KR101817671B1 - Extraction and purification apparatus - Google Patents

Abstract

The present invention relates to an extractive separation and purification apparatus comprising: a sample vessel having a chamber in which a sample or a buffer is received; An orbital shaker for horizontally locating the sample vessel on which the sample vessel is placed; A bore capable of being introduced into the chamber of the sample vessel placed on the orbital shaker; A bore driving part for moving the bore to a chamber of the sample vessel; A magnetic rod inserted into and withdrawn from the bore to attach magnetic beads received in a chamber of the sample container to a surface of the bore or to detach magnetic beads attached to a surface of the bore to a chamber of the sample container; And a rod driving unit for inserting and withdrawing the magnetic rod into the bore.

Description

[0001] EXTRACTION AND PURIFICATION APPARATUS [0002]

The present invention relates to an extract separation and purification apparatus, and more particularly, to an extract separation and purification apparatus for separating or purifying various kinds of biological substances such as nucleic acids or antibodies.

Generally, two key technologies applied to the automatic extraction equipment of extracts contained in solution are a method of inserting a silica membrane according to a substance to which a biological substance such as a nucleic acid or an antibody to be extracted is bound and purified, A spin column method in which a biological substance such as nucleic acid is attached to a substrate and then purified by centrifugal force and a magnetic bead method in which a magnetic bead or a magnetic particle is put into a sample to extract a desired substance .

Although automation equipment using a centrifugal separation method using a silica membrane spin column is popularly used, a magnetic bead method that can be easily implemented without using a centrifuge is progressing gradually.

As described above, the magnetic extraction method used for the automatic nucleic acid extraction uses magnetic particles or paramagnetic particles, and biological materials such as nucleic acid or protein in the high concentration salt solution are added to the magnetic particles Is used to extract high-purity nucleic acid containing impurities in the dissolved cells.

As an example, a specific procedure of nucleic acid extraction using a magnetic bead method is as follows. When magnetic beads are introduced into a sample container in which cells or biological materials are dissolved, and a bore connected to the robot arm moves rapidly in the melt To flow the melt. In this process, nucleic acid or other substances are attached to the magnetic beads by collision of magnetic beads with biological materials such as nucleic acids and antibodies.

After inserting the magnet into the bore, only the magnetic beads in the melt are bound to the surface of the bore which has a magnetic force.

After the magnetic beads are transferred to another sample container containing a washing buffer, the magnet is lifted from the bore, and then the magnet is lifted to immerse the magnetic beads attached to the bore into the cleaning solution contained in the sample vessel. Releasing the magnetic bead by washing with the flow of the fluid by the lifting motion and washing the magnetic bead.

After the washing of the magnetic beads is completed, the bore is inserted into the sample container containing the worn magnetic beads in the state that the magnet is inserted again in the bore, and the magnetic beads, which are worn by the magnetic force of the magnet, are attached again to the surface of the bore .

Through these procedures, the washed magnetic beads are put into a sample container containing an elution buffer to separate biological materials such as nucleic acids and antibodies, and the magnetic beads are collected and finished.

In such extraction processes, a key process for extracting the maximum amount of substance to be extracted and for high purity extraction is to use a biological material such as a nucleic acid, an antibody, and the like to flow the fluid necessary for attachment and separation of magnetic beads, And to maximize the flow required for the binding and releasing of the magnetic beads in the bore with and without the magnetic beads.

However, due to the splashing of the sample during the flow of the sample in the sample vessel due to the lift of the bore, not only the sample is lost but also the periphery of the apparatus is contaminated and the sample is caused to flow due to the limit of the bore lifting speed There is an extremely limited problem. As a result, the adsorption and separation of biological substances such as nucleic acids, antibodies and the like into magnetic beads do not occur effectively, and the maximum extraction of high purity does not occur due to insufficient flow of magnetic beads and related buffers. When a large amount of nucleic acid is extracted from the sample, the sample container, the bore, and the magnet are each increased in proportion to the amount of the sample for sufficient flow of the sample, and a small amount of illu- mination including biological substances such as concentrated nucleic acid, There is a problem that it is not easy to obtain a buffer.

US Patent No. 5,200,084 (entitled " APPARATUS AND METHODS FOR MAGNETIC SEPARATION, " PCT International Application Publication No. WO 2001/68263 A1 (entitled VESSEL AND ROD, published on September 20, 2001).

Disclosure of the Invention The present invention has been made to solve the above problems, and it is an object of the present invention to improve the attachment and separation efficiency of magnetic beads and extracts and to maximize the flow of a sample or buffer even in a sample container having a large or small volume, The present invention also provides a method for separating and purifying an extract.

An object of the present invention is to provide a sample container having a chamber in which a sample or a buffer is accommodated; An orbital shaker for horizontally locating the sample vessel on which the sample vessel is placed; A bore capable of being introduced into the chamber of the sample vessel placed on the orbital shaker; A bore driving part for moving the bore to a chamber of the sample vessel; A magnetic rod inserted into and withdrawn from the bore to attach magnetic beads received in a chamber of the sample container to a surface of the bore or to detach magnetic beads attached to a surface of the bore to a chamber of the sample container; And a rod driving unit for inserting and withdrawing the magnetic rod into the bore.

Here, the bore driving unit includes: a bore supporting bracket for supporting the bore; A screw shaft for a bore, which is screw-rotatably coupled to the bore support bracket, for lifting the support bracket for the bore; And a bore driving motor for rotating the screw shaft for the bore.

The rod driving unit includes: a rod supporting bracket for supporting the magnetic rod; A rod screw shaft rotatably coupled to the rod support bracket for raising and lowering the rod support bracket; And a rod drive motor for rotating the rod screw shaft.

And a reciprocating driving unit for reciprocally moving the bore driving unit and the rod driving unit in the transverse direction with respect to the lifting direction of the bore.

The reciprocating driver includes: a main support bracket for supporting the bore driver and the rod driver; A main screw shaft rotatably coupled to the main support bracket so as to reciprocate the main support bracket in the transverse direction with respect to the lifting direction of the bore; And a main drive motor for rotating the main screw shaft.

The orbital shaker includes a shaking platform on which the sample container is seated; And a shaking motor for horizontally locating the shaking platform.

And an auxiliary bead having a hardness higher than that of the sample and being introduced into the sample container.

The auxiliary beads may be at least one of a stainless bead, a zirconia bead, a ceramic bead, and a glass bead.

According to the present invention, the sample or the buffer contained in the sample container is caused to flow by the horizontal trajectory motion of the orbital shaker, thereby increasing the flow of the sample or buffer contained in the sample container to improve the attachment and separation efficiency of the magnetic bead and the extract, It is possible to separate and purify the extract at a high concentration from a small amount of the sample contained in the sample container having a large or small diameter.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a block diagram of an apparatus for separating and purifying an extract according to an embodiment of the present invention;
FIG. 2 is a view illustrating a process of collecting magnetic beads having a biological material adsorbed by using a bore having a magnetic rod inserted therein using the extraction and purification apparatus of FIG. 1; FIG.
FIG. 3 is a view illustrating a process of washing a magnetic bead in which a biological material is adsorbed by a horizontal trajectory motion while a magnetic rod is separated from a bore using the extractive separation and purification apparatus of FIG. 1;
FIG. 4 is a view showing a process of releasing a substance such as an antibody or a protein of a magnetic bead of a worn magnetic bead into an elution buffer using the extractive separation and purification apparatus of FIG.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will now be described in detail with reference to the accompanying drawings.

FIG. 1 shows an extract separation and purification apparatus according to an embodiment of the present invention.

As shown in these figures, an extractive separation and purification apparatus 10 according to an embodiment of the present invention includes a sample vessel 11, an orbital shaker 21, a bore 31, a bore driving unit 41, (51), and a rod driver (61).

The sample container 11 has a tubular shape with a hollow circular cross section with one side opened. A chamber 13 accommodating a sample or a buffer is formed in the sample vessel 11. The chamber 13 is a chamber for holding a sample. Here, the sample is collectively referred to as a liquid, a solid, or a mixture of a liquid and a solid in a biological or chemical process. The buffer is collectively referred to as a washing buffer, an elusion buffer, and the like.

The orbital shaker 21 seats the sample container 11 and horizontally moves the sample container 11. The orbital shaker 21 includes a shaking platform 23 and a shaking motor 27.

The shaking platform 23 has a flat plate shape and a plurality of seating grooves 25 on which the sample container 11 is seated is recessed. The plurality of seating grooves 25 are formed at regular intervals such that a plurality of sample vessels 11 are seated with an interval therebetween.

The shaking motor 27 is supported on the base 81 and the shaking motor 27 is directly connected to the shaking platform 23 by a coupling (not shown) to horizontally move the shaking platform 23.

The sample or buffer contained in the chamber 13 of the sample container 11 placed on the shaking platform 23 flows on the wall surface of the sample container 11 by the driving of the shaking motor 27, The sample or the buffer can be mixed and flowed. In addition, a high concentration of the extract can be separated and purified from a large amount or a small amount of the sample contained in the sample container 11 having a large or small diameter.

The bore 31 has a tubular shape with a hollow circular cross section with one side opened. The bore 31 has a relatively smaller diameter than the sample container 11 and is thereby insertable into the sample container 11 and is capable of moving the sample or buffer between the sample container 11 and the inner circumference of the sample container 11 Thereby forming a space. In addition, a magnetic rod 51 is inserted into and drawn out from the inside of the bore 31.

The bore driving part 41 moves the bore 31 to the chamber 13 of the sample container 11. The bore driving section 41 includes a bore supporting bracket 43, a bore screw shaft 45, and a bore driving motor 47.

The support bracket 43 for the bore has an arm shape and supports the bore 31 in a suspended manner.

The screw shaft 45 for the bore has a rod shape having a predetermined length and a male screw threaded to the bore support bracket 43 is formed on the outer periphery of the screw shaft 45 for the bore. The screw shaft 45 for the bore is screwed to the support bracket 43 for the bore to raise and lower the support bracket 43 for bore.

The bore driving motor 47 is directly connected to the bore screw shaft 45 by a coupling (not shown) to rotate the bore screw shaft 45 in the forward and reverse directions. The bore support bracket 43 is moved up and down toward the sample container 11 and the bore 31 disposed above the sample container 11 is moved upward and downward relative to the sample container 11 To the chamber 13.

The magnetic rod 51 has a rod shape of a predetermined length and is inserted into and drawn out from the bore 31. The magnetic rod 51 is coaxial with the bore 31 and is disposed above the bore 31. At one end of the magnetic rod 51, a magnetic body 53 for generating a magnetic force for coupling with the magnetic bead by magnetic force is provided. When the magnetic rod 51 is inserted into the bore 31, the magnetic beads received in the chamber 13 of the sample container 11 are bonded to the surface of the bore 31 by the magnetic force of the magnetic rod 51 )do. In contrast, when the magnetic rod 51 is drawn out from the bore 31, the magnetic beads attached to the surface of the bore 31 are separated from the surface of the bore 31 as the magnetic force of the magnetic rod 51 is lost, (13) of the chamber (11).

The rod driver 61 inserts the magnetic rod 51 into the bore 31 and draws it out. The rod drive unit 61 includes a rod support bracket 63, a rod shaft 65 for a rod, and a drive motor 67 for a rod.

The rod support bracket 63 has an arm shape and supports the magnetic rod 51 in a suspended manner.

The rod screw shaft 65 has a rod shape having a predetermined length. On the outer circumference of the rod screw shaft 65, a male screw threadably engaged with the rod support bracket 63 is formed. The rod screw shaft 65 is screwed to the rod support bracket 63 so as to move the rod support bracket 63 up and down.

The rod driving motor 67 is directly connected to the rod screw shaft 65 by a coupling (not shown) to rotate the rod screw shaft 65 in the forward and reverse directions. The rod support bracket 63 is moved up and down toward the sample container 11 as the rod screw shaft 65 rotates in the forward and reverse directions and the magnetic rod 51 disposed above the bore 31 is moved in the bore 31, As shown in Fig.

The apparatus 10 for separating and purifying an extract according to an embodiment of the present invention includes a bore driving part 41 and a reciprocating part for reciprocating the rod driving part 61 in the transverse direction with respect to the lifting direction of the bore 31 71).

The reciprocating unit 71 includes a main support bracket 73, a main screw shaft 75, and a main drive motor 77.

The main support bracket 73 supports the bore driving portion 41 and the rod driving portion 61. More specifically, the bore driving motor 47 and the rod driving motor 67 are supported at one end of the main support bracket 73 in this embodiment, and the bore screw shaft 45 and the rod screw The shaft 65 is threaded through. A main screw shaft 75 is threadedly engaged with the other end of the main support bracket 73.

The main screw shaft 75 has a rod shape having a predetermined length. On the outer periphery of the main screw shaft 75, a male screw threadably engaged with the main support bracket 73 is formed. The main screw shaft 75 is screw-rotatably coupled to the main support bracket 73 so that the main support bracket 73 is reciprocally moved in the transverse direction with respect to the lifting direction of the bore 31, The rod 51 is reciprocally moved in the transverse direction with respect to the lifting direction of the bore 31. [

The main drive motor 77 is directly connected to the main screw shaft 75 by a coupling (not shown) to rotate the main screw shaft 75 in the forward and reverse directions. The bore 31 and the magnetic rod 51 are moved back and forth in the sample container (not shown) by reciprocating the main support bracket 73 in the transverse direction with respect to the lifting direction of the bore 31 as the main screw shaft 75 rotates in the forward / 11, or is spaced apart from the sample container 11. [ The main drive motor 77 is supported on the base 81 and the shaking motor 27 of the orbital shaker 21 is also supported on the base 81.

A process for separating or purifying various kinds of biological substances such as a nucleic acid or an antibody as an extract from a sample using the extractive separation and purification apparatus 10 according to an embodiment of the present invention will now be described. Respectively.

First, a process of separating a biological material such as a nucleic acid or an antibody from a sample 1 containing a biological substance such as a nucleic acid or an antibody will be described with reference to FIG.

A first sample container 11a containing a sample 1 containing biological substances such as a nucleic acid or an antibody to be separated is placed on the shaking platform 23 of the orbital shaker 21. Here, after placing the first sample container 11a on the shaking platform 23, the sample 1 in which biological substances such as nucleic acids or antibodies to be separated are dissolved can be accommodated in the first sample container 11a .

Then, the magnetic beads 7 are received in the chamber 13 of the first sample container 11a. The orbital shaker 21 is horizontally traversed to cause bead beating with the magnetic beads 7 and biological materials such as dissolved nucleic acid or antibody in the sample 1 to cause biological beads such as nucleic acid or antibody Thereby binding the materials to the magnetic beads 7. Further, at least one of a stainless bead, a zirconia bead, a ceramic bead, and a glass bead is further added as an auxiliary bead having a hardness higher than that of the sample, Bacteria, tissue cells, solid materials such as feces, or semi-solid materials are destroyed to extract biological substances such as nucleic acids or antibodies. That is, in addition to the magnetic beads 7, an auxiliary bead having a hardness higher than that of the sample 1 is additionally injected into the first sample container 11a and horizontally traversed to thereby crush the sample such as a solid or semi-solid material A biological material such as an internal nucleic acid or an antibody is released, and then attached to the magnetic beads 7, followed by a washing process and a purification process to remove the nucleic acid or antibody, Antibodies and other biological substances.

Subsequently, the main drive unit is driven to position the bore 31 and the magnetic rod 51 above the first sample container 11a.

Then, the rod drive motor 67 is driven so that the magnetic rod 51 is inserted into the bore 31.

Subsequently, the shaking motor 27 of the orbital shaker 21 is driven to horizontally move the first sample container 11a placed on the shaking platform 23.

At the same time, the bore driving motor 47 and the rod driving motor 67 of the bore driving unit 41 are driven so that the bore 31 and the magnetic rod 51 rise and descend together in the chamber 13 of the first sample container 11a. And rotate them together.

As described above, the sample 1 contained in the first sample container 11a is moved horizontally by the locus motion of the orbital shaker 21 so that the sample 1 of the chamber 13 is moved along with the magnetic beads 7, The cell wall or the outer membrane surrounding the biological material such as the nucleic acid or the antibody contained in the sample 1 is destroyed to separate the biological material such as the nucleic acid and the antibody from the sample 1 The magnetic beads 7 attached to the magnetic beads 7 and adsorbed with biological substances such as nucleic acids and antibodies are attracted to the surface of the bore 31 by the magnetic force of the magnetic rod 51 inserted into the bore 31 For example, one end surface of the bore 31 located in the chamber 13 of the first sample container 11a. At this time, by magnetic force of the magnetic rod 51 ascending and descending in the chamber 13 together with the bore 31, the magnetic beads mixed with the sample 1 flowing in the up and down direction on the wall surface of the first sample container 11a, (7) can be evenly attached to the surface of the bore (31).

When the attachment of the magnetic beads 7 to the surface of the bore 31 is completed, the bore driving motor 47 and the rod driving motor 67 are driven to move the bore 31 and the magnetic rod 51 Together with the chamber 13 of the first sample container 11a.

This completes the process of separating biological material such as nucleic acid or antibody from the sample 1 and attaching it to the magnetic beads 7.

Hereinafter, the process of washing and magnetizing the magnetic beads 7 separated from the sample 1 and adsorbing the biological material as described above will be described.

First, the main driving motor 77 is driven to accommodate the washing buffer 3 and is placed above the second sample container 11b placed on the shaking platform 23 of the orbital shaker 21 as shown in FIG. 3 The bore 31 and the magnetic rod 51 to which the magnetic beads 7 are attached are moved on the surface of the bore 31 as shown in FIG.

Next, the bore driving motor 47 and the rod driving motor 67 are driven to house the bore 31 and the magnetic rod 51 together in the chamber 13 of the second sample container 11b.

Then, the rod drive motor 67 is driven so that the magnetic rod 51 is spaced apart from the bore 31.

The magnetic beads 7 to which the biological substances such as nucleic acids and antibodies are adsorbed can be released from the surface of the bore 31 as the magnetic force of the magnetic rod 51 is lost and the second sample container 11b To the washing buffer 3 of the washing machine.

The shaking motor 27 is driven to horizontally move the second sample container 11b so that the magnetic beads 7 accommodated in the second sample container 11b are detached from the bore 31, . At this time, the foreign substances adsorbed to the magnetic beads 7 together with the biological substances such as nucleic acids and antibodies are separated from the magnetic beads 7 by the washing buffer 3 and separated from the magnetic beads 7 by the washing of the second sample container 11b And remains in the buffer 3.

The bore 31 and the magnetic rod 51 are received in the chamber 13 of the second sample container 11b while the magnetic rod 51 is inserted into the bore 31 again, (See FIG. 4) attached to the surface of the bore 31 by the magnetic force of the first sample container 11c (see FIG. 4) ), And performs the final process of the illusion process.

The main drive motor 77 is driven to drive the bore 31 and the magnetic rod 51 attached with the magnetic beads 9 worn on the surface of the bore 31 to the solution buffer 5, Is moved upward of the third sample container (11c) accommodated. The third sample container 11c may be seated on the shaking platform 23 and the third sample container 11c may be seated on the centrifuge.

Next, the bore driving motor 47 and the rod driving motor 67 are driven to house the bore 31 and the magnetic rod 51 together in the chamber 13 of the third sample container 11c.

Then, the rod drive motor 67 is driven so that the magnetic rod 51 is spaced apart from the bore 31.

At this time, the magnetic force of the magnetic rod 51 is lost as the magnetic rod 51 is separated from the bore 31, and the magnetic beads 9 on which biological substances such as nucleic acid and antibody are adsorbed are removed from the surface of the bore 31 And can be dispersed into the solution buffer 5 of the third sample container 11c.

The shaking motor 27 is driven to horizontally move the third sample container 11c to move the magnetic beads 9 to the surface of the bore 31 in the solution buffer 5 accommodated in the third sample container 11c And biological materials such as nucleic acids and antibodies adsorbed on the magnetic beads 9 are released to separate biological materials such as nucleic acids and antibodies from the magnetic beads 9.

When the solving is completed, the bore 31 and the magnetic rod 51 are positioned above the third sample container 11c, though not shown, by driving the main drive.

Then, the rod drive motor 67 is driven so that the magnetic rod 51 is inserted into the bore 31.

Next, the bore driving motor 47 and the rod driving motor 67 are driven so that the bore 31 and the magnetic rod 51 are received together in the chamber 13 of the third sample container 11c.

At this time, pure magnetic beads (not shown) in which biological substances such as nucleic acids and antibodies are separated by the solution buffer 5 are attached to the surface of one end of the bore 31 by the magnetic force of the magnetic rod 51.

The bore driving motor 47 and the rod driving motor 67 are driven so that the bore 31 and the magnetic rod 51 are spaced apart from the chamber 13 of the third sample container 11c.

As a result, only a biological substance such as a pure nucleic acid or an antibody remains in the third sample container l1c, and the extraction and purification process of a biological substance such as a nucleic acid or an antibody is completed as an extract.

As described above, according to the present invention, the sample or the buffer accommodated in the sample container flows by the horizontal trajectory motion of the orbital shaker, thereby increasing the flow of the sample or buffer contained in the sample container and improving the attachment and separation efficiency of the magnetic bead and the extract .

As the sample or the buffer accommodated in the sample container flows by the horizontal trajectory movement of the orbital shaker, the sample or the buffer accommodated in the chamber of the sample container flows along the wall surface of the sample container, so that the sample or the buffer can be mixed and flowed at high speed , And it is also possible to separate and purify the extract at a high concentration from a large amount or a small amount of sample housed in a sample container having a large or small diameter. Also, even if a magnetic bead having a small magnetic force and a bore having a small diameter are used, a large amount of magnetic beads contained in a sample or a buffer can be attached to the bore, and the efficiency of attachment and separation of the extract can be improved.

In addition, the apparatus for separating and purifying an extract according to the present invention may further comprise a step of additionally adding a hard bead (stainless bead), a zirconia bead, a ceramic bead, a glass bead, etc., Homogenizer function and extraction function are combined into one when the biological material such as nucleic acid or antibody is extracted by breaking the specimen such as solid substance such as bacteria, Enabling one-step automation to be implemented.

On the other hand, in the above-described embodiment, one sample container is placed on the orbital shaker, and the bore is moved up and down simultaneously with horizontal trajectory movement. However, the present invention is not limited to this, and a plurality of sample containers may be placed in line on the orbital shaker, It is possible to horizontally move the sample or the buffer put in the sample container at the same time, and simultaneously raise and lower the bore, so that a large number of samples can be simultaneously separated and purified.

10: Extract separation and purification apparatus 11, 11a, 11b, 11c: Sample container
13: chamber 21: orbital shaker
23: shaking platform 27: shaking motor
31: bore 41: bore drive
43: Support bracket for bore 45: Screw shaft for bore
47: Driving motor for bore 51: Magnetic rod
61: rod driving part 63: rod supporting bracket
65: screw shaft for rod 67: driving motor for rod
71: reciprocating portion 73: main support bracket
75: main screw shaft 77: main drive motor

Claims (8)

A sample container having a chamber in which a sample or a buffer is accommodated;
An orbital shaker for horizontally locating the sample vessel on which the sample vessel is placed;
A bore capable of being introduced into the chamber of the sample vessel placed on the orbital shaker;
A bore driving part for moving the bore to a chamber of the sample vessel;
A magnetic rod inserted into and withdrawn from the bore to attach magnetic beads received in a chamber of the sample container to a surface of the bore or to detach magnetic beads attached to a surface of the bore to a chamber of the sample container;
A rod drive unit for inserting and withdrawing the magnetic rod into the bore;
And a reciprocating part for reciprocating the bore driving part and the rod driving part in the transverse direction with respect to the lifting direction of the bore. The method according to claim 1,
The bore-
A support bracket for the bore for supporting the bore;
A screw shaft for a bore, which is screw-rotatably coupled to the bore support bracket, for lifting the support bracket for the bore;
And a bore driving motor for rotating the screw shaft for bore. The method according to claim 1,
Wherein the rod-
A rod supporting bracket for supporting the magnetic rod;
A rod screw shaft rotatably coupled to the rod support bracket for raising and lowering the rod support bracket;
And a rod driving motor for rotating the screw shaft for rod. delete The method according to claim 1,
The reciprocating unit includes:
A main support bracket for supporting the bore driver and the rod driver;
A main screw shaft rotatably coupled to the main support bracket so as to reciprocate the main support bracket in the transverse direction with respect to the lifting direction of the bore;
And a main drive motor for rotating the main screw shaft. The method according to claim 1,
The orbital shaker may further comprise:
A shaking platform on which the sample container is seated;
And a shaking motor for horizontally locating the shaking platform. The method according to claim 1,
Further comprising an auxiliary bead having a hardness higher than that of the sample and charged into the sample vessel. 8. The method of claim 7,
Wherein the auxiliary beads are at least one of stainless beads, zirconia beads, ceramic beads, and glass beads.

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