KR20240028940A - Tip for Extracting Target Ingredient and Portable Extraction Device Comprising the Same - Google Patents

Abstract

Disclosed is a tip for extracting ingredients and a portable ingredient separation device including the same.
According to one aspect of this embodiment, in the component separation device for extracting genes contained in a sample, a syringe including a cylinder and a piston to provide negative pressure and positive pressure by the piston to inhale and discharge the sample, and a syringe of the syringe. A tip is connected to and fixed at one end, and a tip for extracting a component extracts genes from the sample by sucking in the sample by negative pressure provided from the syringe and discharging the sample by positive pressure, and other functions of the tip for extracting the component. A portable component separation device is provided, which includes a tip sealer coupled to the end to prevent foreign substances from entering the interior of the component extraction tip.

Description

Tip for extracting ingredients and portable ingredient extraction device including the same {Tip for Extracting Target Ingredient and Portable Extraction Device Comprising the Same}

The present invention relates to a tip for extracting ingredients that allows a user to carry it and separate and extract specific ingredients right on site, and a portable ingredient extraction device including the same.

The content described in this section simply provides background information for this embodiment and does not constitute prior art.

Viral infectious diseases such as SARS (Severe Acute Respiratory Syndrome), MERS (Middle East Respiratory Syndrome), or the recently prevalent coronavirus (COVID-19) have a high fatality rate and are highly contagious through the respiratory tract, so early diagnosis is required. It is important to prevent further infection and damage.

Currently, the method mainly used to detect viruses is molecular diagnostics, which detects the cause of the disease and whether or not it is infected by detecting the nucleic acid (DNA/RNA or their variants) of bacteria, viruses, etc. that cause the disease. This is a method for detecting. This molecular diagnostic method involves collecting samples from body fluids, sputum, or tissue samples, extracting genes from the collected samples, and amplifying and analyzing the genes using polymerase chain reaction (PCR). It consists of steps.

At this time, one of the conventional extraction methods required centrifugation to extract genes from the collected sample. However, because centrifugation is difficult to provide at a virus testing site and is located in a specific location, conventional gene extraction methods have had the problem of completely extracting genes on site.

This problem also exists in other conventional extraction methods, such as magnetic separation technology. Magnetic separation technology is a method of adding magnetic particles to a cell mixture to induce bonding between nucleic acids or proteins and magnetic particles to form a complex, and then using an external magnetic field to selectively separate only the nucleic acids or proteins bound to the magnetic particles. am. However, this method also has the problem of making it difficult to immediately extract genes on site in that an external magnetic field must be provided to separate nucleic acids.

Recently, various methods have been attempted to easily extract genes using filter or column structures without centrifugation or magnetic fields, but problems such as reduced gene recovery efficiency and filter clogging due to impurities in the sample still exist. It is not resolved. Therefore, there is still a need to secure a device that can secure recovery efficiency and reliability in a filter or column structure separation method.

One embodiment of the present invention aims to provide a component extraction tip and a portable component extraction device that can easily and completely extract specific components, such as genes, in the field.

According to one aspect of the present invention, in a component separation device for extracting genes contained in a sample, a syringe including a cylinder and a piston to provide negative pressure and positive pressure by the piston to inhale and discharge the sample, and a syringe of the syringe A tip is connected to and fixed at one end, and a tip for extracting a component extracts genes from the sample by sucking in the sample by negative pressure provided from the syringe and discharging the sample by positive pressure, and other functions of the tip for extracting the component. A portable component separation device is provided, which includes a tip sealer coupled to the end to prevent foreign substances from entering the interior of the component extraction tip.

According to one aspect of the present invention, the tip for component extraction includes a tip holder inside one end connected to the syringe tip, and the syringe tip and the tip for component extraction are fixed by the tip holder. Do it as

According to one aspect of the present invention, the tip for extracting ingredients includes a filler, a first opening, and a second opening.

According to one aspect of the present invention, the filler includes an extraction layer for extracting genes contained in a sample, and upper and lower fixation layers disposed at both ends of the extraction layer to fix the extraction layer.

According to one aspect of the present invention, the extraction layer is implemented with silica beads having a particle size of 75 to 150㎛, and when the sample is introduced into the interior by the negative pressure provided from the syringe, only the genes contained in the incoming sample are extracted. It is characterized by filtering and adsorption.

According to one aspect of the present invention, the upper and lower fixed layers are formed of glass fibers, and the glass fibers filled in the upper fixed layer of the filler have an area of 32 mm2 to 64 mm2 for a tip for extracting components with a capacity of 1 ml. It is characterized by having.

According to one aspect of the present invention, the gene is a nucleic acid.

According to one aspect of the present invention, in the component extraction tip for extracting genes contained in a sample, there is a first opening formed at one end to allow the sample to flow into the internal space, and the sample can be introduced through the first opening. A component disposed in the inner space of the tip and a second opening connected to the other end to provide negative pressure, and comprising a filler for extracting genes from the sample flowing in through the first opening. Extraction tips are provided.

According to one aspect of the present invention, the filler includes an extraction layer for extracting genes contained in a sample, and upper and lower fixation layers disposed at both ends of the extraction layer to fix the extraction layer.

According to one aspect of the present invention, the extraction layer is implemented with silica beads having a particle size of 75 to 150㎛, and when the sample is introduced into the interior by the negative pressure provided from the syringe, only the genes contained in the incoming sample are extracted. It is characterized by filtering and adsorption.

According to one aspect of the present invention, the upper and lower fixed layers are formed of glass fibers, and the glass fibers filled in the upper fixed layer of the filler have an area of 32 mm2 to 64 mm2 for a tip for extracting components with a capacity of 1 ml. It is characterized by having.

According to one aspect of the present invention, in the method of extracting genes from a sample by a portable component separation device, the sample is mixed with a crushing solution, a first volume of the mixed crushing solution is sucked, and the sample is disposed in a tip for component extraction. A step of isolating the genes contained in the sample while contacting the filler with the sample, sucking a washing solution, washing the fragmentation solution components remaining inside the filler, and sucking the elution solution to separate the separated genes from the filler. A gene extraction method comprising the step of excreting is provided.

According to one aspect of the present invention, the filler includes an extraction layer for extracting genes contained in a sample, and upper and lower fixation layers disposed at both ends of the extraction layer to fix the extraction layer.

As described above, according to one aspect of the present invention, specific components such as genes can be completely extracted on site without undergoing a process such as centrifugation, which has the advantage of improving test speed and convenience.

In addition, according to one aspect of the present invention, the portable component separation device has the advantage of ensuring the stability of the test by preventing contamination by foreign substances and maintaining the sterile state of the device until the genetic component is extracted.

Figure 1 is a diagram showing the configuration of a portable component separation device according to a first embodiment of the present invention.
Figure 2 is a diagram showing the configuration of a tip for extracting ingredients and a filler in the tip according to an embodiment of the present invention.
Figure 3 is a diagram showing the configuration of a tip sealer coupled to a tip for extracting ingredients according to an embodiment of the present invention.
Figure 4 is a diagram showing the configuration of a tip holder for fixing a tip for component extraction according to another embodiment of the present invention.
Figure 5 is a diagram showing the configuration of a portable component separation device according to a second embodiment of the present invention.
Figure 6 is a flowchart showing a method for separating components by a portable component separation device according to an embodiment of the present invention.
Figure 7 is a diagram showing the results of gene extraction rate (C _q ) analysis according to the amount of extraction layer filled in the filling material of the tip for component extraction according to an embodiment of the present invention.
Figure 8 is a diagram showing the results of analysis of gene extraction performance according to the length of the upper fixed layer among the fillers of the tip for component extraction according to an embodiment of the present invention.
Figure 9 is a diagram showing the results of gene extraction performance analysis of a portable component separation device according to an embodiment of the present invention and a conventional centrifugal component separation device.

Since the present invention can make various changes and have various embodiments, specific embodiments will be illustrated in the drawings and described in detail. However, this is not intended to limit the present invention to specific embodiments, and should be understood to include all changes, equivalents, and substitutes included in the spirit and technical scope of the present invention. While describing each drawing, similar reference numerals are used for similar components.

Terms such as first, second, A, and B may be used to describe various components, but the components should not be limited by the terms. The above terms are used only for the purpose of distinguishing one component from another. For example, a first component may be named a second component, and similarly, the second component may also be named a first component without departing from the scope of the present invention. The term and/or includes any of a plurality of related stated items or a combination of a plurality of related stated items.

When a component is said to be "connected" or "connected" to another component, it is understood that it may be directly connected to or connected to the other component, but that other components may exist in between. It should be. On the other hand, when it is mentioned that a component is “directly connected” or “directly connected” to another component, it should be understood that there are no other components in between.

The terms used in this application are only used to describe specific embodiments and are not intended to limit the invention. Singular expressions include plural expressions unless the context clearly dictates otherwise. In this application, terms such as "include" or "have" should be understood as not precluding the existence or addition possibility of features, numbers, steps, operations, components, parts, or combinations thereof described in the specification. .

Unless otherwise defined, all terms used herein, including technical or scientific terms, have the same meaning as generally understood by a person of ordinary skill in the technical field to which the present invention pertains.

Terms defined in commonly used dictionaries should be interpreted as having a meaning consistent with the meaning in the context of the related technology, and unless explicitly defined in the present application, should not be interpreted in an ideal or excessively formal sense. No.

Additionally, each configuration, process, process, or method included in each embodiment of the present invention may be shared within the scope of not being technically contradictory to each other.

Figure 1 is a diagram showing the configuration of a portable component separation device according to a first embodiment of the present invention.

Referring to Figure 1, the portable component separation device 100 according to the first embodiment of the present invention includes a tip for extracting components (Tip, 110), a tip sealer (120), and a tip holder (Tip Holder, 130). and syringe 140.

The portable component separation device 100 (hereinafter abbreviated as component separation device) is a device that can extract genes, especially DNA or RNA as nucleic acid, from a sample containing the gene of a virus to be detected. Since the component separation device 100 can extract genes without a separate device right at the scene of collecting samples, it can ensure speed and convenience in extracting genes for testing.

The component separation device 100 includes a component extraction tip 110, a tip sealer 120, a tip holder 130, and a syringe 140.

The component extraction tip (Tip, 110, hereinafter abbreviated as tip) includes a filler 116 and openings 113 and 119, and is connected to the syringe 140 to supply negative pressure to the sample. Genes are extracted from the inhaled sample by (140).

The tip 110 is hollow on the inside and includes openings 113 and 119 at both ends, through which a sample (solution containing the sample) containing the virus gene can be introduced. The tip 110 may have a capacity ranging from 200 μl to 1 ml.

An opening 119 is formed at one end of the tip 110, so that the syringe tip 142 of the syringe 140 can be inserted and coupled to the opening 119. An opening 113 is formed at the other end to allow the solution containing the sample to be delivered by a change in pressure of the syringe 140 and flow into the tip 110.

In particular, the shape of the tip 110 is not particularly limited, but is a commonly used conical shape, and the tip 110 is formed from the opening 119 located at one end of the tip 110 to the opening 113 at the other end. The diameter may continuously decrease or decrease intensively in a certain section.

The tip 110 includes a filler 116 at a position close to the opening 113 at the other end (within a preset distance from the opening 113). When the syringe 140 connected to the tip 110 provides negative pressure to the sample and inhales the sample into the interior of the tip 110 (via the opening 113), the filler 116 absorbs only the genes contained in the sample. Filter and adsorb. The specific structure of the filler 116 is shown in FIG. 2.

Figure 2 is a diagram showing the configuration of a filler in a tip for extracting ingredients according to an embodiment of the present invention.

Referring to FIG. 2, the filler 116 according to an embodiment of the present invention includes an upper fixed layer 210, an extraction layer 220, and a lower fixed layer 230.

The upper fixed layer 210 and the lower fixed layer 230 are disposed at both ends of the extraction layer 220 to fix the extraction layer 220.

The upper fixed layer 210 and lower fixed layer 230 have low resistance to fluid, so that the solution containing the sample can be smoothly sucked in and discharged through the upper and lower fixed layers 210 and 230, and the solution containing the sample can be smoothly absorbed and discharged. It can be implemented as a component that can be fixed in place even when inhaled and discharged, and as an example, it can be glass fiber.

In particular, the upper and lower fixed layers 210 and 230 do not lift or move even when the sample is sucked into the tip 110 by the operation of the syringe 140, thereby fixing the extraction layer 220 and allowing the sample to be extracted into the extraction layer 220. Pass the sample smoothly so that the sample can be transferred.

The amount of filling of the upper and lower fixed layers 210 and 230 in the inner space of the tip 110 may also affect the sample suction speed and gene extraction performance. If a relatively small amount of the fixed layer is filled, the resistance during suction and discharge of the sample can be reduced, but it is likely to be lifted by the pressure applied from the syringe 140 during the suction or discharge process, and on the contrary, if a large amount of the fixed layer (210, 230), the resistance increases when inhaling and discharging samples using a syringe, which may affect gene extraction performance.

As described above, the fixing layers 210 and 230 applied to the filler 116 of the component separation device 100 of the present invention may be a conjugate pad made of glass fiber. As an example, for the component extraction tip 110 with a capacity of 1 ml, the upper fixed layer 210 may be a glass fiber pad with an area in the range of 32 to 96 ㎟, and the lower fixed layer 230 may be 10 to 15 ㎟. Fiberglass pads with a range of areas can be used.

The extraction layer 220 is disposed between the upper and lower fixed layers 210 and 230 to extract genes contained in the sample. The extraction layer 220 is implemented with a component that has a large surface area and resistance, such as silica beads, and is capable of contacting the sample with as much surface area as possible and extracting specific components from the sample. For example, the extraction layer 220 may be implemented with silica beads having a particle size in the range of 40 to 150 ㎛, and more specifically, the particle size may be in the range of 75 to 150 ㎛. If the extraction layer 220 is implemented with silica beads having a relatively small particle size, the resistance may increase and the extraction amount may relatively increase. On the other hand, when the extraction layer 220 is implemented with silica beads with a relatively large particle size, the resistance is reduced and the suction speed can be relatively fast.

In addition, the amount of silica beads filled in the extraction layer 220, like the particle size range of the beads, may affect the resistance, speed, and extraction amount when inhaling and discharging the solution containing the sample from the filling material 160. For example, when a 1 ml component extraction tip 110 is used, the silica beads filled in the extraction layer 220 may be filled in the range of 25 to 50 μl, and may be filled in the range of 30 to 45 μl. It can be.

As silica beads having an appropriate size according to the purpose are placed in the extraction layer 220 in an appropriate amount, the extraction layer 220 does not flow between the upper and lower fixed layers 210 and 230, and the upper and lower fixed layers 210 and 230 do not flow. Only the genes to be extracted from the sample that has passed through the fixing units 210 and 230 can be extracted by the extraction layer 220.

Referring again to FIG. 1, the tip 110 for extracting ingredients may have a tip sealer 120 coupled to the opening 113 at the other end.

The tip sealer 120 is a stopper that protects the sample suction portion of the component extraction tip 110, where the tip 110 is in direct contact with the sample and solution for suction of the solution for extracting genes from the sample. It prevents the filler 116 provided inside the opening 113 and the tip 110 from being contaminated.

The tip sealer 120 is coupled to the tip 110 so that the opening 113 at the other end of the component extraction tip 110 can be completely sealed, and the gene is extracted from the sample using the component separation device 100. When extraction and separation are performed, it is separated from the tip 110. Accordingly, the tip sealer 120 may be made of a silicone rubber material having a certain level of elasticity so that it can be completely sealed from the external environment and easily separated during use. The specific structure of the tip sealer 120 is shown in FIG. 3.

Figure 3 is a diagram showing the configuration of a tip sealer coupled to a tip for extracting ingredients according to an embodiment of the present invention.

Referring to FIG. 3, the tip sealer 120 includes a sealer body 310 and a tip insertion hole 320.

Since the tip sealer 120 is a stopper into which the other end of the tip 110 for extracting ingredients is inserted, the sealer body 310 has a tip insertion hole 320 into which the end on the opening 113 side of the tip 110 is inserted. Includes.

As described above, the tip 110 may decrease in diameter continuously or section by section from the opening 119 to the opening 113. Accordingly, the tip insertion hole 320 may be implemented in a shape corresponding to the shape of the end of the tip 100 on the open portion 113 side. Additionally, the sealer body 310 may have a shape whose diameter continuously decreases from one end to the other end, or may have a cylindrical structure. The shape of the sealer body 310 is not limited as long as it has a structure that can completely seal one end of the tip 110 for extracting ingredients.

The total length of the sealer body 320 may be approximately 4 to 6 mm, and the depth of the tip insertion hole 320 into which the end of the tip 110 is inserted may be in the range of 2.5 to 3.5 mm. there is. The size of the tip sealer 120 may be appropriately changed depending on the size of the tip 110 for extracting ingredients.

Referring again to FIG. 1, the tip 110 for extracting ingredients is connected to the syringe 140 by a tip holder (Tip Holder, 130). The tip holder 130 is disposed near the opening 119 at one end of the tip 110 and completely secures the tip 110 to the syringe 140. Since the tip holder 130 must be able to maintain the coupling state between the tip 110 and the syringe 140 despite changes in pressure provided by the syringe 140, the tip holder 130 can be formed of a general sealing material. And, as an example, a silicone rubber material having a certain level of elasticity may be used. The specific structure of the tip holder 130 is shown in FIG. 4.

Figure 4 is a diagram showing the configuration of a tip holder (Tip Holder) for fixing a tip for component extraction according to an embodiment of the present invention.

Referring to FIG. 4 , the tip holder 130 includes a holder body 420 and a through hole 440.

Since the tip holder 130 is disposed inside the tip 110, it is implemented in the same form as the tip 110. As described above, the tip 110 may have a diameter that decreases continuously or section by section from the opening 119 at one end to the opening 130 at the other end. Accordingly, the holder body 410 of the tip holder 130 has an upper surface 410 whose diameter is larger than the diameter of the lower surface 430, so that it can be placed inside the tip 110. In particular, the holder body 410 may have a structure in which the diameter continuously decreases from the upper surface 410 to the lower surface 430.

The holder body 410 includes a through hole 440 in the center, and the syringe tip 142 of the syringe 140 coupled to the opening 119 of the tip 110 is inserted into the through hole 440. Make it possible. The shape of the through hole 440 is the same as that of the syringe tip 142, and the diameter of the through hole 440 on the upper surface 410 side is the same as the diameter of the through hole 440 on the lower surface 430 side, and is cylindrical. It can be formed into a structure. In addition, the diameter of the through hole 440 is the same as or slightly larger than the diameter of the syringe tip 142, so that the syringe 140 and the tip 110 are maintained despite pressure changes due to the movement of the piston 146 of the syringe 140. Make sure to secure the syringe tip 142 so that it does not come off.

Referring again to FIG. 1, the syringe 140 includes a syringe tip 142, a cylinder 144, and a piston 146, and the sample (solution containing the sample) is divided into components by the up and down reciprocating motion of the piston 146. It can be inhaled through the extraction tip 110 or discharged to the outside.

In order to extract specific components such as genes from a sample, the crushing solution, washing solution, and elution solution must be sequentially introduced into the tip 110 for extracting the components. At this time, the crushing solution, washing solution, and elution solution to be introduced. The quantities are different.

A sufficient amount of disruption solution must be sucked into the tip 110 so that sufficient genes can be extracted from the sample, and a sufficient amount of washing solution must also be sucked into the tip 110 so that the disruption solution can be sufficiently washed. Additionally, it is advantageous for the elution solution required to elute the extracted gene to be inhaled only in an amount sufficient to elute the gene. Accordingly, in the present invention, the syringe 140 is used to extract genes from a sample by using a quantitative amount of the solutions required for each step.

The syringe tip 142 is connected to the component extraction tip 110 by being inserted into the through hole 440 of the tip holder 130.

The cylinder 144 accommodates a solution containing a sample and includes a scale marked with numbers on the outer circumferential surface so that the volume of the solution can be recognized as a quantitative amount. Using this, the suction and discharge of the solution for sample and gene separation are quantitatively determined. so that it can be accomplished.

The piston 146 can form negative pressure and positive pressure as it is pushed or pulled through up and down reciprocating motion. Through this process, the solution containing the sample can be sucked in so that genes can be separated from the sample as it passes through the component extraction tip 110, or the elution solution can be discharged to extract the separated genes.

The portable component separation device 100 of the present invention can extract genes from a sample without using centrifugation, and provides accurate results by applying a syringe 140 structure so that each solution required for the extraction process can be used in a quantitative amount. can be derived.

The portable component separation device 100 is equipped with a tip sealer 120 and a tip holder 130 at both ends of the component extraction tip 110, and is a device for extracting genes. It blocks the inflow of foreign substances and is sterilized before use. status can be maintained.

Furthermore, the portable component separation device 100 is a structure of a filler 116 for gene extraction from a sample, and the extraction layer 220 is disposed between the upper and lower fixed layers 210 and 230, using a conventional silica bead or fiber filter. Extraction performance can be improved compared to an extraction layer consisting of only the structure. A method for extracting genes from a sample using the portable component separation device 100 will be described later with reference to FIG. 6.

Figure 5 is a diagram showing the configuration of a portable component separation device according to a second embodiment of the present invention.

Referring to Figure 5, the portable ingredient separation device 500 according to the second embodiment of the present invention includes a tip 510 for ingredient extraction, a tip sealer 520, a tip holder 530, and a syringe 540. , the detailed configuration of each configuration is the same as that of the first embodiment.

However, the syringe 540 of the second embodiment includes three scales marked on the outer peripheral surface of the cylinder 544 to correspond to the usage amounts of the crushing solution, washing solution, and elution solution, respectively. Through this, when using the portable component separation device 500, genes can be easily separated from the sample just by displaying the scale for each solution without having to check the volume of each solution numerically.

Figure 6 is a flowchart showing a method for separating components by a portable component separation device (100, 500) according to an embodiment of the present invention.

A sample is collected and mixed with the crushing solution (S610).

The sample may be saliva, for example, saliva or nasal discharge. Such samples are collected by a specific collection device. If the holder of the collected sample is a carrier, the sample contains the gene of the virus to be tested.

The collected sample is mixed with lysis buffer. Here, the disruption solution is a solution that separates genes (DNA or RNA as nucleic acids) and other proteins contained in the sample. As the collected sample and the crushing solution are mixed, all components in the collected sample are crushed. At this time, mixing of the sample and the crushing solution may be performed without a separate incubation process, but is not limited to this and may further include an incubation process for a preset time (e.g., 10 minutes) if necessary. It may also be carried out. As a result, all components in the collected sample are crushed.

A preset amount of the sample mixed with the disruption solution is sucked and only the genes are separated (S620).

The portable component separation devices 100 and 500 provide negative pressure to the sample crushed by the crushing solution by the reciprocating motion of the piston 146 of the syringe 140. As described above, the piston 146 is pulled and the sample mixed with the crushing solution is sucked into the component extraction tip 110 and passes through the filler 116. The sample passes through the filler 116, and genes in the sample are extracted from the extraction layer 220. The crushing solution containing the sample sucked into the component separation devices 100 and 500 is discharged to the outside through the opening 113 of the tip 110 by pushing the piston 146.

This process can be repeated, and genes in the sample are extracted from the extraction layer 220 within the filler 116. The syringe 140 and the tip 110 can be combined and fixed, negative pressure is transmitted to the sample by the syringe 140, and the sample can be sucked into the tip 110, and the filler 116 allows the gene to be absorbed into the sample. Since can be extracted, the portable component separation devices 100 and 500 can extract genes from the sample even without separately performing centrifugation.

According to the conventional method, the genes in the sample could be extracted only after the disruption solution mixed with the sample was injected into the gene isolation container and the container was centrifuged.

The component separation devices 100 and 500 suck in a preset amount of washing buffer to wash the residue in the filler 116 (S630). After the component separation device (100, 500) extracts the gene for a sufficient period of time, the component separation device (100, 500) sucks the cleaning solution through the opening 113 of the tip 110. As the sample is sucked into the tip 110, not only the gene to be extracted but also the fragmentation solution components may remain in the filler 116. The washing solution is a solution for washing the crushing solution. In order to remove the crushing solution components remaining in the filler 116, the component separation device 100 sucks the washing solution. Similar to the above-described process (S620), the cleaning solution is sucked in and discharged into the tip 110 by the reciprocating motion of the piston 146 in the syringe 140, and through this process, the crushing solution is cleaned.

According to the conventional method, after the washing solution was injected into the gene isolation vessel, additional centrifugation had to be performed before the disruption solution could be washed.

The component separation device 100 sucks in the elution buffer and discharges the separated genes (S640). After completing the above-described washing process, the component separation device 100 inhales the elution solution using the syringe 120. The elution solution separates the genes extracted from the filler 116 from the filler 116 and elutes them. As the elution solution is sucked into the tip 110, the genes extracted from the filler 116 are separated and discharged.

Genes isolated in this way can be tested through an amplification process using polymerase chain reaction (PCR).

As described above, the component separation device 100 includes a tip 110 and a syringe 140, and can extract genes from a sample by simply inhaling and discharging each solution into the tip 110.

Hereinafter, we will look at the gene extraction performance according to the configuration of the filler 116 of the tip 110 for component extraction in the portable component separation devices 100 and 500 through examples.

Example 1: Comparison of gene extraction performance according to the amount of silica beads in the extraction layer

In order to confirm gene extraction performance according to the structure of the filler 116 included in the tip 110 for component extraction of the present invention, gene extraction performance according to the filling amount of the silica beads constituting the extraction layer 220 was measured using a component separation device. It was measured through the example of (100), and the extraction performance according to the filling amount of silica beads can be confirmed in Figure 7.

To compare the extraction performance of genes, the COVID-19 standard sample (ZeptoMetrix, 50 TCID ₅₀ /ml) was used.

The crushing solution was prepared with 4M Guanidine thiocyanate, 27.5mM Tris-Cl (pH 7.4), 12.5mM EDTA, 1.5% Triton It was manufactured with. Additionally, 2mM NaOH was used as the elution solution.

The component separation device 100 used a tip 110 for component extraction and a syringe 140 with a capacity of 1 ml. The upper fixed layer 210 and lower fixed layer 230 included in the filler 116 of the ingredient extraction tip 110 used conjugate pads made of glass fiber, and had an area of 64㎟ and 12㎟, respectively. A fixed layer was constructed with a pad having .

The silica beads constituting the extraction layer 220 are beads with a particle diameter ranging from 75 to 150㎛, and the amount filled between the upper and lower fixed layers 210 and 230 is 9, 18, 27, 36, and 45. It was changed to ㎕ and placed on the component extraction tip 110, and genes were extracted from the sample using this.

Based on the COVID-19 standard sample, qPCR was performed for each amount of silica beads in the filler 116, and the extraction rate (C _q ) and the time required to aspirate 1 ml of sample were measured.

The process of extracting genes from the sample was performed through the processes S610 to S640 described above in FIG. 6. At this time, 14 ㎕ of the sample is mixed with 1,000 ㎕ of the crushing solution, and using the component separation device 100, 1,000 ㎕ of the mixed solution is inhaled so that the sample can pass through the filler 116 in the tip 110, and then again discharged to the outside. Afterwards, the process of sucking and discharging 500 ㎕ of the cleaning solution by the up and down reciprocating motion of the piston 146 was repeated to wash the crushing solution remaining in the filler 160. Next, 60㎕ of the elution solution was inhaled and discharged once to isolate the genes in the sample.

Figure 7 is a diagram showing the results of gene extraction rate (C _q ) analysis according to the amount of extraction layer filled in the filling material of the tip for component extraction according to an embodiment of the present invention.

Referring to FIG. 7, it can be seen that as the amount of silica beads filled with the extraction layer 220 increases, the C _q value decreases. In particular, as the C _q value is significantly lower when the silica beads are filled to 36 ㎕ or more, it can be seen that the extraction performance is excellent when the silica beads for gene extraction are filled to the amount of 36 ㎕ and 45 ㎕. . C _q is an index that measures the relative expression level of the target gene. The lower the C _q value, the higher the expression of the target gene, which means that the speed at which the gene is extracted from the sample is faster. In other words, it can be seen that the gene extraction performance improves as the amount of silica beads provided in the filler 160 to separate genes from the sample increases.

In addition to the extraction speed according to the filling amount of silica beads, the time required to inhale 1 ml was also measured, and the results are summarized in Table 1.

As the amount of beads filled in the extraction layer 220 increases, the performance of extracting genes can improve, but as the internal resistance increases, the time required to separate the sample using the component separation device 100 increases. Alternatively, the pressure for the reciprocating movement of the piston 146 may also increase, which may lower the stability of sample extraction.

silica beads
Volume (㎕) C _q 1ml inhalation
Average time (sec) Measures medium 99 Primary 34.14 34.66 150 Secondary 34.19 3rd 35.65 18 Primary 33.97 34.06 95 Secondary 34.07 3rd 34.14 27 Primary 33.57 33.97 91 Secondary 34.21 3rd 34.14 36 Primary 34.11 34.07 93 Secondary 34.08 3rd 34.02 45 Primary 33.61 33.54 108 Secondary 33.58 3rd 33.44

Referring to Table 1, as the amount of silica beads increases, the error in C _q value according to repeated tests also decreases, and it is judged that extraction of genes with relatively high reproducibility can be performed. On the other hand, it can be seen that as the amount of silica beads increases, the resistance of the structure of the filler 116 increases, and the suction time of the sample using the component separation device 100 also increases. Considering the resistance and extraction speed during suction, 1 ml It can be seen that the silica bead extraction layer 220, which may be included in the filler 116 of the tip 110 for extracting ingredients, can achieve optimal performance in a capacity range of 36 to 45 ㎕.

Example 2: Comparison of gene extraction performance according to the filling amount of the upper fixed layer of filler

Since the tip 110 for extracting ingredients of the present invention performs a process of suctioning and discharging a solution containing a sample several times, the filler 116 must be fixed so as not to flow despite pressure changes inside the tip 110. In particular, since the upper fixed layer 210 directly receives the negative and positive pressures caused by the up-and-down reciprocating motion of the piston 146, it needs to be filled in an amount that does not cause the tip 110 to be excited by the pressure.

However, like the silica beads of the extraction layer, an increase in the amount of glass fiber material filled with the upper fixed layer 210 increases the resistance during sample suction/discharge, thereby reducing the resistance during suction/discharge and preventing it from flowing due to pressure changes. The filling amount of the upper fixed layer 210 must be optimized within a range that is not present.

In Example 2, gene extraction performance was compared according to changes in the filling amount of the upper fixed layer 210, and the results can be seen in FIG. 8.

Figure 8 is a diagram showing the analysis results of gene extraction performance according to the area of the upper fixed layer among the fillers of the tip for component extraction according to an embodiment of the present invention.

Example 2, for comparing the gene extraction performance of the component separation device 100 according to the area of the upper fixed layer 210, included the same sample (COVID-19, ZeptoMetrix, 50 TCID ₅₀ /ml) and crushing solution as Example 1, Washing solution and elution solution were applied, and the process of separating and extracting genes from the sample was also performed through steps S610 to S640 described above in FIG. 6.

The tip 110 for extracting ingredients had a capacity of 1 ml, and the filler 116 was made of the same materials as in Example 1, and the silica beads of the extraction layer 220 were filled with a volume of 45 ㎕. Gene extraction was performed from the sample while varying the filling amount of the upper fixed layer 210 and the area of the glass fiber pad disposed within the tip 110. The area of the upper fixed layer 210 was varied by adjusting the length while maintaining the width of the glass fiber pad filled inside the tip 110 constant (4 mm).

Figure 8 (a) shows the C _q value when extracting genes from a sample while increasing the length of the upper fixed layer 210 from 8 mm to 64 mm, and the extraction amount (End Point) at the time amplification is completed in the qPCR reaction. This is one result, and Figure (b) shows the time required to inhale 1 ml according to the length of the upper fixed layer 210.

Referring to FIG. 8, it can be seen that when a pad with an area of 64 mm2 (4 × 16 mm) is applied to the upper fixed layer 210, the lowest C _q value and fastest suction time are achieved. Afterwards, as the area of the pad increased, the C _q value also increased, and it was confirmed that the time required for suction also increased as the resistance increased. In addition, the filling amount of the upper fixed layer 210 used in the 1 ㎖ ingredient extraction tip is 32㎟ (4 × _8㎜ ) to 96㎟ (4 It is judged that the application with an area of ×24 mm) is the optimal range to secure extraction performance while stably maintaining the filler 116.

From Examples 1 and 2, the optimal range for the structure of the filler 116 of the tip 110 for component extraction was selected to measure the gene separation performance of the component separation device 100 of the present invention for different types of samples. And the results of analyzing the accuracy are shown in Figure 9 by comparing them with the results separated and extracted using a conventional centrifugation method.

Figure 9 shows the results of gene extraction performance analysis of a portable component separation device according to an embodiment of the present invention and a conventional centrifugal component separation device.

Figure 9(a) shows the results of component extraction by centrifugation when an Influenza A standard sample was used as a sample (comparative example) and the component equipped with a tip 110 for component extraction according to an embodiment of the present invention. The results (example) of component extraction using the separation device 100 are shown, and Figure 9(b) shows the extraction results of the comparative example and example when using the COVID-19 standard sample as the sample.

Separation and extraction of components of samples including Influenza A and COVID-19 by the component separation device 100 according to an embodiment of the present invention were performed through the processes of S610 to S640 described above, including crushing solution, washing solution, and elution. The solution was used in the same composition and dosage as in Example 1.

In addition, the filler 116 disposed inside the tip 110 for extracting ingredients is a conjugate pad made of glass fiber with an area of 64 mm2 and 12 mm2 and is disposed on the upper and lower fixing layers 210 and 230. In the extraction layer 220, 45 μl of silica beads with a particle size ranging from 75 to 150 μm were placed.

Referring to FIG. 9, the C _q value of an example in which genes were separated and extracted from a sample using the component extraction tip 110 according to an embodiment of the present invention is the result of separation and extraction using a conventional centrifugation method. It can be seen that they are almost similar, and this can be seen as a value within the error range of the value extracted in the same way. This means that the portable component separation device 100 equipped with the component extraction tip 110 of the present invention has no lower extraction performance compared to a device that separates components through a conventional centrifugal separation method.

That is, the component extraction tip 110 of the present invention and the component separation device 100 including the same are a component extraction tip ( 110), target genes can be separated and extracted from samples simply and quickly.

In Figure 6, each process is described as being sequentially executed, but this is merely an illustrative explanation of the technical idea of an embodiment of the present invention. In other words, a person skilled in the art to which an embodiment of the present invention pertains can change the order depicted in each drawing or perform one or more of the processes without departing from the essential characteristics of an embodiment of the present invention. Since various modifications and variations can be applied by executing in parallel, FIG. 6 is not limited to a time series order.

Meanwhile, the processes shown in FIG. 6 can be implemented as computer-readable codes on a computer-readable recording medium. Computer-readable recording media include all types of recording devices that store data that can be read by a computer system. That is, computer-readable recording media include storage media such as magnetic storage media (eg, ROM, floppy disk, hard disk, etc.) and optical read media (eg, CD-ROM, DVD, etc.). Additionally, computer-readable recording media can be distributed across networked computer systems so that computer-readable code can be stored and executed in a distributed manner.

The above description is merely an illustrative explanation of the technical idea of the present embodiment, and those skilled in the art will be able to make various modifications and variations without departing from the essential characteristics of the present embodiment. Accordingly, the present embodiments are not intended to limit the technical idea of the present embodiment, but rather to explain it, and the scope of the technical idea of the present embodiment is not limited by these examples. The scope of protection of this embodiment should be interpreted in accordance with the claims below, and all technical ideas within the equivalent scope should be interpreted as being included in the scope of rights of this embodiment.

100, 500: Portable component separation device
110, 510: Tips for extracting ingredients
113, 119, 513, 519: opening
116, 516: Filler
120, 520: Tip sealer
130, 530: Tip holder
140, 540: syringe
142, 542: syringe tip
144, 544: cylinder
146, 546: Piston
210: upper fixed layer
220: extraction layer
230: Lower fixed layer
310: Sealer body
320: Tip insertion hole
410: upper surface
420: Holder body
430: If you do
440: Through hole

Claims (13)

In a component separation device for extracting genes contained in a sample,
A syringe including a cylinder and a piston that provides negative and positive pressure by the piston to inhale and discharge the sample;
A syringe tip of the syringe is connected and fixed to one end, and a component extraction tip for extracting genes from the sample by sucking in the sample by negative pressure provided from the syringe and discharging the sample by positive pressure; and
A tip sealer that is coupled to the other end of the ingredient extraction tip to prevent foreign substances from entering the ingredient extraction tip.
A portable component separation device comprising a. According to paragraph 1,
The tip for extracting the above ingredients is,
A portable component separation device comprising a tip holder inside one end connected to the syringe tip, and fixing the syringe tip and the component extraction tip by the tip holder. According to paragraph 1,
The tip for extracting the above ingredients is,
A portable component separation device comprising a filler, a first opening, and a second opening. According to paragraph 3,
The filler is,
A portable component separation device comprising an extraction layer for extracting genes contained in a sample, and upper and lower fixed layers disposed at both ends of the extraction layer to fix the extraction layer. According to paragraph 4,
The extraction layer is,
Implemented with silica beads with a particle size of 75 to 150㎛,
A portable component separation device that filters and adsorbs only genes contained in the incoming sample when the sample is introduced into the interior by negative pressure provided from the syringe. According to paragraph 4,
The upper and lower fixed layers are formed of glass fiber,
A portable component separation device, characterized in that the glass fiber filled in the upper fixed layer of the filler has an area of 32㎟ to 64㎟ for a 1㎖ component extraction tip. According to paragraph 1,
The gene is,
A portable component separation device characterized by nucleic acid. In the component extraction tip for extracting genes contained in a sample,
a first opening formed at one end to allow the sample to flow into the internal space;
a second opening where a syringe that provides negative pressure to allow the sample to flow through the first opening is connected to the other end; and
A tip for extracting ingredients, which is disposed in the inner space of the tip and includes a filler for extracting genes from a sample flowing through the first opening. According to clause 8,
The filler is,
A tip for extracting components, comprising an extraction layer for extracting genes contained in a sample, and upper and lower fixed layers disposed at both ends of the extraction layer to fix the extraction layer. According to clause 9,
The extraction layer is,
Implemented with silica beads with a particle size of 75 to 150㎛,
A tip for component extraction, characterized in that when a sample is introduced into the interior by negative pressure provided from the syringe, only the genes contained in the inflow sample are filtered and adsorbed. According to clause 9,
The upper and lower fixed layers are formed of glass fiber,
A tip for extracting ingredients, characterized in that the glass fibers filled in the upper fixed layer of the filler have an area of 32 mm2 to 64 mm2 for a tip for extracting ingredients with a capacity of 1 ml. In the method of extracting genes from a sample by a portable component separation device,
Mixing a sample with a crushing solution, inhaling a first volume of the mixed crushing solution, and contacting the sample with a filler disposed in a tip for extracting components to separate genes contained in the sample;
Suctioning a cleaning solution to wash the crushing solution components remaining inside the filler; and
Inhaling the elution solution to discharge the separated gene from the packing material
A gene extraction method comprising: According to clause 12,
The filler is,
A gene extraction method comprising an extraction layer for extracting genes contained in a sample, and upper and lower fixed layers disposed at both ends of the extraction layer to fix the extraction layer.

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