Liquid auto-fractionator for column chromatography

Abstract

The present invention relates to a liquid automatic fractionation apparatus for column chromatography. According to the present invention, the liquid automatic fractionation apparatus for column chromatography includes a microtube installation portion in which a plurality of microtubes for accommodating a liquid to be fractionated according to column chromatography are installed, a liquid supply unit configured to sequentially supply the liquid to the plurality of microtubes, a light source unit provided at a rear side of the plurality of microtubes to emit light, an image acquisition unit provided at a front side of the plurality of microtubes to acquire images of the plurality of microtubes at a preset time interval while performing column chromatography, a volume determination unit configured to extract a height of a level of the accommodated liquid by analyzing the images of the plurality of microtubes acquired by the image acquisition unit, and configured to determine whether a volume of the liquid accommodated in the plurality of microtubes reaches a target volume based on the extracted height of the level, and a control unit configured to set a reference value for determining whether the volume reaches the target volume, and configured to control operations of the liquid supply unit and the image acquisition unit according to a determination result of the volume determination unit.

Description

TECHNICAL FIELD

[0001] The present application relates to a liquid automatic fractionation device for column chromatography, and more particularly, to a liquid automatic fractionation device for column chromatography that can more conveniently and accurately obtain a desired amount of liquid fraction by column chromatography. BACKGROUND

[0002] Column chromatography is a size-exclusion chromatography to classify substances mixed in a liquid mixture according to the size of the substances. Specifically, when a stationary phase (e.g., silica beads) is filled in a column and a mobile phase (e.g., plasma or cell culture solution, etc.) passes through the column, substances in the mobile phase are sequentially discharged into different fractions according to their size. In this way, only particles of a desired size are separated.

[0003] When a fraction is obtained by using column chromatography as described above, one usually visually checks whether a desired amount is fractionated. However, when the amount to be fractionated is very small (e.g., several μl), the possibility of an error is high, there are differences depending on the worker, and there is a limitation in that it is difficult to obtain a large amount of sample.

[0004] Although there are liquid automatic fractionation devices on the market, known devices mainly obtain a fraction based on the weight of a liquid, and thus there is a limitation in that it is difficult to fractionate a small amount of liquid in units of μl due to a large error. SUMMARY

[0005] TECHNICAL PROBLEM

[0006] According to the present application, there is provided a liquid automatic fractionation device for column chromatography that can more conveniently and accurately obtain a desired amount of liquid fraction by column chromatography.

[0007] SOLUTION TO PROBLEM

[0008] According to an embodiment of the present invention to achieve the technical object, there is provided a liquid automatic fractionation apparatus for column chromatography, including: a microtube installation portion in which a plurality of microtubes for accommodating a liquid to be fractionated according to the column chromatography are installed; a liquid supply unit configured to sequentially supply the liquid to the plurality of microtubes; a light source unit provided at a rear side of the plurality of microtubes to emit light; an image acquisition unit provided at a front side of the plurality of microtubes to acquire images of the plurality of microtubes at a preset time interval while the column chromatography is performed; a volume determination unit configured to extract a height of a level of the liquid accommodated by analyzing the images of the plurality of microtubes acquired by the image acquisition unit, and configured to determine whether a volume of the liquid accommodated in the plurality of microtubes reaches a target volume based on the extracted height of the level; and a control unit configured to set a reference value for determining whether the volume reaches the target volume, and configured to control operations of the liquid supply unit and the image acquisition unit according to a determination result of the volume determination unit.

[0009] The microtube installation portion can consist of a plurality of grooves formed in a housing to accommodate the plurality of microtubes, and the plurality of grooves can be arranged in a row at a preset interval on a straight line.

[0010] A reference microtube accommodating the liquid of the target volume can be installed in a first groove among the plurality of grooves, and the plurality of microtubes for fractionation can be installed in other grooves.

[0011] The liquid supply unit can include: a column including a stationary phase and a mobile phase; a column support connected to the housing so as to be movable along a linear axis parallel to a straight line in which the plurality of grooves are formed, and configured to support the column so that the column is sequentially movable in positions corresponding to the plurality of microtubes; and a driving unit connected to a lower end of the column support, and configured to move the column so that the column is located at an upper portion of one of the plurality of microtubes while the column chromatography is performed.

[0012] The image acquisition unit can include: a camera installed to be sequentially movable from the positions corresponding to the plurality of microtubes, and configured to acquire images of the plurality of microtubes; a moving axis parallel to the straight line in which the plurality of grooves are formed, and on which the camera moves; and a driving unit configured to move the camera on the moving axis so that the camera is located in front of the plurality of microtubes while the column chromatography is performed.

[0013] The volume determining unit can analyze the images of each of the plurality of microtubes obtained by the image acquisition unit at the preset time interval, and determine whether the volume reaches the target volume by using only the images in which the standard deviation of the height of the horizontal plane is less than or equal to a preset value.

[0014] The volume determining unit can find a boundary surface of a refracted image in the images of the plurality of microtubes, extract the height of the horizontal plane, and compare the height of the horizontal plane with the reference value to determine whether the volume reaches the target volume.

[0015] The volume determining unit can add all R channel values of pixels on a corresponding horizontal line in the images of each of the plurality of microtubes for each horizontal line, extract a position of the horizontal line in which the sum of the R channel values is the largest as the height of the horizontal plane, and determine that the volume reaches the target volume when the height of the horizontal plane exceeds the reference value.

[0016] The liquid automatic fractionation apparatus can further include a display unit implemented as a touch screen to display buttons for receiving signals related to the operation of the liquid automatic fractionation apparatus, visualize a performance state of the column chromatography as a graph, and display the visualized performance state.

[0017] The control unit can acquire and analyze the image of the reference microtube to extract the height of the horizontal plane, and set the extracted height as the reference value.

[0018] The control unit can check whether alignment of the plurality of microtubes and the light source unit is correct based on brightness of the images of the plurality of microtubes, and correct a position of the image acquisition unit when the alignment is not correct.

[0019] The control unit can add R channel values of respective pixels on a central vertical line in the images of each of the plurality of microtubes, and determine that alignment of the plurality of microtubes and the light source unit is correct when the added value is greater than or equal to a preset value.

[0020] Advantageous effects

[0021] As described above, according to the present application, column chromatography that is manually performed in the related art can be more accurately and conveniently performed by a mechanical device, and volume determination can be more accurately and conveniently performed by image analysis. In addition, in terms of performance, an accurate result corresponding to a person having ordinary skill in the art of column chromatography can be obtained. In addition, productivity and convenience can be ensured by automation, so that an accurate fraction of a desired sample can be more easily obtained. BRIEF DESCRIPTION OF DRAWINGS

[0022] Figure 1 is a configuration diagram of a liquid automatic fractionation apparatus for column chromatography according to an embodiment of the present invention.

[0023] Figure 2 is a diagram showing an embodiment example of a liquid automatic fractionation apparatus for column chromatography shown in Figure 1

[0024] Figure 3 is an operation diagram of a liquid automatic fractionation apparatus for column chromatography shown in Figure 2

[0025] Figure 4 is a flowchart showing an operation method of a liquid automatic fractionation apparatus for column chromatography shown in Figure 1

[0026] Figure 5 is a diagram showing an acquired image according to an alignment state of a light source and a microtube according to an embodiment of the present invention.

[0027] Figure 6 is a diagram showing a method of extracting a height of a horizontal plane through image analysis according to an embodiment of the present invention.

[0028] LIST OF REFERENCE NUMERALS

[0029] 100: liquid automatic fractionation apparatus

[0030] 110: microtube mounting portion

[0031] 120: liquid supply unit

[0032] 130: light source unit

[0033] 140: image acquisition unit

[0034] 150: volume determination unit

[0035] 160: control unit

[0036] 170: display unit DETAILED DESCRIPTION

[0037] Hereinafter, preferred embodiments according to the present invention will be described in detail with reference to the accompanying drawings. In this process, the thickness of lines or the size of components shown in the drawings can be exaggerated for the sake of description and convenience.

[0038] Further, terms to be described below are defined in consideration of functions in the present invention, and the terms can be changed according to the intention or habit of a user or an operator. Therefore, the terms must be defined based on the contents throughout the present specification. ​​​

[0039] Figure 1 is a configuration diagram of a liquid automatic fractionation apparatus for column chromatography according to an embodiment of the present invention, Figure 2 is a diagram illustrating Figure 1 an embodiment of a liquid automatic fractionation apparatus for column chromatography shown in FIG. 1, and Figure 3 is Figure 2 an operation schematic diagram of a liquid automatic fractionation apparatus for column chromatography shown in FIG. 1.

[0040] Referring to Figures 1 to 3 , a liquid automatic fractionation apparatus 100 for column chromatography according to an embodiment of the present invention can include a microtube mounting portion 110, a liquid supply unit 120, a light source unit 130, an image acquisition unit 140, a volume determination unit 150, a control unit 160, and a display unit 170.

[0041] The microtube mounting portion 110 is a portion on which a plurality of microtubes 10 that accommodate liquids to be fractionated according to column chromatography are mounted.

[0042] According to one embodiment, the microtube mounting portion 110 can have a plurality of grooves formed in a housing 101 of the apparatus to accommodate a plurality of microtubes 10 used in column chromatography, and the plurality of grooves can be linearly arranged in a row at a preset interval.

[0043] The liquid supply unit 120 performs column chromatography by sequentially supplying liquids to the plurality of microtubes 10 mounted on the microtube mounting portion 110.

[0044] According to one embodiment, the liquid supply unit 120 can be configured to include a column 121 that accommodates a stationary phase and a mobile phase, a column support 122 that supports the column 121, and a driving unit (not shown) connected to the column support 122 to move the column 121.

[0045] Specifically, as Figure 2 shown, the column 121 can be supported by the column support 122 so as to be sequentially moved at positions corresponding to the plurality of microtubes 10 mounted on the microtube mounting portion 110, respectively. The column support 122 can be connected to the housing 101 so as to be movable along a linear axis parallel to a straight line formed by the plurality of grooves constituting the microtube mounting portion 110. In addition, the driving unit (not shown) can be connected to a lower end of the column support 122 within the housing 101 to move the column 121 so that the column 121 supported by the column support 122 is located at an upper portion of the corresponding microtube when column chromatography is performed. The liquid in the mobile phase and stored in the column 121 falls through an outlet formed at a lower portion of the column 121 as column chromatography is performed to be accommodated in the microtube 10 located at the lower portion of the column 121.

[0046] The light source unit 130 can be disposed inside the housing 101 at the rear side of the plurality of microtubes 10 mounted on the microtube mounting portion 110 to emit light, and can be composed of, for example, a red light emitting diode (LED) light source, but is not limited thereto.

[0047] According to one embodiment, the light source unit 130 can be composed of a plurality of LED light sources, as shown in FIG. 2, the LED light sources being disposed at positions respectively corresponding to the plurality of microtubes 10 mounted on the microtube mounting portion 110, and can be composed of a single LED light source that is movable, the single LED light source being disposed at the rear side of the corresponding microtube in which column chromatography is performed. Figure 3

[0048] The image acquisition unit 140 is disposed inside the housing 101 at the front side of the plurality of microtubes 10 mounted on the microtube mounting portion 110 to acquire images of the plurality of microtubes 10 at a predetermined time interval while column chromatography is performed.

[0049] According to one embodiment, the image acquisition unit 140 can include a camera 141 for acquiring images of the plurality of microtubes 10, a movement shaft 142 on which the camera 141 moves, and a driving unit 143 that moves the camera 141 on the movement shaft 142.

[0050] Specifically, the camera 141 can be mounted to sequentially move along the movement shaft 142 at positions respectively corresponding to the plurality of microtubes 10 mounted on the microtube mounting portion 110. Here, the movement shaft 142 can be parallel to a straight line formed by the plurality of grooves constituting the microtube mounting portion 110. In addition, the driving unit 143 can move the camera 141 so that the camera 141 is located in front of the corresponding microtube while column chromatography is performed.

[0051] The volume determination unit 150 can analyze the images of each of the plurality of microtubes acquired by the image acquisition unit 140 to extract the height of the level of the liquid contained, and can determine whether the volume of the liquid contained in the microtube reaches a target volume based on the extracted height of the level.

[0052] In the present application, the height of the level can be extracted by utilizing a phenomenon in which light emitted by the light source unit 130 passes through the microtube 10 and is refracted by the liquid contained in the microtube 10, and it is determined whether the volume of the liquid reaches the target volume based on the extracted height of the level. In other words, when the liquid fills the microtube 10, the refraction pattern of the light changes, and the change in the refraction pattern is acquired as an image and analyzed to find the boundary surface of the refraction image and extract the height of the level, and thus it can be determined that the volume of the liquid reaches the target volume based on the extracted height of the level.

[0053] ​To this end, the volume determining unit 150 analyzes the images of each of the plurality of microtubes 10 acquired by the image acquiring unit 140 at a preset time interval. In this case, it can be difficult to accurately analyze the images of the microtubes 10 acquired at the time when the water droplets fall into the microtubes 10 or at the time when the water droplets adhere to the inner walls of the microtubes 10, and thus only stable images can be used for more accurate analysis. For example, assuming that the volume of one water droplet falling from the column 121 is about 20 μl and the water droplets fall at an interval of about 1 second, the volume determining unit 150 can receive the images of the plurality of microtubes 10 acquired by the image acquiring unit 140 at an interval of 0.1 second, and analyze the images to extract the heights of the horizontal planes, and use only the images of which the standard deviation of the extracted heights of the horizontal planes is less than or equal to a preset value.

[0054] Reference will now be made to Figure 4 The detailed description determines whether the volume determining unit 150 extracts the heights of the horizontal planes to determine whether the volume of the liquid reaches the target volume based on the extracted heights.

[0055] The control unit 160 controls Figure 1 all operations of the respective components of the liquid automatic fractionation apparatus 100 for column chromatography shown.

[0056] According to one embodiment, the control unit 160 can set a reference value for the volume determining unit 150 to determine whether the volume reaches the target volume, and can control the operations of the liquid supplying unit 120 and the image acquiring unit 140 according to the determination result of the volume determining unit 150.

[0057] In particular, as described below with reference to Figure 4 the control unit 160 can control the operations of the liquid automatic fractionation apparatus 100 for column chromatography according to embodiments of the present application.

[0058] The display unit 170 can display information on the operations of the liquid automatic fractionation apparatus 100 for column chromatography, and receive a signal related to the operations of the apparatus, and can be implemented as, for example, a touch screen.

[0059] According to one embodiment, the display unit 170 can display a button for receiving a signal related to the operations of the apparatus, and can further visualize the performance state of the column chromatography as a graph and display the visualized performance state.

[0060] Figure 4 is a flowchart illustrating Figure 1 the operation method of the liquid automatic fractionation apparatus for column chromatography shown in FIG. 1, Figure 5 is a graph illustrating the acquired images according to the alignment state of the light source and the microtube according to embodiments of the present application, and Figure 6is a diagram showing a method of extracting a height of a horizontal plane through image analysis according to an embodiment of the present application.

[0061] Referring to Figure 4 , a column chromatography is first prepared (S210). According to an embodiment, a column 121 accommodating a stationary phase and a mobile phase of the column chromatography is mounted on a column support 122, and a plurality of empty microtubes for fractionation is mounted on a microtube mounting portion 110. In this case, a first recess among a plurality of recesses constituting the microtube mounting portion 110 is left out as not to mount a microtube for fractionation.

[0062] Then, a reference value of a fraction is measured (S220). According to an embodiment, a reference microtube accommodating a target volume of liquid (e.g., water) to be fractionated by column chromatography is mounted in a first recess among a plurality of recesses constituting the microtube mounting portion 110, and when a "Ready" button displayed on the display unit 170 is touched, a height of a horizontal plane is extracted by acquiring and analyzing an image of the reference microtube, and the extracted height of the horizontal plane is set as a reference value (i.e., a target value).

[0063] Then, column chromatography is performed (S230). According to an embodiment, when a "Start" button displayed on the display unit 170 is touched and a cap of the column 121 is removed, the column 121 moves to an upper portion of a next microtube, and the column chromatography is performed. In this case, the camera 141 (and the LED light source if necessary) within the housing 101 also moves to a position corresponding to the next microtube, and an image of the microtube is acquired at a predetermined time interval as the column chromatography is performed.

[0064] Then, alignment of the microtube and the LED light source is checked (S240). When the centers of the microtube and the LED light source are not properly aligned, as Figure 5 indicated in the image shown on the left side, a liquid portion can be dark in the acquired image. Therefore, according to an embodiment, whether the alignment of the microtube and the LED light source is correct is checked based on the brightness of the acquired image, and when the alignment is not correct, the camera 141 can move left or right to correct the position of the camera 141, thereby adjusting the alignment of the microtube and the LED light source. For example, when the light source unit 130 is composed of a red LED light source, an image of a liquid portion is displayed in red, and thus, when Figure 5 the sum of R channel values of respective pixels on an extension line of a vertical line of the center of the acquired image (i.e., an extension line of a vertical line of the center of a black square frame displayed on the upper side of the image) is greater than or equal to a preset value, it can be determined that the alignment of the microtube and the LED light source is correct. Figure 5

[0065] ​Then, the volume is determined by analyzing the image of the microtube (S250). According to one embodiment, the volume determination unit 150 can find the boundary surface of the refracted image by means of image analysis, extract the height of the horizontal plane, and compare the extracted height with a reference value to determine whether the volume of the liquid reaches the target volume. For example, as shown in FIG. 16, the volume determination unit 150 can add all R channel values of the pixels on the respective horizontal line of the image for each horizontal line, and extract the position (value on the y-axis in the figure) of the horizontal line corresponding to the maximum value of the added R channel values (value on the x-axis in the figure) as the height of the horizontal plane. Figure 6 Figure 6

[0066] As a result of the determination, when the volume of the liquid reaches the target volume (S260), that is, when the extracted height of the horizontal plane exceeds the reference value, S230 to S260 are repeated until the last fraction (i.e., the microtube) is reached (S270).

[0067] Although the present application has been described with reference to the embodiments illustrated in the drawings, this is merely an example, and those skilled in the art will understand that various modifications and other embodiments equivalent thereto can be obtained therefrom. Therefore, the true technical scope of the present application should be determined by the technical idea of the appended claims.​

Claims

1. A liquid automatic fractionation apparatus for column chromatography, comprising: a microtube installation portion in which a plurality of microtubes for containing a liquid to be fractionated according to the column chromatography are installed; a liquid supply unit configured to sequentially supply the liquid to the plurality of microtubes; a light source unit provided at a rear side of the plurality of microtubes to emit light; an image acquisition unit provided at a front side of the plurality of microtubes to acquire images of the plurality of microtubes at a preset time interval while the column chromatography is performed; a volume determination unit configured to extract a height of a level of the contained liquid by analyzing the images of the plurality of microtubes acquired by the image acquisition unit, wherein the height of the level is extracted by a refraction phenomenon of light emitted onto the microtubes by the light source unit, and the volume determination unit is configured to determine whether a volume of the liquid contained in the plurality of microtubes reaches a target volume based on the extracted height of the level; and a control unit configured to set a reference value for determining whether the volume reaches the target volume, and configured to control operations of the liquid supply unit and the image acquisition unit according to a determination result of the volume determination unit, wherein the control unit checks whether alignment of the plurality of microtubes and the light source unit is correct based on brightness of the images of the plurality of microtubes; wherein the control unit adds R channel values of respective pixels on a central vertical line in the image of each of the plurality of microtubes, and determines that the alignment of the plurality of microtubes and the light source unit is correct when the added R channel values are greater than or equal to a preset value, and when the alignment is determined to be incorrect, corrects a position of the image acquisition unit by moving the image acquisition unit left or right, wherein the volume determination unit adds all R channel values of pixels on respective horizontal lines in the image of each of the plurality of microtubes to calculate a value of an x-axis, and extracts a position of the horizontal line in which the value of the x-axis is a maximum value as the height of the level. 2.The liquid automatic fractionation apparatus of claim 1, wherein the microtube installation portion is composed of a plurality of grooves formed in a housing to accommodate the plurality of microtubes, and the plurality of grooves are arranged in a row at a preset interval on a straight line. 3.The liquid automatic fractionation apparatus of claim 2, wherein a reference microtube containing a liquid of the target volume is installed in a first groove among the plurality of grooves, and the plurality of microtubes for fractionation are installed in other grooves.

4. The apparatus of claim 2, wherein, the liquid supply unit includes: a column including a stationary phase and a mobile phase; a column support connected to the housing so as to be movable along a linear axis parallel to a straight line in which the plurality of grooves are formed, and configured to support the column so that the column can be sequentially moved at positions corresponding to the plurality of microtubes; and a driving unit connected to a lower end of the column support and configured to move the column so that the column is located at an upper portion of one of the plurality of microtubes when the column chromatography is performed.

5. The liquid auto-fractionator of claim 4, wherein, The image acquisition unit includes: a camera mounted to be movable in sequence from the positions corresponding to the plurality of microtubes and configured to acquire images of the plurality of microtubes; a movement axis parallel to the straight line in which the plurality of grooves are formed, and on which the camera moves; and a driving unit configured to move the camera on the movement axis so that the camera is located in front of the plurality of microtubes when the column chromatography is performed. 6.The liquid automatic fractionation apparatus according to claim 1, wherein the volume determination unit analyzes the images of each of the plurality of microtubes obtained by the image acquisition unit at the preset time interval, and determines whether the volume reaches the target volume by using only the images in which the standard deviation of the height of the horizontal plane is less than or equal to a preset value. 7.The liquid automatic fractionation apparatus according to claim 1, wherein the volume determination unit finds a boundary surface of a refractive image in the images of the plurality of microtubes, extracts the height of the horizontal plane, and compares the height of the horizontal plane with the reference value to determine whether the volume reaches the target volume. 8.The liquid automatic fractionation apparatus according to claim 7, wherein the volume determination unit adds all R channel values of pixels on a corresponding horizontal line in the images of each of the plurality of microtubes for each horizontal line, extracts a position of the horizontal line in which the sum of the R channel values is the largest as the height of the horizontal plane, and determines that the volume reaches the target volume when the height of the horizontal plane exceeds the reference value. 9.The liquid automatic fractionation apparatus according to claim 1, further comprising: a display unit implemented as a touch screen to display buttons for receiving signals related to the operation of the liquid automatic fractionation apparatus, visualize a performance state of the column chromatography as a graph, and display the visualized performance state. 10.The liquid automatic fractionation apparatus according to claim 3, wherein the control unit acquires and analyzes the image of the reference microtube to extract the height of the horizontal plane, and sets the extracted height as the reference value.

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