KR20230101702A - Cartridge type device for biomarker molecular diagnosis - Google Patents

Abstract

A cartridge-type biomarker molecular diagnostic device is disclosed. A cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention includes a cartridge into which a polymerase chain reaction (PCR) sample is put and flows, and a main body on which the cartridge is seated, wherein the cartridge has the sample input inlet; one or more pressure inlets arranged to introduce pressure for the flow of the sample; and a reaction passage in which the sample introduced through the inlet flows or stays in a mixed state with a heating material, a primer, and a polymerase that generate heat when irradiated with reaction light, and the main body includes the one or more pressure one or more pressure supply units supplying pressure for the flow of the sample to the inlet; and a reaction light irradiation unit configured to irradiate the reaction light to at least a portion of the reaction passage, and while the sample is irradiated with the reaction light inside the reaction passage, the exothermic material generates heat so that the sample is heated. And, while the reaction light is not irradiated, the exothermic material does not generate heat and the sample can be cooled.

Description

Cartridge type device for biomarker molecular diagnosis}

The present invention relates to a cartridge-type biomarker molecular diagnosis device, and more particularly, to a cartridge-type biomarker molecular diagnosis device that performs biomarker molecular diagnosis such as PCR (Polymerase Chain Reaction).

A large-scale diagnosis is required in the context of the pandemic of the coronavirus infection (COVID-19). During a disease pandemic, identifying and isolating as many symptomatic or asymptomatic infected people as quickly as possible is the most effective way to prevent disease transmission.

Immunogenic lateral flow assays have small test equipment, rapid results, and cost-effectiveness, but have problems that are not suitable for virus detection in early disease stages. In comparison, the polymerase chain reaction (PCR)-based nucleic acid amplification test (NAAT) has high assay accuracy (~99%) in virus detection. Accordingly, reverse transcription PCR (RT-PCR) is used as a standard in the diagnosis of coronavirus infection.

However, since most PCR diagnoses are performed in laboratories, there are disadvantages in that a lot of cost is required for transporting and preserving samples, and it takes up to several days to obtain results. In order to overcome these disadvantages, there is an attempt to make the PCR equipment site-oriented (POINT OF CARE, POC). However, conventional on-site PCR equipment is not widely used because it is bulky and unsuitable for transport, and the analysis time is rather long, 1 to 2 hours. In addition, it is pointed out as a problem that the accuracy of the test is somewhat limited compared to traditional PCR equipment.

Korean Patent Registration No. 10-2426968

The present invention is to solve the above problems, and an object of the present invention is to provide a cartridge-type biomarker molecular diagnosis device capable of quickly and effectively detecting nucleic acids through PCR.

Another object of the present invention is to provide a cartridge-type biomarker molecular diagnosis device that efficiently performs biomarker molecular diagnosis for a plurality of samples.

The tasks of the present invention are not limited to the tasks mentioned above, and other tasks not mentioned will be clearly understood by those skilled in the art from the description below.

According to one aspect of the present invention, a cartridge for PCR (Polymerase Chain Reaction) is injected and flows, and a body in which the cartridge is seated, wherein the cartridge includes: an inlet through which the sample is introduced; one or more pressure inlets arranged to introduce pressure for the flow of the sample; and a reaction passage in which the sample introduced through the inlet flows or stays in a mixed state with a heating material, a primer, and a polymerase that generate heat when irradiated with reaction light, and the main body includes the one or more pressure one or more pressure supply units supplying pressure for the flow of the sample to the inlet; and a reaction light irradiation unit configured to irradiate the reaction light to at least a portion of the reaction passage, and while the sample is irradiated with the reaction light inside the reaction passage, the exothermic material generates heat so that the sample is heated. and, while not being irradiated with the reaction light, the exothermic material does not generate heat and the sample is cooled.

At this time, the cartridge further includes a first heating passage disposed between the inlet and the reaction passage, and the main body includes the first heating passage so that the sample flows or stays in the first heating passage and is heated to a first temperature. A first heating unit for heating the flow path may be further disposed.

In addition, the cartridge is disposed between the inlet and the reaction passage, the collection body for collecting the sample; and at least one washing solution reservoir storing a washing solution supplied to the collecting body to wash impurities present in the sample, wherein the washing solution is discharged by the pressure supplied by the pressure supply unit through the pressure inlet. It can be transferred to the collection body.

In addition, the cartridge may include a connection passage connecting the inlet and the one or more washing solution reservoirs to the collecting body; and a first valve disposed to selectively connect the connection passage to any one of the inlet and the one or more washing solution reservoirs, wherein a first valve driving unit for driving the first valve is further disposed in the main body. It can be.

In addition, the cartridge further includes a second valve disposed to selectively connect the connection passage to any one of the collection body and the reaction passage, and the main body further includes a second valve driving unit for driving the second valve. can be placed.

In addition, the cartridge further includes a discharged solution reservoir for storing the discharged solution discharged from the collecting body after washing the sample, and the discharged solution is discharged by the pressure supplied by the pressure supply unit through the pressure inlet. can be transferred to the reservoir.

The cartridge may further include a mixed solution reservoir configured to communicate with the connection flow path by the first valve and to store a mixed solution including the exothermic material, the primer, and the polymerase. .

In addition, the cartridge further includes a transfer liquid reservoir for storing a transfer liquid for transferring the sample concentrated in the collecting body to the mixed liquid reservoir after washing, and the transfer by the pressure supplied by the pressure supply unit through the pressure inlet. The liquid may transfer the sample to the mixed liquid reservoir.

In addition, the cartridge further includes a third valve disposed to selectively connect the collection body with any one of the discharged solution reservoir and the transfer liquid reservoir, and the main body includes a third valve driving unit for driving the third valve. may be further placed.

In addition, the cartridge further includes a detection passage connected to a rear end of the reaction passage, and in the main body, a detection light is emitted into the detection passage to detect whether or not a nucleic acid to be detected is present in the sample transferred from the reaction passage. a detection light irradiation unit for irradiating; and a detector configured to detect fluorescence generated by the detection light.

In addition, the exothermic material has magnetism, the cartridge further includes a separation passage disposed between the reaction passage and the detection passage, and the main body generates heat when the sample flows from the separation passage to the detection passage. A magnetic separation unit providing a magnetic force so that the material is separated from the sample and remains in the separation passage may be further disposed.

The cartridge further includes a second heating passage disposed between the inlet and the reaction passage, and the main body heats the second heating passage so that the sample is heated to a second temperature in the second heating passage. A second heating unit that does may be further disposed.

According to the configuration described above, the cartridge-type biomarker molecular diagnosis device of the present invention enables PCR to be performed quickly and efficiently by repeatedly heating and cooling the sample mixed with the pyrogen in the reaction passage of the cartridge.

The cartridge-type biomarker molecular diagnosis device of the present invention has a structure in which samples to be subjected to biomarker molecular diagnosis are placed in a cartridge detachable from the main body, so that biomarker molecular diagnosis for a plurality of samples can be efficiently performed by replacing the cartridge. allow it to be performed

The effects of the present invention are not limited to the above effects, and should be understood to include all effects that can be inferred from the detailed description of the present invention or the configuration of the invention described in the claims.

1 is a diagram showing the configuration of a cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention.
2 is a view showing the upper side of the cartridge of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention.
3 is a view showing the lower side of the cartridge of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention in a state in which the cover member is separated.
4 is a view showing the upper side of the cartridge of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention in a state in which a significant portion of the cartridge body is removed.
5 is a view showing the lower side of the cartridge of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention in a state in which a significant portion of the cartridge body is removed.
6 is a view showing the lower side of the cartridge of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention with a significant portion of the cartridge body removed, but with the first to third valves cut off.
7 is a cross-sectional view of a magneto-plasmonic nanoparticle (MPN).
8 is a graph showing the temperature change of a sample repeating PCR cycles in a reaction flow path of a cartridge-type biomarker molecular diagnostic device according to an embodiment of the present invention.
9 is a view showing a part of the main body of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention.
10 is a view showing first to third valve driving units of a cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention.
11 to 19 are diagrams showing the process of PCR in the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention.

Hereinafter, with reference to the accompanying drawings, embodiments of the present invention will be described in detail so that those skilled in the art can easily carry out the present invention. This invention may be embodied in many different forms and is not limited to the embodiments set forth herein. In order to clearly describe the present invention, parts irrelevant to the description are omitted in the drawings, and the same reference numerals are assigned to the same or similar components throughout the specification.

Words and terms used in this specification and claims are not construed as limited in their ordinary or dictionary meanings, but in accordance with the principle that the inventors can define terms and concepts in order to best describe their inventions. It should be interpreted as a meaning and concept that corresponds to the technical idea. Since the embodiments described in this specification and the configurations shown in the drawings correspond to a preferred embodiment of the present invention and do not represent all of the technical ideas of the present invention, the corresponding configurations are various equivalents to replace them at the time of filing of the present invention. There may be water and variations.

In this specification, terms such as "include" or "have" are intended to describe the presence of features, numbers, steps, operations, components, parts, or combinations thereof described in the specification, but one or more other features It should be understood that the presence or addition of numbers, steps, operations, components, parts, or combinations thereof is not precluded.

A component being in the "front", "rear", "above" or "below" of another component means that it is in direct contact with another component, unless there are special circumstances, and is "in front", "rear", "above" or "below". It includes not only those disposed at the lower part, but also cases in which another component is disposed in the middle. In addition, the fact that certain components are “connected” to other components includes cases where they are not only directly connected to each other but also indirectly connected to each other unless there are special circumstances.

1 shows the configuration of a cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention.

Referring to FIG. 1 , a cartridge-type biomarker molecular diagnosis device 1 according to an embodiment of the present invention includes a cartridge 100 and a body 300. The cartridge-type biomarker molecular diagnosis apparatus 1 according to an embodiment of the present invention has a structure in which a sample to be subjected to biomarker molecular diagnosis is disposed in a cartridge 100 detachable from a main body 300. According to the cartridge-type biomarker molecular diagnosis device 1 according to an embodiment of the present invention, a sample mixed with a pyrogen can flow inside the cartridge 100, and PCR is performed through repeated cycles of heating and cooling. (Polymerase Chain Reaction) can be done quickly and efficiently.

The cartridge 100 is provided so that the sample is injected and flows. For example, the sample may be a sample for PCR (Polymerase Chain Reaction). More specifically, the sample is subject to nucleic acid detection through PCR, and may be prepared by purifying RNA or DNA from saliva of a subject. In this case, the nucleic acid to be detected may be the N1 and N2 genes for detecting the SARS-CoV-2 virus and the human RPP30 gene for confirming that the sample is a human sample. Of course, this is just one example, and the nucleic acid to be detected may vary depending on the type of infection to be diagnosed.

2 shows the upper side of the cartridge of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention, and FIG. 3 covers the lower side of the cartridge of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention. The member is shown in a separated state. 4 shows the upper side of the cartridge of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention with a significant portion of the cartridge body removed, and FIG. 5 is a cartridge-type device according to an embodiment of the present invention. The lower side of the cartridge of the biomarker molecular diagnosis device is shown with a significant portion of the cartridge body removed. Meanwhile, FIG. 6 shows a cutaway view of the first to third valves of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention in a state in which a significant portion of the cartridge body is removed from the lower side of the cartridge.

2 to 6, the cartridge 100 includes a cartridge body 101, an inlet 102, a first heating passage 103, a collecting body 104, and one or more cleaning solution reservoirs 105a, 105b, and 105c. ), connection flow path 106, first valve 107, second valve 108, discharged solution reservoir 109, mixed solution reservoir 110, transported liquid reservoir 111, third valve 112, It may include two heating passages 113, a reaction passage 114, a separation passage 115 and a detection passage 116, and one or more pressure inlets 117a, 117b, 117c, and 117d.

The cartridge body 101 is provided so that it can be seated on the main body 300. In one embodiment of the present invention, the cartridge body 101 may have a rectangular box shape as a whole. The shape of the cartridge body 101 may be determined in consideration of arrangement and operation of components of the cartridge 100 and components of the main body 300 . Accordingly, the shape of the cartridge body 101 may be variously modified.

The inlet 102 is disposed on the cartridge body 101 so that the sample is injected. In one embodiment of the present invention, the inlet 102 is connected to the first heating passage 103, and the sample introduced through the inlet 102 may flow toward the first heating passage 103.

The first heating passage 103 is connected to the inlet 102 and disposed on the cartridge body 101. In one embodiment of the present invention, the first heating passage 103 is disposed between the inlet 102 and the reaction passage 114 on the path of the sample. The sample may be heated to a first temperature while flowing or staying in the first heating passage 103 .

For example, the first heating passage 103 may be formed in a negative shape on a predetermined area of one surface of the cartridge body 101 and covered by the first cover member C1. In this case, the first cover member C1 may be made of a film or the like.

When PCR for detecting a nucleic acid to be detected is performed using the cartridge-type biomarker molecular diagnosis device 1 according to an embodiment of the present invention, the first temperature may be selected from 90 to 100°C. For example, the sample may be heated to 95° C. inside the first heating passage 103 . When the sample is heated to the first temperature, the virus is lysed in the sample, and the nucleic acid to be detected can be extracted.

Collector 104 collects the sample. Concentration of the sample may be performed by washing the sample while the sample is adhered to the collector 104 . In one embodiment of the present invention, the collector 104 is disposed between the inlet 102 and the reaction flow path 114 on the path of the sample. For example, the collection body 104 may be made of resin.

One or more washing solution reservoirs 105a, 105b, 105c store a washing solution supplied to the collector 104 to wash impurities present in the sample. One or more cleaning solution reservoirs 105a, 105b, and 105c may be disposed in a predetermined area of the cartridge body 101. Meanwhile, the cleaning solution stored in the one or more cleaning solution reservoirs 105a, 105b, and 105c may be supplied to the collector 104 by pressure supplied from the main body 300.

In one embodiment of the present invention, the one or more washing solution reservoirs 105a, 105b, and 105c include a first washing solution reservoir 105a in which the first washing solution is stored, and a second washing solution reservoir in which the second washing solution is stored ( 105b), and a third washing solution reservoir 105c in which a third washing solution is stored. In this case, the first washing solution may be ethanol, the second washing solution may be a wash buffer, and the third washing solution may be mineral oil.

The connection passage 106 connects the inlet 102 and one or more washing solution reservoirs 105a, 105b, and 105c to the collecting body 104. In one embodiment of the present invention, the connection flow path 106 includes a first cleaning solution discharged from the first cleaning solution reservoir 105a, a second cleaning solution discharged from the second cleaning solution reservoir 105b, and a third cleaning solution. The third washing solution discharged from the reservoir 105c is formed to be provided to the collector 104, respectively. More specifically, the discharge channels of each of the first to third cleaning solution reservoirs 105a, 105b, and 105c may be formed to converge toward the collecting body 104.

For example, the connection passage 106 may be formed in a negative shape on a predetermined area of one surface of the cartridge body 101 and covered by the second cover member C2. In this case, the second cover member C2 may be made of a film or the like.

Meanwhile, the connection passage 106 may connect the mixed solution reservoir 110 and the collecting body 104 to be described later. In other words, in one embodiment of the present invention, the connection passage 106 may connect the inlet 102, one or more washing solution reservoirs 105a, 105b, and 105c, and the mixed solution reservoir 110 to the collection body 104. .

The first valve 107 is disposed to selectively connect the connection flow path 106 with any one of the inlet 102 and one or more washing solution reservoirs 105a, 105b, and 105c. The first valve 107 may be rotatably disposed inside the cartridge body 101 . Referring to FIG. 6, in one embodiment of the present invention, the first valve 107 is formed by penetrating the first valve body 107a in a cylindrical shape and the first valve body 107a in the transverse direction. A flow path 107b may be included.

Meanwhile, in one embodiment of the present invention, the connection flow path 106 is also connected to the mixed solution reservoir 110 . In this regard, the first valve 107 may communicate the mixed solution reservoir 110 with the connection passage 106 . That is, the first valve 107 may selectively connect the connection passage 106 to any one of the inlet 102, one or more washing solution reservoirs 105a, 105b, and 105c, and the mixed solution reservoir 110.

The second valve 108 is disposed to selectively connect the connection passage 106 with either the collector 104 or the reaction passage 114 . The second valve 108 may be rotatably disposed inside the cartridge body 101 . Referring to FIG. 6, in one embodiment of the present invention, the second valve 108 includes a second valve body 108a made of a cylinder shape and a second valve formed penetrating the second valve body 108a in a T shape. A flow path 108b may be included.

The discharged solution reservoir 109 stores the discharged solution discharged from the collector 104 after washing the sample. The discharged solution reservoir 109 may be disposed in a predetermined area of the cartridge body 101 . In one embodiment of the present invention, the first to third washing solutions may be transferred to the discharged solution reservoir 109 after washing the sample attached to the collector 104 . At this time, the transfer force to the discharged solution reservoir 109 may be formed by the pressure supplied from the main body 300 .

In one embodiment of the present invention, the discharged solution reservoir 109 may be formed in a negative shape on one surface of the cartridge body 101 and covered by a third cover member (C3). At this time, the third cover member (C3) may be made of a film or the like.

The mixed solution reservoir 110 is disposed to communicate with the connection flow path 106 by the first valve 107 and stores a mixed solution including a heating material, a primer, and a polymerase. The exothermic material is used for heating the sample, and the primer and the polymerase are used for reverse transcription or PCR of a nucleic acid to be detected inside the sample. The mixed solution reservoir 110 may be disposed in a predetermined area of the cartridge body 101 . In one embodiment of the present invention, the mixed solution reservoir 110 may be disposed adjacent to the first to third cleaning solution reservoirs 105a, 105b, and 105c.

For example, the exothermic particle may be a magneto-plasmonic nanoparticle (MPN). In this regard, FIG. 7 shows a cross-sectional view of the magnetic plasmon nanoparticles 10 . Referring to FIG. 7 , the magnetic plasmon nanoparticle 10 may include a core 11 and a shell 12 surrounding the core 11 . More specifically, the core 11 may have magnetism. The core 11 may include any one or more of Fe3O4, Zn0.4Fe2.6O4, FexOy, ZnxFeyOz, and MnxFeyOz. In addition, the shell 12 may include at least one of gold (Au), silver (Ag), and copper (Cu).

Magnetoplasmon nanoparticles 10 may have a nanoscale size. For example, the diameter of the core 11 may be 5 to 100 nm, and the thickness of the shell 12 may be 1 to 20 nm.

The transfer liquid reservoir 111 stores a transfer liquid for transferring the sample concentrated in the collecting body 104 to the mixed liquid reservoir 110 after washing. The transfer liquid reservoir 111 may be disposed in a predetermined area of the cartridge body 101 . The transfer liquid stored in the transfer liquid reservoir 111 transfers the sample concentrated in the collector 104 to the mixed liquid reservoir 110 by the pressure provided from the main body 300 . For example, the transfer fluid may be water.

In one embodiment of the present invention, the transfer liquid stored in the transfer liquid reservoir 111 is disposed so that the third valve 112, which will be described later, connects the collector 104 and the transfer liquid reservoir 111, and the second valve ( 108 is disposed to connect the collector 104 with the connection passage 106, and the first valve 107 is arranged to connect the connection passage 106 with the mixed liquid reservoir 110, the collector 104 The concentrated sample may be transferred to the mixed solution reservoir 110.

The third valve 112 is arranged to selectively connect the collecting body 104 with either the discharged solution reservoir 109 or the transfer solution reservoir 111 . The third valve 112 may be rotatably disposed inside the cartridge body 101 . Referring to FIG. 6, in one embodiment of the present invention, the third valve 112 includes a third valve body 112a having a cylindrical shape and a third valve formed by penetrating the third valve body 112a in an L shape. A flow path 112b may be included.

The second heating passage 113 is disposed so that the mixed liquid and the mixed sample are transported and heated to a second temperature. In one embodiment of the present invention, the second heating passage 113 is disposed between the inlet 102 and the reaction passage 114 on the path of the sample. In more detail, the second heating passage 113 may be disposed between the mixed solution reservoir 110 and the reaction passage 114 on the path of the sample. That is, the sample may flow into the second heating passage 113 after being mixed with the exothermic material, the primer, and the polymerase in the mixed solution reservoir 110 .

The sample may be heated to a second temperature while flowing or staying in the second heating passage 113 . For example, the second heating passage 113 may be formed in a negative shape on a predetermined area of one surface of the cartridge body 101 and covered by the first cover member C1. In this case, the first cover member C1 may be made of a film or the like.

When PCR for detecting a nucleic acid to be detected is performed using the cartridge-type biomarker molecular diagnosis device 1 according to an embodiment of the present invention, the second temperature may be selected from 45 to 55°C. For example, the sample may be heated to 52° C. in the second heating passage 113 . When the sample is heated to the second temperature, reverse transcription of RNA in the sample may proceed. When the nucleic acid to be detected is RNA, reverse transcription of RNA is required before PCR is performed in the reaction passage 114, and reverse transcription may be performed inside the second heating passage 113.

On the other hand, when the nucleic acid to be detected in the sample is DNA, reverse transcription is not required. Accordingly, when the target nucleic acid to be detected in the sample is DNA, the second heating passage 113 may be omitted.

The reaction passage 114 is arranged so that the sample introduced through the inlet 102 flows or stays in a mixed state with a heating material generating heat when irradiated with reaction light, the primer, and the polymerase. In one embodiment of the present invention, the reaction passage 114 may be formed in a negative shape on a predetermined area of one surface of the cartridge body 101 and covered by the first cover member C1. In this case, the first cover member C1 may be made of a film or the like.

While the sample is irradiated with the reaction light inside the reaction passage 114, the exothermic material mixed with the sample generates heat to heat the sample, and while the reaction light is not irradiated, the exothermic material generates heat. and the sample is allowed to cool. As described above, when the exothermic particles are magnetoplasmon nanoparticles 10 and the shell 12 is made of gold (Au) having a thickness of 12 nm, it has been confirmed that plasmon resonance is exhibited for light having a wavelength of 535 nm. In this case, for efficient heating of the sample, the peak wavelength of the first light may be 530 to 540 nm (eg, 535 nm). In other words, the reaction light may be laser light having a peak wavelength of 530 to 540 nm. Of course, when different types of particles are used as the exothermic material, the wavelength at which plasmon resonance occurs may be different.

의 온도에서 진행되고, 결합은 50~65

의 온도에서 진행되며, 신장은 75~80

의 온도에서 진행될 수 있다. 본 발명의 일 실시예에 의할 경우, 상기 샘플에 혼합된 발열 물질은 반응 광의 조사 시 열을 발생시켜 샘플을 가열하며, 반응 광을 조사받지 않는 동안에는 발열 물질이 열을 발생시키지 않으며 샘플이 냉각된다. 이에 따라 도 8에 나타난 바와 같은 샘플의 온도 변화 주기가 반복되도록 할 수 있다. 그 결과 변성, 결합 및 신장이 반복되면서 PCR이 수행될 수 있다.PCR of the sample may be performed in the reaction passage 114 . PCR consists of three steps: denaturation, annealing, and elongation. Generally, denaturation is 94 to 98

at a temperature of 50 to 65

It is carried out at a temperature of 75 to 80

can be carried out at a temperature of According to an embodiment of the present invention, the exothermic material mixed in the sample heats the sample by generating heat when the reaction light is irradiated, and the exothermic material does not generate heat while the reaction light is not irradiated, and the sample cools do. Accordingly, the temperature change cycle of the sample as shown in FIG. 8 can be repeated. As a result, PCR can be performed while denaturation, conjugation and elongation are repeated.

Various methods may be considered regarding the flow of the sample in the reaction channel 114 and the irradiation of the reaction light.

First, it may be considered that the reaction light is continuously irradiated to a certain area of the reaction passage 114, and the sample alternately proceeds between an area where the reaction light is irradiated and an area where the reaction light is not irradiated within the reaction passage 114. For example, when there is an area where the reaction light is not irradiated before and after the area where the reaction light is irradiated in the reaction passage 114, the sample passes through the area where the reaction light is irradiated a plurality of times, and the area to which the reaction light is irradiated and its The front and back reaction light can flow back and forth in an unirradiated area. At this time, the flow of the sample may be achieved by the pressure supplied from the body 300.

Next, a method in which the sample stays inside the reaction passage 114 and the reaction light is intermittently irradiated may also be considered. That is, in a state where the sample stays inside the reaction passage 114, the reaction light is turned off after being irradiated for a predetermined time and then irradiated again after a predetermined time has elapsed, so that heating and cooling of the exothermic material can be repeatedly performed.

The separation passage 115 is disposed in connection with the reaction passage 114 . In one embodiment of the present invention, the separation passage 115 is disposed between the reaction passage 114 and the detection passage 116 . Inside the separation passage 115, the caustic sample and the exothermic material may be separated. For example, the separation passage 115 may be formed in a negative shape on a predetermined area of one surface of the cartridge body 101 and covered by the first cover member C1. In this case, the first cover member C1 may be made of a film or the like.

When the cartridge-type biomarker molecular diagnosis device 1 according to an embodiment of the present invention is used for PCR, the exothermic material efficiently induces a PCR reaction by directly heating the sample within the sample, but after completion of the PCR In the process of detecting a nucleic acid to be detected, it may be an obstacle to detection. For example, the pyrogenic substance may interfere with detection of the nucleic acid to be detected by absorbing or reflecting (emitting light) the detection light used during detection. Therefore, the pyrogen is preferably removed from the sample prior to detection of the nucleic acid to be detected.

In one embodiment of the present invention, the separation passage 115 may be formed on the premise that the heating material has magnetism, such as the magneto-plasmon nanoparticles 10 described above. In this case, the heating material is fixed inside the separation passage by the magnetic force provided from the main body 300, and the sample may be separated from the heating material and flow toward the detection passage 116.

Meanwhile, when the heating material does not have magnetism or does not act as an obstacle in the detection process of the nucleic acid to be detected, the separation flow path 115 may be omitted.

The detection passage 116 is disposed connected to the rear end of the reaction passage 114 . In one embodiment of the present invention, irradiation of detection light and detection of fluorescence may be performed while the sample remains in the detection passage 116 . For example, the detection passage 116 may be formed in a negative shape on a predetermined area of one surface of the cartridge body 101 and covered by the first cover member C1. In this case, the first cover member C1 may be made of a film or the like.

One or more pressure inlets 117a, 117b, 117c, 117d are arranged to introduce pressure for the flow of the sample. In one embodiment of the present invention, the cartridge 100 has first to fourth pressure inlets 117a, 117b, 117c and 117d in the cartridge body 101.

The first pressure inlet (117a) is formed on the discharged solution reservoir (109) side. The sample introduced from the inlet 102 may flow toward the first heating passage 103 and the collecting body 104 through the negative pressure introduced through the first pressure inlet 117a. In addition, the washing liquid stored in the one or more washing solution reservoirs 105a, 105b, and 105c may be transferred to the collector 104 through the negative pressure introduced through the first pressure inlet 117a.

The second pressure inlet 117b is formed on the transfer liquid reservoir 111 side. The transfer liquid may be transferred from the transfer liquid reservoir 111 to the mixed liquid reservoir 110 through the collector 104 through the positive pressure introduced through the second pressure inlet 117b. The sample concentrated in the collecting body 104 may be transferred to the mixture liquid reservoir 110 by the transfer liquid.

The third pressure inlet (117c) is formed on the collecting body (104) side. The negative pressure introduced through the third pressure inlet 117c induces the transfer liquid stored in the transfer liquid reservoir 111 to be transferred to the collector 104 side. In one embodiment of the present invention, when the positive pressure introduced through the second pressure inlet 117b is sufficient, the third pressure inlet 117c may be omitted.

The fourth pressure inlet 117d is formed at the rear end of the detection passage 116 and communicates with the detection passage 116 . The sample mixed with the mixed solution in the mixed solution reservoir 110 may flow toward the second heating passage 113 through negative pressure introduced through the fourth pressure inlet 117d.

In one embodiment of the present invention, the first heating passage 103, the second heating passage 113, the reaction passage 114, the separation passage 115 and the detection passage 116 are formed on one surface of the cartridge body 101. It is formed in reverse and can be covered by the first cover member (C1). In addition, a connection passage 106 is formed on one side of a predetermined area of one surface of the cartridge body 101, and the connection passage 106 may be covered by the second cover member C2. In addition, the discharged solution reservoir 109 is disposed in an area adjacent to a predetermined area on one surface of the cartridge body 101, and may be covered by the third cover member C3.

The main body 300 is provided on which the cartridge 100 is seated. The main body 300 may be made in the form of a frame. The main body 300 has a seating portion 301 on which the cartridge 100 is seated. In the cartridge-type biomarker molecular diagnosis device 1 according to an embodiment of the present invention, PCR of samples put into the cartridge 100 is performed while the cartridge 100 is seated on the seat 301 of the main body 300. It can be. More specifically, the seating portion 301 includes a seating groove 301a in which the cartridge 100 is seated, and a seating cover 301b disposed to cover at least a portion of the cartridge 100 seated in the seating groove 301a. can include

Various components for biomarker molecular diagnosis (eg, PCR) of the sample put into the cartridge 100 may be disposed on the main body 300 . Referring to FIG. 9 showing a part of the main body of the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention together with FIG. 1, the main body 300 includes a first heating unit 310, a first valve A driving unit 320a, a second valve driving unit 320b, a third valve driving unit 320c, a second heating unit 330, a reaction light irradiation unit 340, a magnetic separation unit 350, a detection light irradiation unit 360, A detection unit 370 and a pressure supply unit 380 may be disposed.

The first heating unit 310 heats the first heating channel 103 so that the sample flows or stays in the first heating channel 103 and is heated to a first temperature. For example, the first heating unit 310 may include a heater block disposed in contact with the first heating passage 103 . As described above, when PCR is performed to detect a nucleic acid to be detected, the first temperature may be selected from 90 to 100°C. For example, the sample may be heated to 95° C. inside the first heating passage 103 . Accordingly, the virus may be lysed in the sample and the nucleic acid to be detected may be extracted.

The first valve driver 320a, the second valve driver 320b, and the third valve driver 320c drive the first valve 107, the second valve 108, and the third valve 112, respectively. 10 shows first to third valve driving units of a cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention. Referring to FIG. 10 , the first valve driving unit 320a, the second valve driving unit 320b, and the third valve driving unit 320c operate on a first motor 321a, a second motor 321b, and a third valve driving unit 321a generating driving force. It may include a motor 321c, and may include a first driving force transmission unit 322a, a second driving force transmission unit 322b, and a third driving force transmission unit 322c that respectively transmit driving force generated by the motor. The first to third driving force transmitting means 322a, 322b, and 322c may include a pulley structure receiving rotational force of the first motor 321a, the second motor 321b, and the third motor 321c, respectively. It is fastened to the lower side of the first to third valves 107, 108, and 112 to transmit rotational driving force.

The second heating unit 330 heats the second heating passage 113 so that the sample is heated to a second temperature within the second heating passage 113 . For example, the second heating unit 330 may include a heater block disposed in contact with the second heating passage 113 . In one embodiment of the present invention, the first heating unit 310 and the second heating unit 330 may be integrally formed.

As described above, when PCR is performed in which the nucleic acid to be detected is RNA, the second temperature may be selected from 45 to 55°C. For example, the sample may be heated to 52° C. in the second heating passage 113 . When the sample is heated to the second temperature, reverse transcription of RNA in the sample may proceed. On the other hand, when the nucleic acid to be detected in the sample is DNA, reverse transcription is not required. That is, when the nucleic acid to be detected in the sample is DNA, the second heating unit 330 may be omitted.

The reaction light irradiator 340 radiates reaction light to at least a portion of the reaction passage 114 . For example, the reaction light irradiator 340 may irradiate laser light of a set wavelength range. The reaction light has a wavelength for generating heat in the exothermic material. Heat may be generated by light of different wavelengths depending on the type of exothermic material, and the reaction light may have different wavelengths depending on the type of exothermic material. For example, the peak wavelength of the reaction light may have an arbitrary range selected from 400 to 800 nm.

The reaction light irradiator 340 may continuously radiate the reaction light to a certain area of the reaction passage 114 or intermittently radiate the reaction light to at least a portion of the reaction passage 114 . While the reaction light irradiator 340 is irradiating the reaction light, the exothermic material generates heat to heat the sample while the reaction light is irradiated inside the reaction passage 114, and the sample is heated while the reaction light is not irradiated. The exothermic material does not generate heat and the sample is cooled.

As described above, when the exothermic particles are magnetoplasmon nanoparticles 10 and the shell 12 is made of gold (Au) having a thickness of 12 nm, it has been confirmed that plasmon resonance is exhibited for light having a wavelength of 535 nm. In this case, for efficient heating of the sample, the peak wavelength of the first light may be 530 to 540 nm (eg, 535 nm). In other words, the reaction light irradiator 340 may irradiate laser light having a peak wavelength of 530 to 540 nm as the reaction light.

The magnetic separator 350 provides magnetic force so that when the sample flows from the separation passage 115 to the detection passage 116, the exothermic material having magnetism is separated from the sample and remains in the separation passage 115. The magnetic separator 350 includes a magnet, and may further include a transfer means for transferring the magnet.

The detection light irradiation unit 360 radiates detection light to the detection passage 116 to detect whether or not a nucleic acid to be detected is present in the sample transferred from the reaction passage 114 . The detection light may have a wavelength that induces fluorescence excitation of the nucleic acid to be detected. For example, the detection light may have a wavelength selected from 100 to 350 nm. In one embodiment of the present invention, the detection light emitter 360 may include a first detection light source 361, a second detection light source 362, and a third detection light source 363 for irradiating detection light of different wavelengths. can That is, one of the first detection light source 361, the second detection light source 362, and the third detection light source 363 may emit detection light according to the type of target nucleic acid to be detected.

The detector 370 detects fluorescence generated by the detection light. The fluorescence detected by the detector 370 may be displayed by a separate display device. In one embodiment of the present invention, the detection unit 370 may be disposed on the seating cover 301b.

One or more pressure supply units 380a and 380b supply pressure for the flow of the sample to one or more pressure inlets 117a, 117b, 117c, and 117d. In addition, the cleaning solution and the transporting solution may be transported through the pressure supplied through the pressure inlets 117a, 117b, 117c, and 117d. In one embodiment of the present invention, the seat cover 301b has a seat cover pressure inlet at a position corresponding to the one or more pressure inlets 117a, 117b, 117c, and 117d, and the one or more pressure supply units 380a and 380b are Pressure (negative or positive) may be supplied through the seat cover pressure inlet to one or more pressure inlets 117a, 117b, 117c, 117d.

In one embodiment of the present invention, the main body 300 has a first pressure supply unit 380a supplying negative pressure to the first pressure inlet 117a and applying pressure to the second to fourth pressure inlets 117b, 117c, and 117d. A second pressure supply unit 380b is disposed.

The first pressure supply unit 380a may include a pump providing a relatively large negative pressure, and may be connected to the first pressure inlet 117a through a tube (not shown). In addition, the second pressure supply unit 380b includes a cylinder pump 381b for generating pressure and one or more valve blocks 382b for controlling the supply of the pressure generated by the cylinder pump 381b, and the valve block 382b ) and the second to fourth pressure inlets 117b, 117c, and 117d may be connected through tubes (not shown).

Meanwhile, the main body 300 includes a first heating unit 310, a first valve driving unit 320a, a second valve driving unit 320b, a third valve driving unit 320c, a second heating unit 330, and a reaction light irradiation unit. A control unit 390 that controls one or more of the 340 , the magnetic separation unit 350 , the detection light irradiation unit 360 , the detection unit 370 , and the pressure supply unit 380 may be disposed. The controller 390 may be a computer in which a program for automatically performing biomarker molecular diagnosis is stored. In other words, after the sample is injected into the cartridge 100 and the cartridge 100 is seated on the main body 300, biomarker molecular diagnosis (eg, PCR) is automatically performed under the control of the control unit 390. can be performed with

11 to 19 show the process of PCR in the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention. Hereinafter, the process of PCR in the cartridge-type biomarker molecular diagnosis device according to an embodiment of the present invention will be described with reference to FIGS. 11 to 19. At this time, the nucleic acid to be detected may be RNA.

The sample S introduced through the inlet is heated to a first temperature on the first heating passage 103 (see FIG. 11). The sample S may be heated by the first heating unit 310 while being disposed in the first heating passage 103 . As described above, the first temperature of the first heating passage 103 may be selected from 90 to 100 °C. When the sample S is heated to the first temperature, the virus is lysed in the sample S, and the nucleic acid to be detected can be extracted.

At this time, the first valve 107, the second valve 108, and the third valve 112 are arranged so that the first pressure inlet 117a communicates with the first heating passage 103, and the sample S is 1 It may flow by the negative pressure introduced through the pressure inlet (117a). The sample S heated in the first heating passage 103 passes through the first valve 107 and the second valve 108 to the collecting body 104 and may be collected in the collecting body 104 .

Next, the first cleaning solution (S1) is supplied to the collector 104 (see FIG. 12). The first valve 107 connects the first cleaning solution reservoir 105a and the connection flow path 106 and is disposed, and the second valve 108 connects the connection flow path 106 and the collector 104 and is disposed. , In a state where the third valve 112 is disposed to connect the collector 104 and the drained solution reservoir 109, the first cleaning solution S1 stored in the first cleaning solution reservoir 105a is stored in the collector 104 ) can be provided.

At this time, the first washing solution (S1) may be transported by negative pressure introduced through the first pressure inlet (117a), and after washing the sample (S) in the collection body (104) to the discharge solution reservoir (109) It is discharged. As described above, the first cleaning solution S1 may be ethanol.

Next, the second cleaning solution (S2) is supplied to the collector 104 (see FIG. 13). The first valve 107 connects the second cleaning solution reservoir 105b and the connection flow path 106 and is disposed, and the second valve 108 connects the connection flow path 106 and the collecting body 104 and is disposed. , In a state where the third valve 112 connects the collector 104 and the drained solution reservoir 109 and is disposed, the second cleaning solution S2 stored in the second cleaning solution reservoir 105b is stored in the collector 104 ) can be provided.

At this time, the second washing solution (S2) may be transported by negative pressure introduced through the first pressure inlet (117a), and after washing the sample (S) in the collection body (104) to the discharge solution reservoir (109). It is discharged. As described above, the second washing solution S2 may be a wash buffer.

Next, the third washing solution (S3) is supplied to the collector 104 (see FIG. 14). The first valve 107 connects the third cleaning solution reservoir 105c and the connection passage 106 and is disposed, and the second valve 108 connects the connection passage 106 and the collecting body 104 and is disposed, , In a state where the third valve 112 connects the collector 104 and the drained solution reservoir 109 and is disposed, the third cleaning solution S3 stored in the third cleaning solution reservoir 105c is stored in the collector 104 ) can be provided.

At this time, the third washing solution (S3) may be transported by negative pressure introduced through the first pressure inlet (117a), and after washing the sample (S) in the collecting body (104) to the discharge solution reservoir (109). It is discharged. As described above, the third cleaning solution S3 may be mineral oil.

Next, the sample (S) concentrated in the collecting body 104 after washing is transferred to the mixed solution reservoir 110 by the transfer liquid (W) and mixed with the mixed liquid (M) (see FIG. 15). The third valve 112 connects the transfer liquid reservoir 111 and the collecting body 104 and is disposed, and the second valve 108 connects the collecting body 104 and the connection passage 106 and is disposed. 1 valve 107 connects the connection flow path 106 and the mixed solution reservoir 110 and can transfer the sample in which the transfer fluid W is concentrated in the collector 104 to the mixed solution reservoir 110 in a state in which it is disposed. there is.

The transfer liquid W may be transferred by positive pressure introduced through the second pressure inlet 117b. At this time, the negative pressure introduced through the third pressure inlet 117c may be used as an aid to the transfer of the transfer liquid (W). On the other hand, the mixed solution (M) includes a heating material, a primer (primer) and a polymerase. The pyrogen, the primer, and the polymerase are as described above.

Next, the mixed liquid M and the mixed sample S are heated to a second temperature on the second heating passage 113 (see FIG. 16). The sample S may be heated by the second heating unit 330 while being disposed in the second heating passage 113 . As described above, the second temperature may be selected from 45 to 55 °C. For example, the sample may be heated to 52° C. inside the second heating passage 113 . When the sample S is heated to the second temperature, reverse transcription of the nucleic acid to be detected may be performed.

On the other hand, in the sample S mixed with the mixture M, the first valve 107 connects the mixed solution reservoir 110 and the connection flow path 106, and the second valve 108 connects the connection flow path 106. In a state of being connected to the second heating passage 113 and disposed, it may flow into the second heating passage 113 via the connection passage 106 . In addition, the sample (S) may flow by negative pressure introduced through the fourth pressure inlet (117d).

Next, the sample S is transferred to the reaction passage 114, and PCR may be performed in the reaction passage 114 (see FIG. 17). In a state where the sample S is placed in the reaction passage 114 , the reaction light irradiation unit 340 radiates reaction light to at least a portion of the reaction passage 114 . As described above, the reaction light may be continuously irradiated to a certain area of the reaction passage 114 or intermittently irradiated to at least a portion of the reaction passage 114 .

The exothermic material mixed in the sample (S) generates heat when the reaction light is irradiated to heat the sample (S), and while the reaction light is not irradiated, the exothermic material does not generate heat and the sample (S) is cooled. Accordingly, the temperature change cycle of the sample as shown in FIG. 8 is repeated, and PCR may be performed while denaturation, conjugation, and elongation are repeated. Meanwhile, the sample S may flow due to negative pressure introduced through the fourth pressure inlet 117d. In addition, negative pressure and positive pressure may be alternately provided to the fourth pressure inlet 117d so that the sample S repeatedly passes through a region of the reaction passage 114 where the reaction light is irradiated.

Next, the sample S is transferred to the separation passage 115, and the separation of the exothermic material having magnetism may proceed in the separation passage 115 (see FIG. 18). As described above, the exothermic material mixed in the sample S is fixed in the separation passage 115 by the magnetic force provided by the magnetic separator 350, and the sample S can flow into the detection passage 116. there is. At this time, the sample (S) may flow by the negative pressure introduced through the fourth pressure inlet (117d).

Finally, the sample S is transferred to the detection passage 116, and the nucleic acid to be detected is detected through the detection light in the detection passage 116 (see FIG. 19). The detection light irradiation unit 360 irradiates the detection light to the sample S, and the detection unit 370 detects fluorescence generated by the detection light. Meanwhile, the sample S may flow from the separation passage 115 to the detection passage 116 by negative pressure introduced through the fourth pressure inlet 117d.

Although the embodiments of the present invention have been described, the spirit of the present invention is not limited by the embodiments presented herein, and those skilled in the art who understand the spirit of the present invention may add or change components within the scope of the same spirit. Other embodiments can be easily proposed by adding, deleting, adding, etc., but this will also be said to be within the scope of the present invention.

1: Cartridge type biomarker molecular diagnosis device
100: cartridge
300: body

Claims (12)

A cartridge for PCR (Polymerase Chain Reaction) sample is injected and flows, and a main body on which the cartridge is seated,
the cartridge,
an inlet through which the sample is injected;
one or more pressure inlets arranged to introduce pressure for the flow of the sample; and
A reaction passage in which the sample introduced through the inlet flows or stays in a mixed state with a heating material, a primer, and a polymerase that generate heat when irradiated with reaction light;
In the body,
one or more pressure supply units supplying pressure for the flow of the sample to the one or more pressure inlets; and
A reaction light irradiator for irradiating the reaction light to at least a portion of the reaction passage is disposed,
While the sample is irradiated with the reaction light inside the reaction passage, the exothermic material generates heat and the sample is heated, and while the sample is not irradiated with the reaction light, the exothermic material does not generate heat and the sample is A cooling cartridge-type biomarker molecular diagnostic device. According to claim 1,
The cartridge further includes a first heating passage disposed between the inlet and the reaction passage,
A cartridge-type biomarker molecular diagnostic device further disposed on the main body, wherein a first heating unit for heating the first heating channel so that the sample flows or stays in the first heating channel and is heated to a first temperature. According to claim 1,
the cartridge,
a collecting body disposed between the inlet and the reaction passage and collecting the sample; and
Further comprising; one or more washing solution reservoirs for storing a washing solution supplied to the collecting body to wash impurities present in the sample;
A cartridge-type biomarker molecular diagnostic device in which the washing solution is transferred to the collection body by the pressure supplied by the pressure supply unit through the pressure inlet. According to claim 3,
the cartridge,
a connection passage connecting the inlet and the one or more washing solution reservoirs to the collecting body; and
A first valve disposed to selectively connect the connection flow path to any one of the inlet and the one or more washing solution reservoirs; further comprising,
A cartridge-type biomarker molecular diagnostic device further disposed in the main body with a first valve driving unit for driving the first valve. According to claim 4,
The cartridge further includes a second valve disposed to selectively connect the connection passage to any one of the collecting body and the reaction passage,
A cartridge-type biomarker molecular diagnostic device further disposed on the main body with a second valve driving unit for driving the second valve. According to claim 4,
The cartridge further comprises a discharge solution reservoir for storing the discharge solution discharged from the collection body after washing the sample,
The cartridge-type biomarker molecular diagnosis device in which the discharged solution is transferred to the discharged solution reservoir by the pressure supplied by the pressure supply unit through the pressure inlet. According to claim 6,
The cartridge is disposed to be in communication with the connection flow path by the first valve, and further includes a mixed solution reservoir for storing a mixed solution including the heating material, the primer, and the polymerase. Molecular diagnostic device. According to claim 7,
The cartridge further includes a transfer liquid reservoir for storing a transfer liquid for transferring the sample concentrated in the collection body to the mixed liquid reservoir after washing,
The cartridge-type biomarker molecular diagnosis device in which the transfer liquid transfers the sample to the mixed liquid reservoir by the pressure supplied by the pressure supply unit through the pressure inlet. According to claim 8,
The cartridge further includes a third valve disposed to selectively connect the collection body with any one of the discharged solution reservoir and the transported liquid reservoir,
A cartridge-type biomarker molecular diagnostic device further disposed in the main body with a third valve driving unit for driving the third valve. According to claim 1,
The cartridge further includes a detection passage connected to a rear end of the reaction passage,
In the body,
a detection light irradiation unit for radiating detection light to the detection passage in order to detect whether a nucleic acid to be detected is present in the sample transferred from the reaction passage; and
A cartridge-type biomarker molecular diagnostic device further disposed; a detection unit for detecting fluorescence generated by the detection light. According to claim 10,
The exothermic material has magnetism,
The cartridge further includes a separation passage disposed between the reaction passage and the detection passage;
A cartridge-type biomarker molecular diagnostic device further disposed on the main body to provide a magnetic force so that the exothermic substance is separated from the sample and remains in the separation passage when the sample flows from the separation passage to the detection passage. According to claim 1,
The cartridge further includes a second heating passage disposed between the inlet and the reaction passage,
A cartridge-type biomarker molecular diagnostic device further disposed on the main body, a second heating unit for heating the second heating channel so that the sample is heated to a second temperature in the second heating channel.

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