KR102076809B1 - Chip for Sample Analysis and Device for Sample Analysis containing the same, and Catridge Mounted on chip for Sample Analysis - Google Patents

Abstract

Provide a cartridge. The cartridge according to the embodiment of the present application is mounted on a chip for analyzing a sample, and inserted into at least one of a sample storage unit, a first eluent storage unit, a second eluent storage unit, and a wash solution storage unit of the sample analysis chip. It may include at least one inlet for injecting the solution to be.

Description

Chip for Sample Analysis, Device for Sample Analysis Comprising the Same, and Cartridge Mounted on Chip for Sample Analysis {Chip for Sample Analysis and Device for Sample Analysis containing the same, and Catridge Mounted on chip for Sample Analysis}

The present application relates to a device for sample analysis.

Genetic diagnosis is essential for the diagnosis of bacterial or viral diseases.

Genetic diagnosis consists of four steps: sampling, sample pretreatment (DNA or RNA extraction), gene amplification, and detection.

However, conventionally, since each step is performed by a separate equipment or device, an expensive analysis device and a large amount of samples are required, a lot of time is required for analysis, and a sample is likely to be contaminated during diagnosis, There is a problem that rapid diagnosis is difficult.

In order to solve this problem, an integrated genetic analysis apparatus using a microchip based on microfluidic has been recently developed. It is called lab on a chip, meaning that the lab environment is implemented on one chip.

However, the conventional integrated genetic analysis device also has high production costs due to complex chip structure, metal electrode patterning and silicon / glass substrate based structure, complexity of operation due to the need of external inflow pump and multiple tube systems, and highly integrated chip driving. Problems such as low reproducibility of the device, automation of system driving, and difficulty in on-site diagnosis due to miniaturization limitations require improvement.

For example, a recently developed rotary gene diagnosis device and system is a device capable of performing pretreatment, gene amplification and detection of a sample introduced thereto through rotation of the device. However, the conventional rotary gene diagnosis device has the following problems.

First, the sample and the wash solution must be sequentially introduced into the capture passage for DNA or RNA capture in the sample, which is a problem that the sample and the wash solution are simultaneously introduced into the capture passage when the device is rotated.

Since the washing liquid washes the remaining material except DNA or RNA which is not captured in the capture passage, it is preferable that the washing solution is introduced after all the DNA or RNA is captured in the capture passage. If the wash is introduced before the DNA or RNA in the sample is captured in the capture passage, the accuracy of the assay is compromised.

Secondly, the diagnosis of multiple diseases in one device is impossible. In the case of avian influenza, various influenza viruses exist, and it is necessary to analyze which virus caused the disease.

However, the conventional device can only analyze one virus per unit processing unit. Therefore, it is difficult to meet the needs of the on-site diagnosis that requires rapid diagnosis because a large number of devices, manpower and time is required to determine the presence of the virus in the sample.

Third, the solution must be injected into each reservoir manually for sample analysis. On-site diagnostics (POCT) is the most important factor, but there is a risk of contamination if the solution is manually injected into each reservoir and takes a long time.

The above problems cause a decrease in the accuracy of gene diagnosis, and the need for a device that solves these problems is increasing.

Korean Patent Publication No. 10-2015-0031912 (2015.03.25) Japanese Patent Laid-Open No. 2004-012290 (2004.01.15)

The present application has been made to solve the above problems. More specifically, it is possible to simultaneously diagnose a plurality of viruses with a single device, and to provide a sample analysis chip that allows the washing solution to reach the capture passage later than the sample when rotated to analyze the sample.

In addition, the present application is to provide a cartridge for automatically injecting a solution to the chip for sample analysis.

One embodiment of the present application for solving the above problems, the sample storage unit, configured to capture the analysis target material contained in the sample, in communication with the sample storage unit, the outer side in the radial direction than the sample storage unit A capture passage located in the radially inward direction than the sample reservoir, the wash liquid reservoir communicating with the capture passage and between the capture passage and the wash liquid reservoir directed from the wash liquid reservoir to the capture passage; A retardation section through which the wash liquor passes, and when rotated to analyze the sample, the retardation unit is configured to delay the passage of the wash liquor entering the wash liquor reservoir such that the wash liquor reaches the capture passage later than the sample. Provides an analysis chip.

The delay unit may include a delay chamber in which the washing liquid from the washing liquid storage unit to the capture passage may be stored, and a predetermined amount of the washing liquid stored in the delay chamber between the delay chamber and the capture passage. In case of abnormality it may include a delay channel to be discharged to the capture passage.

In one embodiment, the radially outermost side of the delay chamber is provided with an outlet, the delay channel may be connected to the outlet of the delay chamber.

In one embodiment, the delay channel comprises a first passage extending radially inward at the exit of the delay chamber and a second passage extending radially outward at the end of the first passage; The predetermined amount may be an amount when the height of the washing liquid stored in the delay chamber coincides with the length of the first passageway.

In one embodiment, the delay unit is provided between the washing liquid reservoir and the delay chamber, when rotating to analyze the sample, the inlet channel configured to introduce the washing liquid stored in the washing liquid reservoir into the delay chamber It may further include.

In one embodiment, the inlet channel, the third passage extending away from the delay chamber in a direction parallel to the width direction of the delay chamber and the delay in a direction parallel to the third passage at the end of the third passage It may include a fourth passage extending close to the chamber.

In one embodiment, the first eluent storage portion located in the radial direction than the sample storage portion, in communication with the capture passage, the first eluent for storing the analysis material to be captured in the capture passage therein And a second eluent storage unit located radially outward from the sample storage unit and having a second eluent stored therein for detection of the analyte.

In one embodiment, a connecting chamber located radially outward from the capture passage and the second eluent reservoir, and in communication with the outlet of the capture passage and the second eluent reservoir, radially outward from the connection chamber. In a radial direction than the waste chamber and the connection chamber in communication with the connection chamber, wherein the sample is not captured in the capture passage and the wash liquid passing through the capture passage is rotated to analyze the sample. Communicate with the connection chamber so as to be located outside, and when rotated to analyze the sample, an analyte separated by the first eluent, a first eluate that has passed through the capture passage and the second eluent are stored; It may further comprise a collection chamber.

In one embodiment, an injection hole for solution injection may be formed in each of the sample reservoir, the wash liquid reservoir, the first eluent reservoir and the second eluent reservoir.

In addition, the present application is configured to perform a pre-treatment process including a sample storage unit, capture, washing and separation of the analyte contained in the sample, and the pre-treatment unit and the pre-treatment unit configured to communicate with the sample storage unit Located in the outer side in the radial direction, the analyte to be pre-treated by the pre-treatment unit is configured to flow, and includes a plurality of detection chambers are stored inside each different primer for the detection of the analyte, for sample analysis Provide chips.

In one embodiment, located in the radially inner side than the plurality of detection chamber, when the sample is rotated in a first direction with a first rotational speed to analyze the sample, the analyte material pre-treated in the pretreatment unit is added The input channel and the plurality of outlets of the input channel, respectively, extending to the plurality of detection chambers, and having a second rotational speed to rotate the first direction to analyze the sample, from the input channel to the detection chamber. The apparatus may further include a plurality of connection channels through which the analyte to be directed passes, and the input channel may extend toward the first direction at an inlet from which the analyte is introduced from the pretreatment unit.

In one embodiment, when rotating in the first direction with the second rotational speed, the width of at least one connection channel of the plurality of connection channels corresponds to a width of the analyte to be introduced into the detection chamber. It may be narrower than the width of the outlet of the input channel.

In some embodiments, the plurality of connection channels may be shorter as the connection channels are farther from the preprocessor.

In one embodiment, the second rotational speed may be faster than the first rotational speed.

In one embodiment, a hole corresponding to a rotation axis is formed in the center of the sample analysis chip, when rotating to analyze the sample, the distance between the at least two detection chamber and the through hole of the plurality of detection chambers is the same In addition, the plurality of connection channels may be shorter as the connection channels are farther from the preprocessor.

In addition, the present application is a sample analysis chip, the sample analysis chip is seated, the lower member is provided with a temperature control unit on the upper surface, the upper member mounted on the lower member so as to be movable to the lower member and the lower member And a drive motor mounted to the rotating device for rotating the sample analysis chip, wherein the sample analysis chip is configured to capture a sample storage material and an analysis target material contained in the sample, and communicates with the sample storage device. A capture passage located radially outward from the sample reservoir, a wash liquid reservoir positioned radially inward from the sample reservoir, and in communication with the capture passage, and between the capture passage and the wash liquid reservoir; A retardation section through which the wash liquor from the reservoir to the capture passage passes; To the delay unit provides a sample analysis device for configured to delay the passage of cleaning fluid coming from the cleaning fluid storage portion to a later washing liquid reaches the trapping passage than the sample.

In one embodiment, a hole corresponding to a rotation axis is formed in the center of the sample analysis chip, the hole is coupled to a column extending upward from the temperature control portion of the lower member to rotate the sample analysis chip. Can act as a rotation axis.

In addition, the present application is mounted on the chip for sample analysis, and inserted into at least one of the sample storage unit, the first eluent storage unit, the second eluent storage unit and the wash solution storage unit of the sample analysis chip to inject the corresponding solution It provides a cartridge, comprising at least one inlet for.

In one embodiment, the at least one inlet can be detachably inserted.

In one embodiment, each of the at least one inlet and in communication with each other, further comprising at least one solution reservoir for storing a corresponding solution, wherein the at least one inlet, the at least one injection hole when the sample analysis chip is rotated It may be in communication with the at least one solution reservoir at the position farthest from the axis of rotation so that the corresponding solution can be injected through one inlet.

In one embodiment, the at least one solution reservoir is provided with a first inlet that can be inserted into the sample reservoir, the Lysis Buffer Solution for the dissolution of the analyte to be contained in the sample is stored A first solution reservoir and a second injection hole which may be inserted into the washing solution storage unit may be provided, and may include a second solution storage unit in which the washing solution is stored.

In one embodiment, there is provided a third inlet that can be inserted into the first eluent reservoir, and a third inlet that can be inserted into the third solution reservoir and the second eluent reservoir in which the first eluent is stored. The apparatus may further include a fourth solution reservoir in which the second eluent is stored.

In one embodiment, the first to fourth injection port, the injection hole formed in the sample storage unit, the injection hole formed in the washing liquid storage unit, the injection hole formed in the first eluent storage unit and the second eluent storage unit Can be inserted into each of the injection holes formed in the detachable.

In one embodiment, the cartridge may be manufactured by a 3D printing method.

In one embodiment, the cartridge may be made of a disposable plastic material.

According to the present application, the cleaning solution is introduced into the capture passage later than the sample through the delay unit provided between the wash liquid storage unit and the capture passage, so that the accuracy of the diagnosis is improved because the wash liquid is introduced after the sample is sufficiently captured in the capture passage. Is improved.

In addition, a plurality of detection chambers are provided and different detection primers are stored in each detection chamber, so that diagnosis of several pathogens can be all performed in one device.

In addition, by using a discolored EBT indicator when the gene amplification is performed, it is possible to easily observe the presence or absence of the analyte in the sample.

In addition, the solution can be automatically injected into each reservoir through a cartridge detachably mounted on the chip for sample analysis, thereby reducing the time required for the conventional diagnosis and improving convenience.

1 is an exploded perspective view illustrating a state in which a sample analysis chip and a cartridge are coupled according to an embodiment of the present application.
FIG. 2 is a plan view of the sample analysis chip of FIG. 1. FIG.
3 is an enlarged view of a portion of the sample analysis chip of FIG. 2.
4 is an enlarged view of the exit of the capture passage.
5 is a view for explaining a delay unit for delaying the passage of the washing liquid flowing from the washing liquid storage unit.
6 is a perspective view for explaining a device for sample analysis according to an embodiment of the present application.
7 is a perspective view illustrating a cartridge according to an embodiment of the present application.
8 is a view showing the centrifugal force applied to the solution stored in the cartridge as the sample analysis chip equipped with the cartridge rotates.
9 is a view showing a cartridge coupled to the sample analysis chip, cartridge and the sample analysis chip according to an embodiment of the present application.
10 to 12 are diagrams showing the results of diagnosing pathogens using a sample analysis chip, a cartridge and a sample analysis device according to an embodiment of the present application.
13A to 13I are views for explaining a process of analyzing an analysis target material using a chip for analyzing a sample according to an embodiment of the present application.

Hereinafter, gene amplification means that the gene to be analyzed is amplified. Examples of gene amplification may include PCR (Polymerase Chain Reaction), Real-time PCR, and Isothermal Amplification Reaction, and any reaction that amplifies a gene to be analyzed is not limited thereto. Explain in advance.

1. Sample Analysis Chip

With reference to the accompanying drawings, it will be described in detail for the chip 1000 for analyzing the sample according to an embodiment of the present application.

Referring to FIG. 2, the sample analyzing chip 1000 may have a disc-shaped body 100. Through hole (H) is formed in the center of the body (100). The through hole H is coupled to the sample analyzing device 2000, which will be described later, and corresponds to a rotating shaft during rotation.

The sample analyzing chip 1000 includes a plurality of integrated process units 200a and 200b sequentially disposed along the circumferential direction. Hereinafter, the integrated process unit will be described as two, but is not limited to this, it is also possible to have two or more integrated process unit. Hereinafter, only one integrated process unit will be described in detail.

For example, the chip 1000 for analyzing the sample may be manufactured by machining a CNC milling machine on one surface of a disk-shaped PMMA having a thickness of 3 mm to form a predetermined pattern in a groove shape, and then attaching the PSA to the processed surface. .

The configuration of the integrated process unit will be described in more detail with reference to FIGS. 2 and 3.

Integrated process unit (200a, 200b) according to an embodiment of the present application is formed in a pattern on the body 100, respectively, sample storage unit 210, capture passage 220, wash liquid storage unit 230, delay unit ( 240, the first eluent reservoir 250, the second eluent reservoir 260, the connection chamber 270 and the plurality of detection chambers 280a, 280b, 280c, 280d, 280e, 280f, 280g, 280h, 280i, 280j, 280k, 280l, 280m, 280n, 280o, 280p, 280q, 280r, 280s, 280t).

The sample reservoir 210 is formed in a chamber shape and is located radially inward from the inlet 221 of the capture passage 220, which will be described later, and is connected to the inlet 221 of the capture passage 220. A sample injection hole 211 is formed in the sample storage unit 210 to inject a sample from the outside. After the sample is injected into the sample storage unit 210 through the sample injection hole 211, the sample injection hole 211 is sealed by the coupling with the cartridge 3000 to be described later. The sample stored in the sample storage unit 210 flows into the inlet 221 of the capture passage 220 by centrifugal force generated when the sample analyzing chip 1000 rotates.

The capture passage 220 is a portion for capturing the analyte to be contained in the sample. It is located outside the sample storage unit 210 in the radial direction and communicates with the sample storage unit 210. It may have a zigzag shape as shown in FIGS. 2 and 3 and has a catching means 223 such as silica beads therein. Inlets 221 and outlets 222 are positioned at both ends of the capture passage 220, respectively. The inlet 221 is located radially inward with respect to the through hole H and the outlet 222 is located radially outward. The capture means 223 adsorbs a substance containing the analyte to be contained in the sample. As shown in FIG. 4, the size of the capture means 223 is greater than the width of the outlet 222 to prevent departure from the capture passage 220.

The washing liquid storage unit 230 has a chamber shape and is located radially inward from the sample storage unit 210 and connected to the inlet 221 of the capture passage 220. The washing liquid stored in the washing liquid storage unit 230 removes the remaining components except for the analyte from the trapping material from the trapping means 223 by washing them. The washing solution storage unit 230 has a washing solution injection hole 231 for injecting the washing solution from the outside. After the washing liquid is injected into the washing liquid storage unit 230 through the washing liquid injecting hole 231, the washing liquid injecting hole 231 is sealed by a double-sided tape or the like. The washing solution stored in the washing solution storage unit 230 is introduced into the inlet 221 of the capture passage 220 by centrifugal force generated when the sample analyzing chip 1000 rotates.

The delay unit 240 is provided between the capture passage 220 and the wash liquid storage unit 230 to pass through the wash liquid from the wash liquid storage unit 230 to the capture passage 220. The delay unit 240 delays the passage of the washing liquid introduced from the washing liquid storage unit 230 so that the washing liquid reaches the capture passage 220 later than the sample when the sample analyzing chip 1000 rotates to analyze the sample. Let's do it.

Referring to FIG. 5, the configuration of the delay unit 240 will be described in more detail. The delay unit 240 includes an inlet channel 241, a delay chamber 242, and a delay channel 243.

The inflow channel 241 connects the washing liquid storage unit 230 and the delay chamber 242. The washing liquid stored in the washing liquid storage unit 230 flows into the delay chamber 242 when the sample analyzing chip 1000 rotates. This is to make it possible. The inflow channel 241 extends away from the delay chamber 242 in a direction parallel to the width direction of the delay chamber 242 and the third passage 241a at the end of the third passage 241a. And a fourth passage 241b extending closer to the delay chamber 242 in a direction parallel to the second chamber 242.

The delay chamber 242 is in the form of a chamber and is a part for storing the washing liquid introduced by the inflow channel 241. As the capacity of the delay chamber 242 increases, the inflow of the cleaning liquid toward the capture passage 220 may be delayed. At the radially outermost side of the delay chamber 242, an outlet 242b through which the washing liquid flows is formed.

The delay channel 243 is connected to the outlet 242b of the delay chamber 242, and is a portion to allow the washing liquid stored in the delay chamber 242 to delay inflow into the capture passage 220. Specifically, the delay channel 243 analyzes the sample at the end of the first passage 243a and the first passage 243a extending radially inward of the sample analyzing chip 1000 at the outlet 242b of the delay chamber. And a second passage 243b extending toward the radially outer side of the dragon chip 1000.

The delay channel 243 allows the washing liquid stored in the delay chamber 242 to be discharged to the capture passage 220 when the amount of the washing liquid is more than a predetermined amount. That is, in order for the washing liquid to pass through the first passage 243a and flow into the second passage 243b, the washing liquid stored in the delay chamber 242 should be more than a predetermined amount. Here, the predetermined amount is an amount when the height h of the washing liquid stored in the delay chamber 242 coincides with the length L of the first passage 243a. In this case, the capillary force is greater than the centrifugal force generated by the rotation of the sample analyzing chip 1000, so that the washing liquid may flow into the second passage 243b through the first passage 243a. That is, since the washing liquid does not flow into the capture passage 220 until the washing liquid reaches a predetermined amount in the delay chamber 242, the washing liquid is delayed reaching the capture passage 220 rather than the sample. Therefore, it is possible to prevent the cleaning liquid from flowing into the capture passage 220 before the substance including the analysis target material is captured in the capture passage 220.

The first eluent storage unit 250 is in the form of a chamber and is located in the radial direction of the sample analysis chip 1000 in the radial direction of the sample storage unit 210 and communicates with the inlet 221 of the capture passage 220. The first eluent is stored in the first eluent storage unit 250. The first eluent is a solution that separates the substance adsorbed by the trap means 223 from the trap means 223. As an example, the first eluent may be water. In the first eluent storage unit 250, a first eluent injection hole 251 for injecting the first eluent from the outside is formed. After the first eluent is injected into the first eluent reservoir 250 through the first eluent injecting hole 251, the first eluent injecting hole 251 is sealed by coupling with the cartridge 3000 described later. The first eluent stored in the first eluent storage unit 250 is introduced into the first eluent introduction passage 253 by centrifugal force generated when the sample analysis chip 1000 rotates.

The first eluent introduction passage 253 is connected to the outlet 252 of the first eluent reservoir located in the radially outermost side of the first eluent reservoir 250 and extends radially inward of the sample analysis chip 1000. And a sixth passage 253b connected to an end of the fifth passage 253a and the fifth passage 253a and extending radially outward of the sample analyzing chip 1000. Through the configuration of the fifth passage 253a and the sixth passage 253b, when the sample analyzing chip 1000 is rotating, the first eluent does not flow into the capture passage 220, and the sample analyzing chip 1000 does not flow. When the rotation is stopped after the rotation stop, the first eluent flows into the capture passage 220. Therefore, after both the sample and the washing liquid are introduced into the capture passage 220, the first eluent may flow into the capture passage 220.

The second eluent storage unit 260 is in the form of a chamber and is located outside the sample storage unit 210 in the radial direction of the sample analysis chip 1000 and communicates with the connection chamber 270 which will be described later. The second eluent storage unit 260 stores the second eluent. The second eluate is a solution containing an enzyme required for gene amplification of an analyte and an indicator. Here, the indicator may be used Eriochrome Black T (EBT) indicator that discolors when the gene is amplified. A second eluent injection hole 261 is formed in the second eluent storage unit 260 to inject the second eluent from the outside. After the second eluent is injected into the second eluent reservoir 260 through the second eluent injecting hole 261, the second eluent injecting hole 261 is sealed by engagement with the cartridge 3000 described later. The second eluent stored in the second eluent storage unit 260 flows into the second eluent introduction passage 263 by centrifugal force generated when the sample analysis chip 1000 rotates.

The second eluent introduction passage 263 is connected to the outlet 262 of the second eluent storage unit located at the outermost in the radial direction of the second eluent storage unit 260 and extends inward in the radial direction of the chip 1000 for sample analysis. And an eighth passage 263b connected to an end of the seventh passage 263a and the seventh passage 263a and extending radially outwardly of the sample analyzing chip 1000. Through the configuration of the seventh passage 263a and the eighth passage 263b, when the sample analyzing chip 1000 is rotating, the second eluent does not flow into the connection chamber 270, and the sample analyzing chip 1000 is applied. When stopped during this rotation, the second eluent flows into the connection chamber 270. Therefore, after both the sample and the washing liquid are introduced into the capture passage 220, the second eluent may be introduced into the connection chamber 270.

The connection chamber 270 is in the form of a chamber and is located outside the capture passage 220 and the second eluent reservoir 260 in the radial direction of the sample analysis chip 1000 and the outlet 222 of the capture passage 220. And the eighth passage 263b of the second eluent storage unit 260. Therefore, when the sample analyzing chip 1000 rotates, the material passing through the capture passage 220 or the second eluent introduction passage 263 passes through the connection chamber 270 to be described later. ) And the collection chamber 272.

The waste liquid chamber 271 is in the form of a chamber, and is located outside the connecting chamber 270 in the radial direction of the sample analysis chip 1000 and communicating with the connecting chamber 270. The waste liquid chamber 271 stores unnecessary components except for the analyte. The waste liquid chamber 271 may have a capacity of the sum of the sample and the washing liquid. A capillary valve is formed in the connection passage 273 connecting the connection chamber 270 and the waste liquid chamber 271 so that the waste liquid stored in the waste liquid chamber 271 is returned to the connection chamber 270. Inflow can be prevented.

The collection chamber 272 is in the form of a chamber, and is located outside the connecting chamber 270 in the radial direction of the sample analysis chip 1000 and communicating with the connecting chamber 270. The collection chamber 272 stores the analyte separated from the capture passage 220 and the first and second eluents. The analyte, the first eluent and the second eluent stored in the collection chamber 272 are introduced into the detection chamber introduction passage 276 by centrifugal force generated when the sample analyzing chip 1000 rotates.

The detection chamber introduction passage 276 is connected to the outlet 275 located at the outermost radial direction of the collection chamber 272 and extends in the radially inward direction of the chip 1000 for sample analysis. And a tenth passage 276b connected to the end of the ninth passage 276a and extending radially outward of the sample analyzing chip 1000. Through the configuration of the ninth passage 276a and the tenth passage 276b, when the sample analyzing chip 3000 is rotating, the substance stored in the collection chamber 272 does not flow into the connection chamber 270, and the sample analysis is performed. When the dragon chip 1000 is stopped during rotation, the material stored in the collection chamber 272 flows into the input channel 281.

The detection chamber 280 is a part where amplification and detection of analyte are performed. In order to detect a plurality of viruses or pathogens at one time using one sample analyzing chip 1000, a plurality of detection chambers 280 may be provided. 2 shows 20 detection chambers 280a, 280b, 280c, 280d, 280e, 280f, 280g, 280h, 280i, 280j, 280k, 280l, 280m, 280n, 280o, 280p, 280r, 280r, in one integrated process. 280s, 280t), but it is possible to have less than 20 or more than 20 detection chambers 280, without being limited thereto. Each detection chamber 280 may store different primers from each other. Preferably, the distance between each detection chamber and the through hole H is the same.

The input channel 281 is connected to the end of the tenth passage 276b of the detection chamber introduction passage 276 and has a first rotational speed to analyze the sample and rotates in the first direction when the substance is in the collection chamber 272. This is the part that is being injected. The input channel 281 may extend toward the first direction at the inlet through which material is introduced from the collection chamber 272. Here, the first direction is clockwise, the first rotational speed may be 1000 RPM.

The input channel 281 may have an aliqoting structure so that the material may be sequentially introduced by centrifugal force. The dispensing structure is a structure in which a plurality of outlets extend radially outward of one input channel 281 as shown in FIG. 2. Accordingly, the material may be sequentially introduced toward the first direction at the inlet through which the material is introduced from the collection chamber 272 to the plurality of outlet portions of the input channel 281 by the centrifugal force generated by the sample analyzing chip 1000. have.

The connecting channel 282 extends from the plurality of outlets of the input channel 281 to the plurality of detection chambers 280, respectively, so as to rotate in the first direction with a second rotational speed to analyze the sample. ) Is the portion through which the analyte to be directed to the detection chamber 282 passes. Here, the second rotational speed may be 5000 RPM. It is positioned outside the input channel 281 in the radial direction of the sample analysis chip 1000 and is formed to be narrower than the width of the outlet of the input channel 281. Here, the second rotational speed may be 5000 RPM. Therefore, when the sample analyzing chip 1000 rotates at a second rotational speed that is faster than the first rotational speed, the centrifugal force that the material receives radially outward is greater than the capillary force exerted radially inwardly from the detection chamber 280. The material contained in each of the plurality of outlets of the input channel 281 may be introduced into the detection chamber 280. As shown in Fig. 2, a plurality of connection channels 282a, 282b, 282c, 282d, 282e, 282f, 282g, 282h, 282i, 282j, 282k, 282l, 282m, 282n, 282o, 282p, 282q, 282r, 282s , 282t, is shorter for the connecting channel further away from the collection chamber 272. This is because the centrifugal force applied to the material contained at the outlet of the input channel 281 located in the circumferential direction decreases, so that the centrifugal force can be introduced into the detection chamber 280 even with the reduced centrifugal force.

The detection chambers 280a, 280b, 280c, 280d, 280e, 280f, 280g, 280h, 280i, 280j, 280k, 280l, 280m, 280n, 280o, 280p, 280q, 280r, 280s, 280t are connected channels It is a portion located outward in the radial direction of the sample analysis chip 1000 from 282. Each detection chamber 280a, 280b, 280c, 280d, 280e, 280f, 280g, 280h, 280i, 280j, 280k, 280l, 280m, 280n, 280o, 280p, 280q, 280r, 280s, 280t primer). Therefore, each of the detection chambers 280a, 280b, 280c, 280d, 280e, 280f, 280g, 280h, 280i, 280j, 280k, 280l, 280m, 280n, 280o, 280p, 280q, 280r, 280s, 280t The analyte may be reacted with or not reacted with a primer to observe the discoloration of the indicator contained in the second eluent, thereby analyzing the analyte included in the sample.

2. Sample analysis device

Referring to FIG. 6, a device for analyzing a sample 2000 according to an embodiment of the present application will be described in more detail.

The device for analyzing a sample 2000 according to the exemplary embodiment of the present application includes a lower member 310, an upper member 320, and a driving motor 330.

The temperature control part 311 is provided on the upper surface of the lower member 310. The temperature controller 311 is a portion that provides a temperature required for a gene amplification process of the analyte to be introduced into the detection chamber 280 of the sample analysis chip 1000. At the center of the temperature control unit 311 is formed a column 312 extending upward from the temperature control unit 311. The through hole H is combined with the pillar 312 to act as a rotation axis when the sample analysis chip 1000 is rotated by the driving motor 330 which will be described later.

The upper member 320 is a portion mounted above the lower member 310 so as to be movable relative to the lower member 310. The sample analysis chip 1000 is accommodated in the upper member 320. The upper member 320 is moved with respect to the lower member 310 until the sample analyzing chip 1000 accommodated in the upper member 320 contacts the temperature control part 311.

The driving motor 330 is a portion that is mounted below the lower member 310 to rotate the sample analysis chip 1000 seated on the temperature controller 311. As the sample analyzing chip 1000 is rotated by the driving motor 330, preprocessing, amplification, and detection of the sample may be performed.

3. Cartridge

A cartridge 3000 according to an exemplary embodiment of the present application will be described in detail with reference to FIGS. 7 and 8.

The cartridge 3000 according to the embodiment of the present application is mounted on the above-described sample analysis chip 1000, and the sample storage unit 210, the washing solution storage unit 230, and the first eluent of the sample analysis chip 1000. The solution is injected into at least one reservoir of the reservoir 250 and the second eluent reservoir 260.

Referring to FIG. 7, the cartridge 3000 according to the exemplary embodiment of the present application may include a first solution reservoir 410, a second solution reservoir 420, a third solution reservoir 430, and a fourth solution reservoir. 440.

The first solution storage unit 410 stores a Lysis Buffer Solution for eluting the analyte to be contained in the sample. The first solution storage unit 410 has a first injection hole 411 protruding from one surface to be inserted into the sample storage unit 210 of the sample analysis chip 1000. The first injection hole 411 may be detachably inserted into the sample storage unit 210. The first injection hole 411 may be in communication with the first solution storage part 410 at a position farthest from the rotation axis so that the solution may be injected through the first injection hole 411 when the sample analyzing chip 1000 rotates. . As a result, since the centrifugal force may be applied to the solution stored therein, as shown in FIG. can do. A first injection groove 411a is formed in the first injection hole 411 so that the solution stored in the first solution storage part 410 may be discharged. When the sample analysis chip 1000 rotates, the solution stored in the first solution storage unit 410 is discharged to the outside through the first injection groove 411a.

The second solution storage unit 420 stores a washing solution for washing and removing the remaining components except the analyte from the trapping material from the trapping means 223. The second solution storage unit 420 has a second injection hole 421 protruding from one surface to be inserted into the cleaning solution storage unit 230 of the sample analysis chip 1000. The second injection hole 421 may be detachably inserted into the washing liquid storage unit 230. The second injection hole 421 may be in communication with the second solution storage part 420 at a position farthest from the rotation axis so that the solution may be injected through the second injection hole 421 when the sample analyzing chip 1000 rotates. As a result, centrifugal force may be applied to the solution stored therein as shown in FIG. 8 when the sample analyzing chip 1000 rotates, thereby preventing a so-called dead space phenomenon, in which the solution does not discharge from the inside to the outside. can do. A second injection groove 421a is formed in the second injection hole 421 so that the solution stored in the second solution storage part 420 may be discharged. When the sample analysis chip 1000 is rotated, the solution stored in the second solution reservoir 420 is discharged to the outside through the second injection groove 421a.

In the third solution storage unit 430, the first eluent for separating the analyte to be adsorbed by the capture unit 233 is stored. The third solution storage unit 430 has a third injection hole 431 protruding from one surface to be inserted into the first eluent storage unit 250 of the sample analysis chip 1000. The third injection hole 431 may be detachably inserted into the first eluent storage part 250. The third injection hole 431 may be in communication with the third solution storage part 430 at a position farthest from the rotation axis so that the solution may be injected through the third injection hole 431 when the sample analyzing chip 1000 rotates. As a result, centrifugal force may be applied to the solution stored therein as shown in FIG. 8 when the sample analysis chip 1000 is rotated, thereby preventing a so-called dead space phenomenon in which the solution does not discharge from the inside to the outside. can do. A third injection groove 431a is formed in the third injection hole 431 so that the solution stored in the third solution storage part 430 may be discharged. When the sample analyzing chip 1000 rotates, the solution stored in the third solution storage unit 430 is discharged to the outside through the third injection groove 431a.

The fourth solution storage unit 440 stores an enzyme required for gene amplification of the analyte and a second eluent including an indicator. The fourth solution storage unit 440 has a fourth injection hole 441 protruding from one surface of the fourth solution storage unit 440 so as to be inserted into the second eluent storage unit 260 of the sample analysis chip 1000. The fourth injection hole 441 may be detachably inserted into the second eluent storage unit 260. The fourth injection hole 441 may be in communication with the fourth solution reservoir 440 at a position farthest from the rotation axis so that the solution may be injected through the fourth injection hole 441 when the sample analysis chip 1000 is rotated. As a result, centrifugal force may be applied to the solution stored therein as shown in FIG. 8 when the sample analysis chip 1000 is rotated, thereby preventing a so-called dead space phenomenon in which the solution does not discharge from the inside to the outside. can do. A fourth injection groove 441a is formed through the fourth injection hole 441 so that the solution stored in the fourth solution reservoir 440 may be discharged. When the sample analysis chip 1000 is rotated, the solution stored in the fourth solution reservoir 440 is discharged to the outside through the fourth injection groove 441a.

4. Verification Experiment 1

Experiments were performed to verify the superiority of the diagnosis using the sample analysis chip 1000, the sample analysis device 2000, and the cartridge 3000 according to the exemplary embodiment of the present application. In the following verification experiments, gene amplification was performed using an isothermal amplification reaction.

First, an experiment was performed to detect E. coli O157: H7 bacteria contained in milk. The sample used 25 μL of milk and the concentration of E. coli O157: H contained in the sample was 4 × 10 ³ cells / μL.

First, a sample prepared in 75 μL Lysis Buffer Solution stored in the first solution reservoir 410 of the cartridge 3000 was loaded, and E. coli O157: H7 bacteria were dissolved in the lysis buffer for 1 minute.

Each of the injection holes 411, 421, 431, and 441 of the cartridge 3000 is inserted into the storage units 210, 230, 250, and 260 of the sample analysis chip 1000. The cartridge 3000 is mounted on the sample analyzing chip 1000 so that a solution is injected into each of the storage parts 210, 230, 250, and 260 of the sample analyzing chip in the solution storage part of the cartridge 3000. FIG. 13A illustrates a state in which a solution is injected into each of the reservoirs 210, 230, 250, and 260 after mounting the cartridge 3000.

Next, as illustrated in FIG. 13B, the sample analysis chip 1000 was rotated at a rotation speed of 5000 RPM (ω ₀ ) in the counterclockwise direction. The sample stored in the sample storage unit 210 is introduced into the capture passage 220, and unnecessary substances other than the analyte to be adsorbed by the capture means 223 of the capture passage 220 are connected to the connection chamber 270 and the connection channel. Into the waste liquid chamber 271 through 273.

Next, as shown in FIG. 13C, the washing solution stored in the washing solution storage unit 230 is introduced into the capture passage 220 through the delay unit 240, except for the substance to be analyzed from the substance captured by the capture unit 223. The remaining components were washed out of the capture means 223 and introduced into the waste liquid chamber 271 through the connection chamber 270 and the connection channel 273, too.

Next, as shown in FIG. 13D, the rotation of the sample analyzing chip 1000 was stopped (ω ₁ ). Since the centrifugal force applied to the solution is removed when the sample analysis chip 1000 is stopped, the first eluent and the second eluent stored in the first eluent storage 250 and the second eluent storage 260 by capillary force are removed. The eluent passed through the first eluent introduction passage 253 and the second eluent introduction passage 263.

Next, the sample analyzing chip 1000 was rotated at a rotational speed of 5000 RPM (ω ₂ ) in the clockwise direction as shown in FIG. 13E. The first eluent flowed into the capture passage 220 to separate the material adsorbed to the capture means 223 from the capture means 223 and introduced into the collection chamber 272 through the connection chamber 270. It entered the collection chamber 272 through the connection chamber 270.

Next, as shown in FIG. 13F, the rotation of the sample analyzing chip 1000 was stopped (ω ₁ ). When the rotation of the sample analyzing chip 1000 is stopped, the centrifugal force applied to the solution is removed, and the substance stored in the collection chamber 272 has passed through the detection chamber introduction passage 276 by capillary force.

Next, as shown in FIG. 13G, the sample analyzing chip 1000 was rotated at a rotational speed of 1000 RPM (ω ₃ ) in the clockwise direction. The material passing through the detection chamber introduction passage 276 flows into the input channel 281, and the material introduced into the input channel 281 extends along the circumferential direction in a clockwise direction as the sample analyzing chip 1000 rotates. A plurality of outlets of the input channel 281 was introduced.

Next, as shown in FIG. 13H, the sample analyzing chip 1000 was rotated at a rotational speed of 5000 RPM (ω ₂ ) in the clockwise direction. The material stored in the plurality of outlets of the input channel 281 passes through the plurality of connecting channels 282 having a narrower width than the outlets, so that the respective detection chambers 280a, 280b, 280c, 280d, 280e, 280f, 280g, 280h, 280i, 280j, 280k, 280l, 280m, 280n, 280o, 280p, 280q, 280r, 280s, 280t) was confirmed that the flow into.

Next, the detection chamber 280a, 280b, 280c, 280d, 280e, 280f, 280g, 280h, 280i, 280j, 280k, 280l, 280m, 280n, for 1 hour at a temperature of 63 ° C through the temperature control unit 311 280o, 280p, 280q, 280r, 280s, 280t) were heated. In this case, when the gene amplification process is performed, discoloration (purple-> blue) of the EBT indicator included in the second eluate occurs as shown in FIG. 13I.

E. coli O157: H primers were stored in the first to fourth detection chambers 280a, 280b, 280c, and 280d of the plurality of detection chambers, and the sixth to ninth detection chambers 280f, 280g, 280h, and 280i. Salmonella Typhimurium primers were stored, Vibrio parahaemolyticus primers were stored in the 11th to 14th detection chambers (280k, 280l, 280m, and 280n), and 16th and 19th detection chambers (280p, 280q, 280r, and 280s). Listeria monocytogenes primers were stored. No primers were stored in the fifth, 10, 15, and 20 detection chambers 280d, 280i, 280o, and 280t.

As a result, as shown in FIG. 10, only the colors of the first to fourth detection chambers 280a, 280b, 280c, and 280d were changed to blue. Therefore, the results of using the sample analyzing chip 1000, the sample analyzing device 2000, and the cartridge 3000 according to the exemplary embodiment of the present application can accurately and easily diagnose specific pathogens.

5. Verification Experiment 2

Experiments were performed to determine whether the pathogen can be diagnosed at the same time using the sample analysis chip 1000, the sample analysis device 2000, and the cartridge 3000 according to the exemplary embodiment of the present application.

The experimental procedure was applied in the same manner as the above 4. Validation Experiment 1. Only the sample (1) 25μL milk: E. coli O157: H7 and S.Typhimurium were present at 4x10 ³ cells / μL, respectively (2) 25μL Milk: E. coli O157: H7, S.Typhimurium, and V.parahaemolyticus are present at 4x10 ³ cells / μL, respectively. (3) 25 μL milk: E.coli O157: H7, S.Typhimurium, V. Parahaemolyticus and L.monocytogenes were changed to 4x10 ³ cells / μL, respectively.

As a result of the experiment, as shown in Figure 11, even if the type of pathogen contained in the sample was confirmed that the diagnosis is possible for each pathogen.

6. Verification Experiment 3

An experiment was performed to identify a limit of detection (LOD) of pathogens using the sample analyzing chip 1000, the sample analyzing device 2000, and the cartridge 3000 according to the exemplary embodiment of the present application.

The experimental procedure was applied in the same manner as in 4. Validation Experiment 1 described above, using only 25 μL of milk containing E. coli O157: H7, S.Typhimurium, V.parahaemolyticus, and L.monocytogenes. (1) 10 ⁴ cells of pathogens contained in the sample (2) 10 ³ cells of pathogens contained in the sample (3) 10 ² cells of pathogens contained in the sample were tested using a total of three samples. .

As a result, as shown in FIG. 12, even if the sample contained only 100 cells of the pathogen, it was confirmed that the diagnosis of the pathogen was possible.

That is, the limit of detection of pathogens using the sample analyzing chip 1000, the sample analyzing device 2000, and the cartridge 3000 according to the exemplary embodiment of the present application was 10 ² cells. .

In the present specification, the present invention has been described with reference to the embodiments shown in the drawings so that those skilled in the art can easily understand and reproduce the present application, which is merely exemplary, and those skilled in the art can make various modifications and equivalents from the embodiments of the present application. It will be appreciated that embodiments are possible. Therefore, the protection scope of the present application will be defined by the claims.

100: body
200a, 200b: integrated process unit
210: sample storage unit
211: sample injection hole
220: capture passage
221: entrance
222 exit
223: capture means
230: washing liquid storage unit
231: cleaning liquid injection hole
240: delay unit
241: inflow channel
241a: third passage
241b: fourth passage
242: delay chamber
242a: entrance
242b: exit
243: delay channel
243a: first passage
243b: second passage
250: first eluent storage unit
251: first eluent injection hole
253: first eluent introduction passage
253a: fifth passage
253b: sixth aisle
260: second eluent storage unit
261: second eluent injection hole
262: exit
263: second eluent introduction passage
263a: seventh aisle
263b: eighth passage
270: connection chamber
271 waste liquid chamber
272: collection chamber
273: connecting passage
275: exit
276: detection chamber introduction passage
276a: 9th passage
276b: passage 10
280: detection chamber
281 input channel
282: connection channel
310: lower member
311: temperature controller
312: pillar
320: upper member
330: drive motor
410: first solution reservoir
411: first injection hole
420: second solution reservoir
421: second injection hole
430: third solution reservoir
431: third inlet
440: fourth solution reservoir
441: fourth inlet
1000: chip for sample analysis
2000: device for sample analysis
3000: cartridge
H: through
S: sample
W: washing liquid
E ₁ : first eluent
E ₂ : second eluent

Claims (8)

A solution corresponding to the storage unit which is mounted on the sample analysis chip and inserted into the sample storage unit, the first eluent storage unit, the second eluent storage unit, and the wash solution storage unit formed at different positions of the sample analysis chip; It includes a plurality of injection holes each through which the injection groove is formed for injecting and in communication with each injection groove, and a plurality of solution storage for storing the corresponding solution, respectively,
The plurality of injection holes are injected into the sample storage unit, the first eluent storage unit, the second eluent storage unit and the washing solution storage unit together with a corresponding solution through the plurality of injection holes when the sample analysis chip is rotated. Communicating with the at least one solution reservoir at a position farthest from the axis of rotation of the sample analysis chip,
cartridge.
The method of claim 1,
The plurality of inlet is detachably inserted,
cartridge.
delete The method of claim 1,
The plurality of solution storage unit,
A first solution storage unit having a first injection hole which can be inserted into the sample storage unit and storing a lysis buffer solution for dissolution of the analyte to be contained in the sample; And
It includes; a second injection port is provided that can be inserted into the washing liquid storage unit, the second solution storage unit storing the washing liquid;
cartridge.
The method of claim 4, wherein
The plurality of solution storage unit,
A third solution storage part having a third injection hole which can be inserted into the first eluent storage part and storing the first eluent solution; And
A fourth injection port is provided that can be inserted into the second eluent storage unit, and further comprising a fourth solution storage unit in which the second eluent is stored.
cartridge.
The method of claim 5,
The first to fourth injection holes may include a sample injection hole formed in the sample storage part, a washing liquid injection hole formed in the washing liquid storage part, a first eluent injection hole and a second eluent storage part formed in the first eluent storage part. Removably inserted into the formed second eluent injection hole,
cartridge.
The method of claim 1,
The cartridge is characterized in that produced by the 3D printing method,
cartridge.
The method of claim 1,
The cartridge is made of a disposable plastic material,
cartridge.

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