01L7/52—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples
Abstract
The invention provides a temperature field and light path double-channel detection device for a real-time fluorescence quantitative PCR instrument, which comprises a temperature data acquisition module connected with a structural circuit board, wherein a standard light source capable of simulating fluorescence brightness change is arranged between the temperature data acquisition module and the structural circuit board; the standard light source is wrapped by a light-permeable shell and is connected with the temperature acquisition module through the shell to form a detection end together. The device can collect the temperature in the porous metal heating module hole of the real-time fluorescence quantitative PCR instrument and control the brightness of the standard light source according to the temperature cycle so as to simulate the brightness change of the fluorescent group amplification reaction.
Description
Technical Field
The invention relates to the technical field of detection of real-time fluorescence quantitative PCR instruments, in particular to a temperature field and light path double-channel detection device for a real-time fluorescence quantitative PCR instrument.
Background
The real-time fluorescent quantitative PCR technology is a method of adding fluorescent groups into a PCR reaction system, monitoring the whole PCR reaction process in real time by utilizing the accumulation of fluorescent signals, and finally carrying out quantitative analysis on an unknown template through a standard curve. At present, the real-time fluorescence quantitative PCR instrument is widely applied to the fields of clinical examination, inspection and quarantine, forensic identification, disease control, in-vitro diagnostic reagent manufacture and research and development, biotechnology product research and development and the like. The accuracy of the experimental result of the real-time fluorescent quantitative PCR instrument is mainly influenced by the temperature accuracy of the metal heating module and the reliability of the optical path system.
The prior art can realize the detection of the temperature field part of the real-time fluorescence quantitative PCR instrument, but aiming at the light path system, the detection method of a biological reagent test disc is mainly used by a chemical method, and the linear error of a sample is detected by adopting a plasmid DNA standard substance with known concentration; the method for calibration cannot explain the influence relationship of the temperature and fluorescence of the fluorescence quantitative PCR instrument on the final quantitative result, the error amount, the error source and the like; this method can only measure the combined linearity between wells, and the results obtained can only represent the average result between wells of a PCR instrument, and cannot represent the linearity of a single well.
At present, no physical method can simultaneously detect the temperature accuracy of a metal heating module of a real-time fluorescence quantitative PCR instrument and the reliability of an optical path system.
Disclosure of Invention
In order to solve the problems, the invention provides a temperature field and light path dual-channel detection device for a real-time fluorescence quantitative PCR instrument, which can acquire the temperature in a hole of a porous metal heating module of the real-time fluorescence quantitative PCR instrument by introducing a thermistor element and a standard light source, controls the luminance of the standard light source according to temperature cycle to simulate the luminance change of fluorophore amplification reaction, receives and detects the luminance change by the real-time fluorescence quantitative PCR instrument, and detects and analyzes the accuracy and the sensitivity of the temperature field and the light path system of the real-time fluorescence quantitative PCR instrument by combining the acquired temperature and the luminance of the standard light source through fluorescence detection data generated by the real-time fluorescence quantitative PCR instrument.
In order to achieve the purpose, the invention provides the following specific technical scheme:
a temperature field and light path double-channel detection device for a real-time fluorescence quantitative PCR instrument comprises a temperature data acquisition module connected with a structural circuit board, wherein a standard light source capable of simulating fluorescence brightness change is arranged between the temperature data acquisition module and the structural circuit board, the spectral range of the standard light source is preferably 320-780 nm, the peak wavelength is 400-500 nm, and more preferably the peak wavelength is 460 nm; the standard light source is wrapped by a light-permeable shell and is connected with the temperature acquisition module through the shell to form a detection end together.
Preferably, the outer contour of the detection end is matched with the inner contour in the porous metal heating module hole of the real-time fluorescent quantitative PCR instrument, so that the outer contour and the inner contour are effectively attached, and the accuracy of the acquired temperature data is guaranteed.
Preferably, a light-transmitting point which can enable light of the standard light source to pass through is arranged at the connection position of the detection end and the structural circuit board, and is used for receiving and detecting the light by the real-time fluorescence PCR instrument and generating fluorescence detection data.
Preferably, the structural circuit board comprises an acquisition circuit connected with the temperature data acquisition module, a storage circuit and a control circuit connected with the standard light source.
Preferably, the structural circuit board is further provided with a wireless communication module, and the wireless communication module is connected with the control circuit; wireless communication module adopts BlueTooth communication protocol, can with PC end or APP end communication, right temperature field and light path binary channels detection device start, set up, stop and read etc..
Preferably, a support bottom plate is further covered on the structural circuit board, and the structural circuit board is fixedly connected with the support bottom plate; the support base plate is preferably a carbon fiber base plate.
Preferably, the temperature data acquisition module comprises a thermistor, and the thermistor is fixedly connected with the shell through a heat-conducting sealant; the detection range of the thermistor is 0-120 ℃.
Preferably, the number of the detection ends is 7-15.
Preferably, a power supply is also included in communication with the structural circuit board.
The invention has the beneficial effects that:
(1) the system can simultaneously detect the temperature field and the light path system of the real-time fluorescence quantitative PCR instrument and analyze the correlation of the temperature field and the light path system.
(2) And when the temperature field detection is carried out, the embedding and excitation control of a traceable standard light source are adopted to simulate biochemical fluorescence reaction by a physical means so as to comprehensively evaluate the performance reliability of the real-time fluorescence quantitative PCR instrument.
(3) And analyzing the correlation of the temperature field and the light path system of the real-time fluorescent quantitative PCR instrument. Finally, detection results of the fluorescence quantitative PCR instrument such as temperature accuracy, inter-pore temperature difference, temperature overshoot, temperature rising and falling rate, Ct value indicating error, Ct value uniformity, Ct value precision, channel peak height consistency, linear sensitivity coefficient, melting temperature drift, melting temperature ratio and the like can be obtained.
(4) The device can be suitable for the detection of temperature fields and light path systems of real-time fluorescent quantitative PCR instruments of various types, provides technical support for the accuracy and reliability of the real-time fluorescent quantitative PCR instruments in the fields of clinical examination, inspection and quarantine, forensic identification, disease control, in-vitro diagnostic reagent manufacturing and research and development, biotech product research and development and the like, and provides a standard device for a metering mechanism to carry out detection and calibration of temperature and fluorescence.
Drawings
FIG. 1 is a schematic structural diagram of a preferred embodiment of the present application;
FIG. 2 is a sectional view taken along line A-A of FIG. 1;
FIG. 3 is a schematic view of the structure of the detection end in a preferred embodiment of the present application;
fig. 4 is a schematic deployment diagram of the present application in practical use.
Wherein, 1-structure circuit board; 2-supporting the bottom plate; 3-detection end, 31-thermistor, 32-standard light source, 33-shell and 34-light-transmitting point; 4-screws.
Detailed Description
As used in the specification and in the claims, certain terms are used to refer to particular components. As one skilled in the art will appreciate, manufacturers may refer to a component by different names. This specification and claims do not intend to distinguish between components that differ in name but not function. In the following description and in the claims, the terms "include" and "comprise" are used in an open-ended fashion, and thus should be interpreted to mean "include, but not limited to. The description which follows is a preferred embodiment of the present application, but is made for the purpose of illustrating the general principles of the application and not for the purpose of limiting the scope of the application. The protection scope of the present application shall be subject to the definitions of the appended claims.
Referring to fig. 1 to 4, in a preferred embodiment of the present invention, a dual-channel temperature field and optical path detection device for a real-time fluorescence quantitative PCR instrument is provided, which includes a structural circuit board 1, a supporting base plate 2, a detection end 3, a wireless communication module and a power supply. The detection end 3 comprises a thermistor 31 and a standard light source 32; the structural circuit board 1 comprises an acquisition circuit, a storage circuit and a control circuit.
Further, the structural circuit board 1 and the bottom surface of the supporting base plate 2 have the same shape and are fixed by screws 4, and the supporting base plate 2 is preferably made of a carbon fiber material.
Furthermore, the outer contour of the detection end 3 is matched with the inner contour in the porous metal heating module hole of the real-time fluorescent quantitative PCR instrument so as to ensure that the two are effectively attached; one end of the detection end 3 is a thermistor 31, the other end is a standard light source 32, the outer side of the standard light source 32 is wrapped by a shell 33 made of PE transparent materials, and the thermistor 31 and the shell 33 are fixed by heat conduction sealant. The detection end 3 is fixedly connected with the structural circuit board 1, and a light transmission point 34 which can enable light of the standard light source 32 to pass through is arranged at the connection position, so that the real-time fluorescence PCR instrument receives and detects the light and generates fluorescence detection data.
Further, the total height of the detection end 3 is 21.5-26.5mm, preferably 21.5 mm; the size of the thermistor 31 is 0.5mm, and the temperature measuring range is preferably 0-120 ℃; the spectrum range of the standard light source 32 is preferably 320-780 nm, the peak wavelength is 400-500 nm, and the peak wavelength is more preferably 460 nm; the diameter of the light transmission point 34 is phi 2-phi 3 mm. Specifically, the number of the detection terminals 3 is 7-15, and as shown in fig. 4, 7 dots and 15 dots are taken as an example.
Further, the acquisition circuit is connected with the thermistor 31, and the control circuit is connected with the standard light source 32 and the wireless communication module. Specifically, wireless communication module adopts BlueTooth communication protocol, can with PC end or APP end communication, right temperature field and light path binary channels detection device start, set up, stop and read etc..
Further, the power supply comprises a lithium battery, and the lithium battery is connected with the structural circuit board 1 through a data line to supply power for the device.
In this embodiment, when the dual-channel detection device for the temperature field and the optical path of the real-time fluorescence quantitative PCR instrument is used, the detection end 3 is inserted into the porous metal heating module hole of the real-time fluorescence quantitative PCR instrument, the temperature collected by the detection end 3 is transmitted through the wireless communication module, the light output of the standard light source 32 is controlled, so that the fluorescence change in the gene amplification process is simulated, the fluorescence change is received and detected by the real-time fluorescence quantitative PCR instrument, and the accuracy and the sensitivity of the temperature field and the optical path system of the real-time fluorescence quantitative PCR instrument are detected and analyzed by combining the fluorescence detection data generated by the real-time fluorescence quantitative PCR instrument and the brightness of the standard light source 32, so as to realize the comprehensive evaluation of the performance of the real-time fluorescence quantitative PCR instrument.
Compared with the prior art, the beneficial effect of this application lies in:
(1) the system can simultaneously detect the temperature field and the light path system of the real-time fluorescence quantitative PCR instrument and analyze the correlation of the temperature field and the light path system.
(2) And when the temperature field detection is carried out, the embedding and excitation control of a traceable standard light source are adopted to simulate biochemical fluorescence reaction by a physical means so as to comprehensively evaluate the performance reliability of the real-time fluorescence quantitative PCR instrument.
(3) And analyzing the correlation of the temperature field and the light path system of the real-time fluorescent quantitative PCR instrument. Finally, detection results of the fluorescence quantitative PCR instrument such as temperature accuracy, inter-pore temperature difference, temperature overshoot, temperature rising and falling rate, Ct value indicating error, Ct value uniformity, Ct value precision, channel peak height consistency, linear sensitivity coefficient, melting temperature drift, melting temperature ratio and the like can be obtained.
(4) The device can be suitable for the detection of temperature fields and light path systems of real-time fluorescent quantitative PCR instruments of various types, provides technical support for the accuracy and reliability of the real-time fluorescent quantitative PCR instruments in the fields of clinical examination, inspection and quarantine, forensic identification, disease control, in-vitro diagnostic reagent manufacturing and research and development, biotech product research and development and the like, and provides a standard device for a metering mechanism to carry out detection and calibration of temperature and fluorescence.
The foregoing description shows and describes several preferred embodiments of the present application, but as aforementioned, it is to be understood that the application is not limited to the forms disclosed herein, but is not to be construed as excluding other embodiments and is capable of use in various other combinations, modifications, and environments and is capable of changes within the scope of the application as described herein, commensurate with the above teachings, or the skill or knowledge of the relevant art. And that modifications and variations may be effected by those skilled in the art without departing from the spirit and scope of the application, which is to be protected by the claims appended hereto.
Claims (9)
1. A temperature field and light path double-channel detection device for a real-time fluorescence quantitative PCR instrument comprises a temperature data acquisition module connected with a structural circuit board, and is characterized in that a standard light source capable of simulating fluorescence brightness change is arranged between the temperature data acquisition module and the structural circuit board; the standard light source is wrapped by a light-permeable shell and is connected with the temperature acquisition module through the shell to form a detection end together.
2. The dual-channel detection device for the temperature field and the optical path of the real-time fluorescence quantitative PCR instrument as claimed in claim 1, wherein the outer contour of the detection end is matched with the inner contour in the hole of the porous metal heating module of the real-time fluorescence quantitative PCR instrument.
3. The apparatus of claim 2, wherein a light-transmitting point is disposed at the connection between the detection end and the structural circuit board for transmitting the light of the standard light source.
4. The apparatus of claim 3, wherein the structural circuit board comprises an acquisition circuit connected to the temperature data acquisition module, a storage circuit, and a control circuit connected to the standard light source.
5. The apparatus according to claim 4, wherein a wireless communication module is further disposed on the structural circuit board, and the wireless communication module is connected to the control circuit.
6. The apparatus of claim 5, wherein a support base plate is covered on the structural circuit board, and the structural circuit board is fixedly connected with the support base plate.
7. The dual-channel detection device for the temperature field and the optical path of the real-time fluorescence quantitative PCR instrument as recited in claim 6, wherein the temperature data acquisition module comprises a thermistor, and the thermistor is fixedly connected with the housing through a heat-conducting sealant.
8. The apparatus of claim 7, wherein the number of the detection ends is 7-15.
9. The apparatus of claim 8, further comprising a power supply connected to the structural circuit board.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010966729.XA CN112011455A (en) | 2020-09-15 | 2020-09-15 | Temperature field and light path double-channel detection device for real-time fluorescence quantitative PCR instrument |
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| CN202010966729.XA CN112011455A (en) | 2020-09-15 | 2020-09-15 | Temperature field and light path double-channel detection device for real-time fluorescence quantitative PCR instrument |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113528315A (en) * | 2021-08-03 | 2021-10-22 | 北京林电伟业电子技术有限公司 | Method for simulating gene amplification standard curve |
| CN113564232A (en) * | 2021-08-03 | 2021-10-29 | 北京林电伟业电子技术有限公司 | Method for simulating fluorescent group amplification reaction |
-
2020
- 2020-09-15 CN CN202010966729.XA patent/CN112011455A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113528315A (en) * | 2021-08-03 | 2021-10-22 | 北京林电伟业电子技术有限公司 | Method for simulating gene amplification standard curve |
| CN113564232A (en) * | 2021-08-03 | 2021-10-29 | 北京林电伟业电子技术有限公司 | Method for simulating fluorescent group amplification reaction |
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