CN103412029B - Tablet electrochromatography separation for amino acids device based on chip level and its application method - Google Patents

Abstract

The invention relates to a chip-level flat plate electrochromatographic amino acid separation device and its use method, which includes a CCD camera and a fluorescence microscope imaging system, an electrode module, a silica gel chromatography plate, a bottom support component and a power supply; the electrode module is placed horizontally on The two ends of the silica gel chromatography plate and the bottom support part are used to fix the chromatography plate and the electrode module, which constitute the carrier for separation and analysis; the CCD camera and the fluorescence microscope imaging system are placed directly above the overall device, and the power supply is connected to the electrode module through the power line middle. The positive effects of the present invention are: simple and low device cost, portable device size; and it is proved by experiments that the method is stable, reliable, effective and highly compatible; by introducing an electric field in the original one-dimensional chromatography process The effect turns the whole process into a two-dimensional separation, and the principle of chromatographic separation and electrophoretic separation are combined and acted on the separation and analysis of amino acids at the same time.

Description

Chip-level plate electrochromatographic amino acid separation device and its application method

【Technical field】

The invention relates to the technical field of amino acid separation devices, in particular to a chip-level flat plate electrochromatographic amino acid separation device and its application method.

【Background technique】

Amino acids are the most basic substances that constitute biological proteins and are related to life activities, and are closely related to life activities of organisms. As the basic unit of protein molecules, amino acids play an extremely important research role in the detection of protein sequences in genetic engineering. (Henikoff, S.; Henikoff, J.; 1992, Proc. Natal. Acad. Sci. USA, 89, 10915-10919). Since the importance of amino acids has been widely concerned, scholars at home and abroad have invested a lot of scientific research and attention on the separation and analysis of amino acids, and have made a lot of progress. For example, the determination of amino acids by near-infrared method (Zhao Chen; Qu Haibin; 2004, Spectroscopy and Spectral Analysis, 24), high-performance liquid phase method (Shi Chunzhen; Zhang Hongman; 2012, Analytical Chemistry, 4), rapid detection method of amino acid automatic analyzer, etc. ( Jiang Tao; Feng Yongjian; 2012, Chemical Application and Research, 7) etc. Most of the methods and instruments rely on commercial large-scale devices for analysis and testing, and there are strict requirements for instrument requirements, testing environment, sample purity and many other conditions. Some methods are even cumbersome to operate, and the detection cost is relatively high. Therefore, it is necessary to propose a simple and fast reliable method for the separation and analysis of amino acids based on low-cost chip technology.

At present, there are some analytical methods that can simply and quickly determine a certain specific amino acid in a complex sample, such as the combined technique of capillary electrophoresis (Schulta C.L., Moini M.2003, Anal.Chem., 75, 1508-1513) and Mass spectrometry (Soga T., Heiger D.N., 2000, Anal. Chem., 75, 1508-1513). Although these methods can accurately determine amino acids, their specificity and practicality can be interfered by one or more external factors, so the detection throughput of amino acids is not very high. In addition, these methods often require expensive instruments and equipment, complicated operation and long measurement time.

After more than ten years of rapid development, chip technology has been widely used in many fields of analytical chemistry. Such as isoelectric focusing IEF chip (Cui H.C., Keisuke H.2005, Anal.Chem., 77, 1303-1309) and capillary electrophoresis CE chip (Carlo S.E., Gerard J.M.B., 1997, Anal.Chem., 69, 3451-3457 ). The above-mentioned chip technology is aimed at the analysis and detection of capillary tubes, and has relatively strict and specific requirements for detectors, such as fluorescence detection or ultraviolet detection. Both need to use photomultiplier tubes or special CCDs to collect signals and convert data information. Moreover, in the field of chips, there is still a relatively large gap in the research on analytical chips in the field of chromatography.

【Content of invention】

The purpose of the present invention is to overcome the deficiencies of the prior art, and provide a chip-based flat plate electrochromatographic amino acid separation device and its application method.

The purpose of the present invention is achieved through the following technical solutions:

A chip-level flat-panel electrochromatographic amino acid separation device, which includes a CCD camera and a fluorescence microscope imaging system, an electrode module, a silica gel chromatography plate, a bottom support component and a power supply; the electrode module is placed horizontally at both ends of the silica gel chromatography plate , the bottom supporting part is used to fix the chromatographic plate and the electrode module, which constitutes the carrier for separation and analysis; the CCD camera and the fluorescence microscope imaging system are placed directly above the overall device, and the power supply is connected to the electrode module through the power line.

The CCD camera and fluorescence microscope imaging system have 3-6 million pixels, 300-1000 DPI fluorescence image resolution, 1-4 square centimeter CCD imaging area, and 50-500 times magnification of the microscope platform.

The electrode module includes an electrode carrier made of PMMA polymethyl methacrylate with a size of 2 to 3 square centimeters, the electrode ports carved at both ends have a size of 1 to 2 square millimeters, and the metal platinum electrode wire has a size of 2 to 3 square centimeters. 30-60 mm.

The silica gel chromatographic plate includes a glass plate whose length is 30-40 square centimeters, the thickness of the silica gel coating of the solid chromatography silica gel is 0.1-0.2 mm thick, and the particle size of the silica gel is 200-300 mesh.

The bottom supporting part is a supporting bottom plate made of PMMA polymethyl methacrylate, which is used to fix the electrode module and the chromatographic plate, and the specification is 30-40 square centimeters.

The power supply is a 0-600V DC power supply device.

A method for using a chip-level flat-panel electrochromatographic amino acid separation device, the specific steps of which are that the electrode modules with built-in platinum metal electrode wires are horizontally placed on both ends of the glass silica gel chromatography plate, and the bottom support parts are used for Fix the chromatographic plate and the electrode module to form the carrier for separation and analysis; put the sample into the sample loading area of the silica gel plate by point loading; the CCD camera and the fluorescence microscope imaging system are placed directly above the overall device, and the power supply is connected to the electrode through the power line In the module; during use, the lower end of the silica gel chromatography plate is immersed in a 5-10mm deep buffer solution, and an electric field is introduced to achieve the purpose of two-dimensional separation during chromatography.

This chip-scale device can be used for two-dimensional separation of various fluorescently labelable amino acids.

Compared with prior art, positive effect of the present invention is:

First, simple and low device cost, portable device size.

Second, it is proved by experiments that the method is stable, reliable, effective, and highly compatible.

Third, the whole process becomes a two-dimensional separation by introducing an electric field into the original one-dimensional chromatography process. The principles of chromatographic separation and electrophoretic separation act together and simultaneously in the separation and analysis of amino acids.

【Description of drawings】

Fig. 1 is a schematic diagram of the principle of the present invention;

Fig. 2 is a device structural diagram of the present invention;

Fig. 3 is the separation result figure of the present invention, what No. 1 picture illustrates is the moving band of the sample under the electric field; What No. 2 picture illustrates is to carry out buffer solution siphon effect first, add electric field subsequently, the electrophoresis of sample on silica gel plate Moving state; Picture No. 3 shows the state of the electrophoretic bands of the components to be separated under the simultaneous action of siphon and electric field; Picture No. 4 is a repetitive experiment, and the conditions are the same as those of Picture No. 3.

The marks in the drawings are: 1 CCD camera and fluorescence microscope imaging system, 2 Capillary sample loading part, 3 Electrode module, 4 Silica gel chromatography plate, 5 Sample sample loading port, 6 Bottom support part.

【Detailed ways】

The following provides a specific embodiment of a chip-based plate electrochromatographic amino acid separation device and its usage method of the present invention.

Example 1

A chip-level flat-panel electrochromatographic amino acid separation device, which includes a CCD camera and a fluorescence microscope imaging system, an electrode module, a silica gel chromatography plate, a bottom support component and a power supply; the electrode module is placed horizontally at both ends of the silica gel chromatography plate , the bottom supporting part is used to fix the chromatographic plate and the electrode module, which constitutes the carrier for separation and analysis; the CCD camera and the fluorescence microscope imaging system are placed directly above the overall device, and the power supply is connected to the electrode module through the power line.

The CCD camera and fluorescence microscope imaging system have 3-6 million pixels, 300-1000 DPI fluorescence image resolution, 1-4 square centimeter CCD imaging area, and 50-500 times magnification of the microscope platform.

The electrode module includes an electrode carrier made of PMMA polymethyl methacrylate with a size of 2 to 3 square centimeters, the electrode ports carved at both ends have a size of 1 to 2 square millimeters, and the metal platinum electrode wire has a size of 2 to 3 square centimeters. 30-60 mm.

The silica gel chromatographic plate includes a glass plate whose length is 30-40 square centimeters, the thickness of the silica gel coating of the solid chromatography silica gel is 0.1-0.2 mm thick, and the particle size of the silica gel is 200-300 mesh.

The bottom supporting part is a supporting bottom plate made of PMMA polymethyl methacrylate, which is used to fix the electrode module and the chromatographic plate, and the specification is 30-40 square centimeters.

The power supply is a 0-600V DC power supply device.

A method for using a chip-level flat-panel electrochromatographic amino acid separation device, the specific steps of which are that the electrode modules with built-in platinum metal electrode wires are horizontally placed on both ends of the glass silica gel chromatography plate, and the bottom support parts are used for Fix the chromatographic plate and the electrode module to form the carrier for separation and analysis; put the sample into the sample loading area of the silica gel plate by point loading; the CCD camera and the fluorescence microscope imaging system are placed directly above the overall device, and the power supply is connected to the electrode through the power line In the module; during use, the lower end of the silica gel chromatography plate is immersed in a 5-10mm deep buffer solution, and an electric field is introduced to achieve the purpose of two-dimensional separation during chromatography.

As shown in Figure 1, this embodiment includes: a CCD camera 1, a capillary sample loading part 2, an electrode module 3, a silica gel chromatography plate 4, a sample loading port 5, and a bottom support part 6.

As shown in Figure 2, the length of the electrode module is shown in the separation schematic diagram; the direction of the buffer siphon action is described in the black dotted box; the colored arrows simulate the difference in the electrophoretic direction of various charged substances under the action of an electric field; the top marks Properties of the applied electric field.

As shown in Figure 3, picture No. 1 shows the moving band of the sample without an electric field; picture No. 2 shows the electrophoresis movement state of the sample on the silica gel plate after siphoning the buffer solution first, and then adding an electric field; Picture No. 1 shows the state of electrophoretic bands of the components to be separated under the simultaneous action of siphon and electric field; picture No. 4 is a repetitive experiment, and the conditions are the same as picture No. 3.

The above is only a preferred embodiment of the present invention, it should be pointed out that for those of ordinary skill in the art, without departing from the concept of the present invention, some improvements and modifications can also be made, and these improvements and modifications should also be considered Within the protection scope of the present invention.

Claims (2)

1. A flat plate electrochromatographic amino acid separation device based on chip level, which includes a CCD camera and a fluorescence microscope imaging system, an electrode module, a silica gel chromatography plate, a bottom support component and a power supply; it is characterized in that the electrode module is placed horizontally on the silica gel The two ends of the chromatographic plate and the bottom supporting part are used to fix the chromatographic plate and the electrode module, which constitute the carrier for separation and analysis; the CCD camera and the fluorescence microscope imaging system are placed directly above the overall device, and the power supply is connected to the electrode module through the power line ; The method for using the chip-level plate electrochromatographic amino acid separation device, the specific steps are: the electrode module with built-in platinum metal electrode wire is horizontally placed on the two ends of the glass silica gel chromatography plate, and the bottom support part is used to fix the chromatography plate and The electrode module constitutes the carrier for separation and analysis; the sample is loaded into the sample loading area of the silica gel plate; the CCD camera and the fluorescence microscope imaging system are placed directly above the overall device, and the power supply is connected to the electrode module through the power line; the use process Immerse the lower end of the silica gel chromatography plate in a buffer solution with a depth of 5 to 10 mm, and introduce an electric field to achieve the purpose of two-dimensional separation during chromatography; The CCD camera and fluorescence microscope imaging system have pixels of 3 to 6 million, fluorescence image resolution of 300 to 1000 DPI, CCD imaging area of 1 to 4 square centimeters, and microscope platform magnification of 50 to 500 times; The electrode module includes an electrode carrier made of PMMA polymethyl methacrylate with a size of 2 to 3 square centimeters, the electrode ports carved at both ends have a size of 1 to 2 square millimeters, and the metal platinum electrode wire has a size of 2 to 3 square centimeters. 30-60 mm; The silica gel chromatography plate includes a glass plate with a size of 30-40 square centimeters, a silica gel coating thickness specification of 0.1-0.2 mm thick for solid chromatography silica gel, and a silica gel particle size of 200-300 mesh; The bottom supporting part is a supporting bottom plate made of PMMA polymethyl methacrylate, which is used to fix the electrode module and the chromatographic plate, and the specification is 30-40 square centimeters. 2 . The chip-level electrochromatography amino acid separation device based on claim 1 , wherein the power supply is a 0-600V DC power supply device. 3 .