JP2011072276A - Reagent cartridge and nucleic acid purification kit - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a reagent cartridge and a nucleic acid purification kit in which liquid sticking to the outer surface of dispensing tips is properly removed with a simple structure. <P>SOLUTION: The reagent cartridge 100 containing the liquid for separating and purifying the nucleic acid from specimen includes an oil removing part 128 which removes the oil 127A sticking to the outer surface of the cylindrical dispensing tip 201 which carries out sucking, holding and discharging the liquid. The oil removing part 128, into which the dispensing tip 201 is inserted and in which a groove 129C making contact with the outer surface of the dispensing tip 201 is formed, includes the wiping filter 129A which absorbs the oil 127A. <P>COPYRIGHT: (C)2011,JPO&INPIT

Description

  The present invention relates to a reagent cartridge and a nucleic acid purification kit.

  With recent developments in genetic testing technology, for example, nucleic acid can be extracted from a sample such as a biological sample collected from a patient, and genetic differences such as single nucleotide polymorphisms can be detected to predict drug sensitivity in advance. Sex has been suggested.

  When extracting nucleic acids from a specimen, in order to prevent cross-contamination between specimens or to prevent the spread of infection sources, the reagents necessary for extracting nucleic acids from the specimen are contained in a reagent cartridge, Every time nucleic acid is extracted from a subject, these reagents are used up and the reagent cartridge is discarded (see, for example, Patent Document 1).

  Further, the liquid contained in the reagent cartridge is sucked and held by a dispensing tip having a cavity inside, and dispensed or stirred according to a predetermined process. At this time, the liquid may adhere to the outer surface of the dispensing tip in the step of sucking the liquid into the dispensing tip or the step of discharging the liquid from the dispensing tip. When the liquid adheres to the outer surface of the dispensing tip, there is a possibility that the accuracy of measuring the liquid using the dispensing tip may be reduced, or the contamination due to the liquid adhering to the outer surface of the dispensing tip may spread. For this reason, an automatic analyzer for analyzing nucleic acids is equipped with an apparatus for cleaning the outer surface of a dispensing tip (see, for example, Patent Document 2).

International Publication No. 2005/118772 JP 2003-215828 A

However, since the nucleic acid analyzer described in Patent Document 1 cannot remove the liquid adhering to the outer surface of the dispensing chip, a measurement error may occur due to the liquid adhering to the outer surface of the dispensing chip.
Moreover, in the automatic analyzer described in Patent Document 2, there is a problem that the apparatus configuration becomes complicated because the dispensing tip is vibrated to remove the cleaning liquid adhering to the outer surface of the dispensing tip.

  The present invention has been made in view of the above-described circumstances, and an object thereof is to provide a reagent cartridge and a nucleic acid purification kit that can remove liquid adhering to the outer surface of a dispensing chip with a simple configuration.

In order to solve the above problems, the present invention proposes the following means.
The reagent cartridge of the present invention is a reagent cartridge that contains a liquid for separating and purifying nucleic acid from a specimen and dispenses the liquid using a dispensing chip, and is attached to the outer surface of the dispensing chip. A liquid removing unit that removes excess liquid is provided, and the liquid removing unit includes a wiping member that can insert the dispensing tip and is formed with a through-passage that contacts the outer surface of the dispensing tip and absorbs the excess liquid. It is characterized by having.

  Moreover, it is preferable that the said wiping member is formed with the porous material which has elasticity.

  Moreover, it is preferable that the said wiping member is formed with the cut | notch part expanded by the said dispensing tip.

  Moreover, it is preferable that the said wiping member has a through-hole which has an internal diameter smaller than the largest external shape of the area | region which can contact the said liquid in the said dispensing tip.

  Moreover, it is preferable that the said liquid is oil and the said wiping member has lipophilicity.

  In addition, the reagent cartridge includes a sample storage unit that stores the sample, a liquid storage unit that stores the liquid, a waste liquid storage unit that stores a waste liquid generated in the separation and purification, and the nucleic acid of the sample. It is preferable to further have an extraction filter cartridge for purifying the above.

  The nucleic acid purification kit of the present invention comprises the reagent cartridge of the present invention and a dispensing chip container for housing a plurality of the dispensing chips.

  According to the reagent cartridge and the nucleic acid purification kit of the present invention, since the excess liquid is absorbed by the wiping portion in the liquid removing portion provided in the reagent cartridge, the liquid adhering to the outer surface of the dispensing tip can be removed with a simple configuration.

It is a perspective view which shows the reagent cartridge of one Embodiment of this invention. It is a perspective view which shows the structure of the reagent cartridge and the dispensing tip rack. It is sectional drawing which shows the structure of the extraction filter cartridge in the reagent cartridge. (A) is sectional drawing which shows the structure of the liquid removal part in the reagent cartridge, (B) is a disassembled perspective view which shows the structure of a part of liquid removal part. (A) thru | or (C) is a top view which shows the structure of the wiping filter in the liquid removal part. (A) And (B) is explanatory drawing for demonstrating the effect | action of the liquid removal part in the reagent cartridge.

Hereinafter, a reagent cartridge and a nucleic acid purification kit according to an embodiment of the present invention will be described with reference to FIGS.
FIG. 1 is a perspective view showing a reagent cartridge 100 according to an embodiment of the present invention. FIG. 2 is a perspective view showing the configuration of the reagent cartridge 100 and the dispensing tip rack 200. FIG. 3 is a cross-sectional view showing the configuration of the extraction filter cartridge 150 in the reagent cartridge 100. 4A is a cross-sectional view showing a configuration of the liquid removing unit 128 in the reagent cartridge 100, and FIG. 4B is an exploded perspective view showing a partial configuration of the liquid removing unit 128. 5A to 5C are plan views showing the configuration of the wiping filter 129A in the liquid removing unit 128. FIG. 6A and 6B are explanatory diagrams for explaining the operation of the liquid removing unit 128 in the reagent cartridge 100. FIG.

  As shown in FIGS. 1 and 2, the nucleic acid purification kit 10 contains a plurality of reagent cartridges 100 containing reagents for extracting nucleic acids from a specimen and a plurality of dispensing chips 201 for dispensing liquids. And a dispensed tip rack (dispensing tip container) 200. In this embodiment, the dispensing tip rack 200 includes a plurality of dispensing tips 201 of the same shape and the same size, and the liquid stored in the reagent cartridge 100 is dispensed or stirred by any of the plurality of dispensing tips 201. In operation, the dispensing tip 201 prevents cross contamination between liquids. The dispensing tip rack 200 is also a container for collecting the dispensing tips 201 after use, and dispenses the dispensing tips 201 as infectious waste after the use of the dispensing tips 201 in the nucleic acid analyzer 1 is finished. Note: The entire tip rack 200 can be discarded.

The reagent cartridge 100 includes a main body 101 formed in a substantially box shape and a claw portion 102 formed so as to protrude from the outer surface of the main body 101. For example, when the reagent cartridge 100 is set in an automatic analyzer or the like, the claw portion 102 can be engaged with a part of the automatic analyzer so that the reagent cartridge 100 does not fall down.
A thin sealing film 103 that is removed when the nucleic acid purification kit 10 is used is attached to a part of the outer surface of the main body 101. The opening of the main body 101 is sealed by the sealing film 103, so that an extraction filter cartridge 150, which will be described later, disposed inside the main body 101 does not fall from the main body 101 and foreign substances such as dust are present inside the main body 101. It can be prevented from mixing.

  Inside the main body 101, a sample well (subject housing portion) 110 into which a subject such as a biological sample is put, a reagent well portion 120 containing a reagent for extracting nucleic acid from the subject, A waste liquid well (waste liquid storage unit) 130 for discarding an unnecessary solution separated in the step of extracting nucleic acid from the subject and a recovery well 140 for recovering the nucleic acid extracted from the subject are integrally formed. .

  The reagent cartridge 100 has an extraction filter cartridge 150 containing a carrier that adsorbs nucleic acids, and the reagent cartridge 100 is integrally formed with a holding portion 160 in which the extraction filter cartridge 150 is accommodated.

  The reagent well section 120 includes a plurality of reagent wells (reagent storage sections) 121, 122, 123, 124, 125, 126, an oil well (oil storage section) 127, and an oil removal section (liquid removal section) 128. is doing. In the reagent well portion 120, the openings of the plurality of reagent wells 121, 122, 123, 124, 125, 126 and the oil well 127 are sealed with the sealing film 104. The sealing film 104 is preferably configured such that gas permeation is suppressed and the film can be broken by piercing the dispensing tip 201. For example, a metal thin film or a plastic film is used. Can do.

The reagent wells 121 to 126 include a lysis solution 121A that dissolves biological materials such as cell membranes, a lysis solution 122A that dissolves biological materials such as cytoplasm that cannot be dissolved by the lysis solution 121A and clogs the carrier, and carriers. Washing solutions 123A and 124A for washing away unnecessary substances other than the nucleic acid adsorbed on the eluate, an eluent 125A for eluting nucleic acid from the carrier, and a diluting solution 126A for adjusting the nucleic acid concentration in the eluate are provided in each reagent well It is housed individually.
In the usage mode described later, the analysis reagent is arranged on the nucleic acid analysis chip. Alternatively, the analysis reagent may be stored in a reagent well. For example, an analysis reagent premix in which a part of reagents for performing PCR and SNP measurement by Invader (registered trademark) method on nucleic acid is mixed in advance can be individually accommodated in each reagent well.

  The oil well 127 contains, for example, a well-known oil 127A that is used as a layer on a reaction solution in a PCR reaction. As the oil 127A, for example, mineral oil or silicon oil can be preferably used.

  As shown in FIG. 4A, the oil removing unit 128 has a wiping portion 129 for removing oil 127A adhering to the outer surface of the dispensing tip 201 (see FIG. 2).

As shown in FIGS. 4A and 4B, the wiping portion 129 includes a wiping filter 129A having lipophilicity and a cylindrical shape that supports the wiping filter (wiping member) 129A inside the oil removing portion 128. And supporting portions 129B and 129D. The wiping filter 129A is formed in a substantially cylindrical shape or a substantially disc shape along the inner diameter of the oil removing portion 128, and a cut portion formed so as to penetrate the wiping filter 129A in the center axis direction of the wiping filter 129A. 129C is provided.
The wiping filter 129A is preferably formed of a porous member having elasticity. For example, a wiping filter formed by compressing a sponge filter or the like can be used. The main thing to be used is rubber sponge, urethane sponge, polyethylene foam, EVA sponge, etc. However, this is not limited as long as it is porous and can be elastically deformed.

  As shown in FIG. 5A, the cut portion 129C is located in the center of the wiping filter 129A when the wiping filter 129A is viewed in the central axis direction, and is formed in a cross shape. As shown in FIG. 5B, instead of the cut portion 129C, it may have a cut portion 129E formed in a true circle when the wiping filter 129A is viewed in the central axis direction. Further, as shown in FIG. 5C, instead of the cut portion 129C, a circular cut portion 129F having a bulging portion bulging radially inward of the wiping filter 129A may be provided.

  As shown in FIG. 2, the waste well 130 is a recess formed along the outer diameter shape of the extraction filter cartridge 150 and has a shape capable of supporting the extraction filter cartridge 150. In the state where the extraction filter cartridge 150 is attached to the waste well 130, the extraction filter cartridge 150 does not fall within the reagent cartridge 100.

  The recovery well 140 can support the extraction filter cartridge 150 in the same manner as the waste well 130. The bottom of the recovery well 140 has a container shape that can store the nucleic acid solution eluted from the carrier of the extraction filter cartridge 150 by the eluent 125A.

  The waste liquid well 130 and the recovery well 140 are provided adjacent to each other in the reagent cartridge 100. This is to shorten the flow line of the extraction filter cartridge 150 when the extraction filter cartridge 150 is moved to the recovery well 140 after the extraction filter cartridge 150 is washed in the waste liquid well 130. Thereby, the possibility that the extraction filter cartridge 150 passing over the reagent cartridge 100 contaminates the reagent cartridge 100 or the like can be reduced.

  As shown in FIG. 3, the extraction filter cartridge 150 includes a substantially cylindrical main body 151 serving as an outer frame of the extraction filter cartridge, and an extraction filter unit 152 provided inside the main body 151.

The main body 151 has an upper end 151A and a lower end 151B that are both open. Further, the main body 151 is formed in a funnel shape having an opening diameter smaller than the opening on the upper end 151A side on the lower end 151B side with respect to the extraction filter unit 152, and a nozzle-like discharge port 151C is formed on the lower end 151B in the downward direction. Protrusively provided.
The opening on the upper end 151A side is supplied with the lysing solutions 121A and 122A, the cleaning solutions 123A and 124A, the eluent 125A and the like in a state where the specimen is dissolved. These liquids pass through the filter unit 152 and are discharged from the outlet 151C.

  The filter unit 152 includes an adsorption filter 152A containing a carrier having the property of adsorbing nucleic acids, and a support member 152B that is disposed on the lower end 151B side of the adsorption filter 152A and prevents deformation of the adsorption filter 152A.

  The adsorption filter 152A is formed in a film shape with a porous material capable of adsorbing nucleic acids. The material of the adsorption filter 152A is preferably a material having a property that the nucleic acid is adsorbed in the cleaning liquids 123A and 124A, and the adsorbed state of the nucleic acid is weakened in the eluate 125A. The adsorption filter 152A is preferably a porous material in which a hydroxyl group is introduced as a hydrophilic group. Specifically, the adsorption filter 152A is formed of silica or by combining silica on another substance. The material of the adsorption filter 152A is not particularly limited as long as it is a material that can adsorb a biological substance in the presence of an organic substance. Further, the adsorption filter 152A may be porous by forming a fiber material such as glass wool on top of each other.

  The support member 152B is preferably formed of a material that has at least low adsorptivity to nucleic acids and does not inhibit the reaction of extracting nucleic acids from the specimen. For example, the support member 152B can be formed by baking and hardening resin particles. The rigidity of the support member 152B is higher than that of the adsorption filter 152A, and the support member 152B prevents the adsorption filter 152A from being deformed in the main body 151.

  As shown in FIG. 2, the holding unit 160 is in an initial position where the extraction filter cartridge 150 is accommodated in the reagent cartridge 100. In addition, an absorber (not shown) that absorbs liquid can be provided on the bottom of the holding unit 160. The absorbent body comes into contact with the outer surface of the extraction filter cartridge 150 on the discharge port 151 </ b> C side when the extraction filter cartridge 150 is accommodated in the holding unit 160. For this reason, for example, when the cleaning liquid 123A adheres to the outer surface of the outlet 151C when the cleaning liquid 123A is supplied into the extraction filter cartridge 150, the cleaning liquid 123A can be removed by absorbing the cleaning liquid 123A into the absorber.

  The operation of the reagent cartridge 100 and the nucleic acid purification kit 10 of the present embodiment having the above-described configuration will be described focusing on the operation of the liquid removal unit 128. Hereinafter, as an automatic analyzer to which the nucleic acid purification kit 10 is applied, an automatic analyzer that has a dispensing conveyance mechanism that conveys the dispensing chip 201 and performs PCR analysis after purifying nucleic acid will be described as an example. explain.

  First, the sealing film 103 of the reagent cartridge 100 shown in FIG. 1 is removed manually by the user. Subsequently, for example, a whole blood sample is injected into the sample well 110 of the reagent cartridge 100 manually by the user.

  Subsequently, various reagents stored in the reagent wells 121 to 126 are dispensed and mixed by the dispensing transport mechanism of the automatic analyzer according to a predetermined procedure. Thereby, the cells in the whole blood sample supplied to the sample well 110 are lysed to obtain a cell lysate. When the liquid is sucked into the dispensing tip 201 from the reagent wells 121 to 126, the tip of the dispensing tip 201 is inserted into the sealing film 104 that seals the reagent wells 121 to 126. Then, a through hole is formed in the sealing film 104, and various reagents inside the reagent wells 121 to 126 can be sucked by the dispensing tip 201.

  First, since it is necessary to collect the waste liquid, the extraction filter cartridge 150 is conveyed to the waste liquid well 130. The solution in which the cells are lysed is supplied to the extraction filter cartridge 150. The extraction filter cartridge 150 can feed the gas from the upper end 151A and pressurize the main body 151 to increase the speed at which the liquid passes through the adsorption filter 152A. Then, the solution in which the cells are dissolved passes through the adsorption filter 152A, and the nucleic acid is adsorbed on the adsorption filter 152A. Thereafter, the adsorption filter 152A is washed with a dissolving solution 122A that dissolves biological substances such as cytoplasm that cannot be completely dissolved by the dissolving solution 121A and clog the carrier.

  Further, the cleaning liquids 123A and 124A are supplied to the adsorption filter 152A, and the adsorption filter 152A is cleaned with the cleaning liquids 123A and 124A. Thereafter, the extraction filter cartridge 150 is transported to the recovery well 140, and the eluate 125A is supplied to the adsorption filter 152A. Thereby, the nucleic acid adsorbed by the adsorption filter 152A is eluted in the eluent 125A, and the nucleic acid solution containing the nucleic acid is recovered in the recovery well 140.

Furthermore, the diluent 126A and the eluate 125A from which the recovered nucleic acid is recovered are mixed together, and the sample preparation is completed.
The nucleic acid separation and purification by the nucleic acid purification kit 10 is thus completed.

  Subsequently, using the nucleic acid separated and purified by the nucleic acid purification kit 10, PCR analysis is performed on the nucleic acid. The automatic analyzer sucks the nucleic acid solution with the dispensing chip 201 and supplies the nucleic acid solution to a predetermined reaction container (for example, a microtube). Subsequently, the dispensing tip 201 is inserted into the oil well 127 of the nucleic acid extraction kit 10 shown in FIG. 1, and the oil 127 </ b> A accommodated in the oil well 127 is sucked into the dispensing tip 201. At this time, the oil 127A may adhere to the outer surface of the dispensing tip 201 in the form of droplets because the material of the dispensing tip 201 has a high affinity with the oil 127A.

  The dispensing transport mechanism of the automatic analyzer moves the dispensing tip 201 to the oil removing unit 128 as shown in FIG. Furthermore, as shown in FIG. 6B, the dispensing conveyance mechanism inserts the tip 201A of the dispensing tip 201 into the cut portion 129C of the wiping filter 129A of the oil removing unit 128. Then, the oil 127A adhering to the outer peripheral surface of the tip 201A of the dispensing tip 201 is sucked by the wiping filter 129A and removed from the outer peripheral surface of the dispensing tip 201. By pulling out the dispensing tip 201 from the cut portion 129C, the outer peripheral surface of the dispensing tip 201 comes into contact with the wiping filter 129A again, and the oil 127A attached to the outer peripheral surface of the dispensing tip 201 is wiped off.

The oil 127A is superposed on the nucleic acid solution in the reaction vessel described above by the dispensing tip 201 in which the oil 127A is held and the oil 127A on the outer peripheral surface is removed. At this time, since the excess oil on the outer surface of the dispensing tip 201 is removed, the oil does not adhere to the wall surface of the reaction vessel.
By superposing the oil 127A on the nucleic acid solution in the reaction vessel, the nucleic acid solution is prevented from evaporating in the PCR reaction, and the PCR reaction can be suitably performed.

  As described above, according to the reagent cartridge 100 and the nucleic acid purification kit 10 of the present embodiment, the oil removal unit 128 provided in the reagent cartridge 100 wipes the oil 127A and the filter 129A absorbs it. The oil 127A attached to the outer surface can be suitably removed with a simple configuration.

  Moreover, since the nucleic acid purification kit 10 includes the oil removing unit 128, the oil 127A attached to the outer peripheral surface of the tip 201A of the dispensing tip 201 can be removed by the wiping filter 129A of the oil removing unit 128. It is possible to prevent the liquid from unintentionally adhering to the wall of the reaction vessel and the like, and to improve the accuracy and reproducibility when examining nucleic acids.

  Further, since the wiping filter 129A has elasticity, the oil 127A attached to the outer surface of the dispensing tip 201 can be wiped off by pressing the wiping filter 129A against the outer surface of the dispensing tip 201. Furthermore, since the wiping filter 129A is formed of a porous material, the oil 127A can be suitably absorbed and held in the wiping filter 129A. For this reason, even if the wiping filter 129A is used a plurality of times, the oil 127A can be suitably absorbed.

  In addition, since the reagent and filter unit necessary for extracting the nucleic acid and the oil removing unit 128 are provided inside the reagent cartridge 100 and integrated as a kit, an analyte is added to the sample well 110 for automatic analysis. Since only the operation of setting in the apparatus has only to be performed manually, genetic testing can be performed easily. Further, since the waste liquid well 130 is integrally provided in the reagent cartridge 100, the reagent cartridge 100 may be simply removed from the automatic analyzer and discarded after the genetic test is completed. For this reason, waste liquid processing is simple and there is no possibility that the surroundings will be contaminated by residual liquid such as a subject.

As mentioned above, although embodiment of this invention was explained in full detail with reference to drawings, the concrete structure is not restricted to this embodiment, The design change etc. of the range which does not deviate from the summary of this invention are included.
For example, in the above-described embodiment, an example in which the oil removing unit (liquid removing unit) 128 removes the oil attached to the outer surface of the dispensing tip 201 has been described. Moreover, the liquid adhering to the outer surface of the dispensing tip 201 can be suitably removed. For example, the liquid removal unit of the present invention can be suitably applied when operating glycerol, a surfactant, or the like with high viscosity.

10 Nucleic Acid Purification Kit 100 Reagent Cartridge 128 Oil Remover (Liquid Remover)
129 Wiping part 129A Wiping filter (wiping member)
129C Notch 200 Dispensing tip rack (dispensing tip container)
201 dispensing tips

Claims (7)

A reagent cartridge that contains a liquid for separating and purifying nucleic acid from a specimen and dispenses the liquid using a dispensing chip,
A liquid removing unit for removing excess liquid adhering to the outer surface of the dispensing tip;
The liquid removal section has a wiping member that can insert the dispensing tip and that has a through-passage that contacts the outer surface of the dispensing tip and absorbs the excess liquid.   The reagent cartridge according to claim 1, wherein the wiping member is made of a porous material having elasticity.   The reagent cartridge according to claim 2, wherein the wiping member is formed with a cut portion that is pushed and spread by the dispensing tip.   The reagent cartridge according to claim 2, wherein the wiping member has a through hole having an inner diameter smaller than a maximum outer shape of an area where the liquid can contact in the dispensing tip. The liquid is oil;
The reagent cartridge according to claim 1, wherein the wiping member has lipophilicity. The reagent cartridge is
A subject storage section for storing the subject;
A liquid container for containing the liquid;
A waste liquid storage unit for storing a waste liquid generated in the separation and purification;
An extraction filter cartridge for purifying the nucleic acid of the analyte;
The reagent cartridge according to any one of claims 1 to 5, further comprising: The reagent cartridge according to any one of claims 1 to 6,
A dispensing tip container for housing a plurality of the dispensing tips;
A nucleic acid purification kit comprising:

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