CN1187601C - Micro fluid control chip detecting system - Google Patents

Abstract

The present invention discloses a micro-fluid control chip detection system which comprises an optical part, an interface circuit used for processing electrical signals output by a photomultiplier tube and an embedded system with display, storage and printing functions, wherein the optical part comprises a laser excitation unit and a detection unit; the laser excitation unit is vertical to an optical axis of the detection unit. Through a signal processing and data conversion unit, the embedded system is connected to the detection unit and controls electrophoresis voltage and the negative high-voltage output of the photomultiplier tube. The whole detection unit adopts an inverse structure, and an optical system keeps still with a folded optical path so as to realize the miniaturization of the whole system. In addition, the detection system is convenient for placing chips and regulating chip positions, meets the confocal principle to obtain signals with high a signal-to-noise ratio, and can be independently operated or can communicate with a computer.

Description

Microfluidic chip detection system

Technical field

The invention relates to a microfluidic chip detection system.

Background technique

Micro Total Analysis Systems (μ-TAS) is a new interdisciplinary field, whose goal is to realize the overall miniaturization of chemical analysis systems from sample processing to detection by means of micro-electromechanical processing (MEMS) technology and biotechnology , Integration and portability. It has become an important direction and frontier in the development of analytical instruments.

The current micro-full analysis system can be divided into two categories: chip type and non-chip type. At present, the chip type is the focus of development, which can be divided into two categories according to the chip structure and working mechanism: microarray chip (Microarray chip) and microfluidic chip (Microfluidic chip). Although there is a small amount of overlap between the two technologies, they have basically experienced their own development process.

Microarray chips, also known as biochips, are mainly based on biotechnology, with affinity binding technology as the core, and a series of addressable recognition molecule arrays immobilized on the chip surface as structural features. It is easy to use and fast to measure, but it is generally used for one-time use and has strong specificity. Another type of chip, the microfluidic chip, is mainly based on chemical analysis and analytical biochemistry, supported by micro-electromechanical processing technology, and characterized by a micro-pipeline network, which is the focus of the development of the current micro-total analysis system. It integrates sampling, dilution, reagent addition, reaction, separation, etc. on the chip, and can be used multiple times, so it has wider applicability.

The emergence of the microchip analysis system can not only greatly reduce the consumption of precious biological samples and reagents to microliters or even nanoliters, but also increase the analysis speed by ten times and a hundred times, and reduce the cost by ten times and a hundred times.

Biomedicine is the main application field of the current microfluidic analysis system. For the research on the relationship between human genes and diseases, the development of capillary electrophoresis suitable for single nucleotide polymorphism (SNP) detection, DNA sequencing and protein sequencing in the post-genome era Microfluidic chips are a top priority. Microfluidic chips used for clinical testing will have the most extensive market in my country. The synthesis and screening of new drugs is another area where microfluidic chips can play an important role. Other important application areas include food and commodity inspection, environmental monitoring, criminal science and aerospace science.

The microfluidic chip targeted by the present invention is an integrated capillary electrophoresis chip, and its detection methods include ultraviolet absorption detection, fluorescence detection, mass spectrometry detection, amperometric detection and the like. Among them, fluorescence detection, also known as Laser Induced Fluorescence (LIF) detection method, is used to detect compounds that can fluoresce, and is a highly sensitive and selective detection method. Certain compounds with special structures can emit light with a wavelength longer than that of ultraviolet light or laser after being irradiated by ultraviolet light or laser, generally in the range of visible light. This light is called fluorescence, and ultraviolet light or laser with a shorter wavelength becomes the excitation The resulting fluorescence is called emitted light. A fluorescence detector is a system for measuring fluorescence intensity. When the experimental conditions are fixed, the fluorescence intensity has a linear relationship with the sample concentration. Because the fluorescence detection method directly measures the fluorescence intensity and has high sensitivity, it is especially suitable for trace analysis, and the equipment is relatively simple. Therefore, when considering the detection method of capillary electrophoresis type chip, the fluorescence detection method is usually selected.

Figure 1 is a schematic diagram of a fluorescence detector. The laser light emitted by the laser 1 is converged by the lens 2 to generate a light spot at a fixed position of the capillary channel of the microfluidic chip 3 . When the fluid in the capillary flows through the position of the light spot, the fluorescent substance carried in it is excited by the laser and emits fluorescence. These fluorescent signals are collected by the microscope objective lens 4, and the unnecessary wavelength information is filtered out by the emission narrow-band color filter 5, and the received fluorescent signal is converted into an electrical signal by the photomultiplier tube 6 (PMT), which is output to subsequent equipment to display.

There are three kinds of optical paths for concrete realization: oblique incident type, transmissive type and reflective type detection optical path. The oblique-incidence optical path was mostly used in the early stage. The laser beam is obliquely incident at a certain angle. The fluorescent signal generated by the fluorescent material flowing through the laser spot is directly collected by the objective lens, processed and displayed by the photomultiplier tube. This structure is relatively simple, but it is inconvenient to adjust the angle, and the signal-to-noise ratio of the signal is not high, so it is rarely used now.

Figure 2 is a schematic diagram of the reflective optical path. The structure of the transmissive detection optical path is similar to that of the reflective detection optical path. The biggest difference between the two is that the transmissive optical path does not use a dichromatic mirror, and the loss of light energy is low. In the transmissive optical path, the laser light enters from the bottom of the chip through the reflector and the converging lens, and the main objective lens collects the fluorescence emitted from the designated area on the upper part of the chip. Build up the light path.

In the reflective optical path ( FIG. 2 ), the laser beam emitted by the laser 1 is reflected by the dichromatic mirror 7 , and under the action of the main objective lens 8 , a converging spot is generated at a fixed position of the microfluidic chip 3 . During electrophoresis, fluorescent substances flow through the laser spot and are excited to generate fluorescent signals. These scattered fluorescent signals are collected by the main objective lens 8, passed through the dichromatic mirror 7, converged by the lens tube lens 9, filtered by the color filter 5 to filter non-fluorescent signals, received by the photomultiplier tube 6 and converted into electrical signals. The electrical signal generated by the photomultiplier tube 6 is sent to the computer for recording and display through subsequent processing such as electronic filtering. The reflective optical path structure uses the dichromatic mirror 7 to realize the partial folding of the laser introduction optical path and the fluorescent signal optical path. The main objective lens 8 not only plays the role of converging the laser beam, but also can collect the fluorescent signal, so that the volume of the entire system is reduced and the miniaturization of the instrument is achieved. requirements. However, the dichroic mirror brings a certain degree of light energy loss and reduces the signal. Therefore, confocal technology is considered to improve the signal-to-noise ratio of the signal. In this way, even if the system loses a part of light energy, better quality images can still be obtained. Signal.

Figure 3 is a schematic diagram of the confocal principle. Confocal technology, that is, the light source, the object point and the point detector are in the conjugate position of the corresponding optical object image. The laser beam expanded and collimated by the laser 1 is reflected by the dichroic mirror 7, and then focused on the microfluidic chip 3 by the main objective lens 8 (the numerical aperture is NA) to form a diffraction-limited spot (spot diameter d=1.22*λ/NA) , the fluorescence generated by excitation is collected by the main objective lens 8, passed through the dichroic mirror 7, and imaged on the plane of the spatial filter 10 (detection pinhole) through the lens tube lens 9, and received by the photodetector near the image plane. Reflected (scattered) signal from chip 3. In the confocal technology, only the reflection (scattering) signal of the focal plane of the microfluidic chip 3 is received through the filtering effect of the detection pinhole 10 , and the reflection (scattering) signal of the non-focal plane of the microfluidic chip 3 is filtered out. It can be seen that the biggest advantage of confocal technology for microfluidic chip detection systems is that it can effectively reduce noise and improve signal quality. Compared with the ordinary optical path, the biggest feature of the introduction of confocal technology is the use of the detection pinhole 10, its size and position have a crucial impact on the performance of the entire system, both theoretically and experimentally. It embodies this.

Contents of Invention

The purpose of the present invention is to provide a microfluidic chip detection system that introduces confocal technology and adopts an inverted structure for optical components.

The technical scheme that the present invention adopts is as follows:

It includes optical components, used to process the electrical signal output by the photomultiplier tube, an interface circuit with functions of amplification, filtering, A/D sampling, and D/A control of the output voltage of the high-voltage module, and an embedded circuit with display, storage, and printing functions. formula system. Said optical components are composed of a laser excitation unit and a detection unit, the laser excitation unit is perpendicular to the optical axis of the detection unit; the laser excitation unit includes a semiconductor laser, and the excitation light narrow-band color filter connected on the same optical axis from right to left , a beam expander objective lens and a collimator objective lens; the detection unit includes dichroic mirrors inclined on the same optical axis, the main objective lens is installed above the dichromatic mirror, and the lens tube lens is installed sequentially below the dichromatic mirror from top to bottom , Rotating reflector, eyepiece, detection pinhole, emission narrow-band color filter and photomultiplier tube.

The beneficial effects that the present invention has are:

The laser excitation unit is perpendicular to the optical axis of the detection unit. The embedded system is connected to the detection unit through the signal processing and data conversion unit, and also controls the electrophoretic voltage and the negative high voltage output of the photomultiplier tube through the signal processing and data conversion unit. The entire detection unit adopts an inverted structure, and the optical system remains still. Not only the optical path is folded, the miniaturization of the entire system is realized, but also it is convenient to place the chip, adjust the position of the chip, meet the confocal principle, and obtain a signal with a high signal-to-noise ratio. Can operate independently or communicate with a computer.

Description of drawings

Figure 1 is a schematic diagram of the principle of fluorescence detection;

Fig. 2 is a schematic diagram of the detection optical path of the reflective microfluidic chip;

Figure 3 is a schematic diagram of the confocal principle;

Fig. 4 is the schematic diagram of parallel light optical path of the present invention;

Fig. 5 is the overall frame diagram of circuit system;

FIG. 6 is a refinement of the circuit in the small dashed box I in FIG. 5 .

Explanation of symbols in the figure:

1-semiconductor laser 2-lens

3-Microfluidic chip 4-Microscopic objective lens

5-Narrow-band emission filter 6-Photomultiplier tube

7-Dichromatic mirror 8-Main objective lens

9-Tube lens 10-Detection pinhole

11-Excitation light narrow-band color filter 12-Beam expander objective lens

13-collimating objective lens 14-mirror

15-eyepiece

Detailed ways

As shown in Figure 4, it includes optical components, used to process the electrical signal output by the photomultiplier tube, an interface circuit with functions of amplification, filtering, A/D sampling, and D/A control of the output voltage of the high-voltage module, and a display, Embedded system with storage and printing functions. Said optical components are composed of a laser excitation unit and a detection unit, and the laser excitation unit is perpendicular to the optical axis of the detection unit; the laser excitation unit includes a semiconductor laser 1, and the excitation light is sequentially connected on the same optical axis from right to left. Sheet 11, beam expander objective lens 12 and collimator objective lens 13; The detection unit includes the dichroic mirror 7 that is inclined on the same optical axis, and the main objective lens 8 is housed above the dichromatic mirror 7, and the bottom of the dichroic mirror 7 is viewed from above. And down, lens barrel lens 9, rotatable reflector 14, eyepiece 15, detection pinhole 10, emission light narrow-band color filter 5, photomultiplier tube 6 are housed successively.

The semiconductor laser 1 can be a laser with an excitation wavelength of 635nm, or multiple lasers with different wavelengths, which are coupled to the optical path of the laser excitation unit by switching brackets. The dichromatic mirror 7 reflects light with a wavelength of 635±410nm, and transmits light with a wavelength above 670nm. The magnification of the main objective lens 8 is 25-40, and the numerical aperture is 0.4-0.65. The diameter of the detection pinhole 10 is 200-500 μm.

The present invention adopts the optical path of the structure shown in FIG. 4 . Based on the principle in Figure 2, an inverted structure is adopted, combined with confocal technology, to make its structure more reasonable and more in line with actual needs. The laser beam emitted by the laser 1 is expanded by the beam expanding objective lens 12 through the excitation light narrow-band color filter 11 and collimated by the collimating objective lens 13, and then reflected by the dichromatic mirror 7, and the main objective lens 8 directs the beam on the microfluidic chip 3 Converge into a spot of light. Fluorescent substances flow through this light spot, and the fluorescence generated by excitation is still collected by the main objective lens 8, and focused through the dichroic mirror 7 through the lens tube lens 9. If the reflector 14 is in the solid line position shown in Figure 4, the fluorescence is reflected by the reflector 14 and enters the eyepiece 15. The operator can directly observe through the eyepiece 15 whether fluorescence is generated and adjust the specific position of the spot of the laser beam on the chip; If the reflector 14 is switched to the dotted line position shown in Figure 4, the fluorescent signal passes through the detection pinhole 10 to filter the non-detection surface information, then passes through the emission narrow-band color filter 5 to filter the non-fluorescence wavelength signal, and finally is received by the photomultiplier tube 6 .

In the present invention, an inverted structure is adopted, and the microfluidic chip is placed on a special platform, which can move in the X and Y directions, which makes it more convenient to install the chip, and is also conducive to the alignment adjustment of the electrophoretic electrode position. The introduction of the reflector 14 can not only ensure that the photomultiplier tube receives the fluorescent signal, but also help the operator to observe whether the fluorescent signal is generated, whether the position of the light spot is wrong, and whether the experiment is normal.

The circuit part includes signal processing and data conversion unit (interface circuit) and embedded system unit. The block diagram of the circuit part is shown in Figure 5 and Figure 6. The signal processing and data conversion unit mainly amplifies, filters and samples the electrical signal output by the photomultiplier tube 6 . In addition, it also includes the negative high voltage control of the photomultiplier tube 6 and the output high voltage control of the chip electrophoresis high voltage module. In order to meet the requirements of instrumentation, miniaturization and portability, an embedded system is adopted, combined with a liquid crystal display and a dedicated liquid crystal drive circuit, so that the instrument itself has the functions of display, storage, and printing, and can be used alone, or a standardized communication interface can be used with a computer. Interconnection, further testing results can be transmitted through the Internet, and the operation is more convenient.

Its working principle is as follows: the photomultiplier tube 6 converts the collected optical signals into electrical signals. After pre-amplification and filtering, these electrical signals are A/D sampled by the data acquisition card, and the results are sent to the embedded system for storage or show. The embedded system sends a signal according to the instruction, and controls the output of the high-voltage package of the photomultiplier tube 6 and the high voltage of the electrophoresis through D/A conversion to adjust the performance of the photomultiplier tube and the progress of the electrophoresis.

In the circuit module, the specific diagram of the signal channel, that is, the preamplification and electronic filtering part is shown in Figure 7. The output of the photomultiplier tube 6 is a current signal, which is converted into a voltage signal by the current-voltage conversion module and amplified, then passes through the buffer follower, and then enters the instrument amplifier for secondary amplification, and is filtered by a voltage-controlled filter. According to the performance of the selected data acquisition card, the filtered signal is subjected to corresponding voltage bias processing, and is collected and converted by the data acquisition card through the buffer follower.

Claims (1)

1. fluidic chip detecting system, it comprises optics, be used for handling the electric signal of photomultiplier output, interface circuit with amplification, filtering, A/D sampling and D/A control high-pressure modular output voltage function, with embedded system with demonstration, storage, printing function, it is characterized in that: said optics is made up of laser excitation unit and probe unit, and the laser excitation unit is vertical with the optical axis of probe unit; The laser excitation unit comprises semiconductor laser (1), the exciting light narrow-cut filter (11) that connects at same optical axis successively from a right left side, expansion bundle object lens (12) and collimator objective (13); Probe unit comprises dichroscope tilting on the same optical axis (7), in the top of dichroscope (7) principal goods mirror (8) is housed, the catoptron (14) that below at dichroscope (7) is equipped with tube lens (9) from top to down successively, can be rotated, eyepiece (15), detecting pinhole (10), emission light narrow-cut filter (5), photomultiplier (6).

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