CN114214178A

CN114214178A - Exosome filtering and extracting device and exosome extracting method

Info

Publication number: CN114214178A
Application number: CN202111536643.4A
Authority: CN
Inventors: 黄治国; 方静纯; 姚美玲
Original assignee: Shenzhen Betis Biotechnology Co ltd
Current assignee: Shenzhen Betis Biotechnology Co ltd
Priority date: 2021-12-16
Filing date: 2021-12-16
Publication date: 2022-03-22

Abstract

The invention discloses an exosome filtering and extracting device and an exosome extracting method, relates to the field of biological pharmacy, and aims at solving the problems of low exosome filtering and extracting purity, complex filtering and extracting means and the like. According to the invention, the large-aperture filter membrane and the small-aperture filter membrane are stably placed in the inner wall of the internal bin of the extraction reagent tube, cells are filtered out through the large-aperture filter membrane, impurities such as protein are filtered out through the small-aperture filter membrane, preliminary filtration is realized, the rotation speed of the driving motor is regulated to be high speed, the ultra-separation method is realized through a high-speed and low-speed matched mode, the alternate mode of low-speed centrifugation and high-speed centrifugation is adopted, the operation is simple and convenient, the fault tolerance rate is low, and after the filtration and extraction are finished, the filtered exosome solution is filtered through the 0.1-micron porous membrane, the filtration concentration is improved, and the concentration of the exosome after filtration is ensured.

Description

Exosome filtering and extracting device and exosome extracting method

Technical Field

The invention relates to the field of biological pharmacy, in particular to an exosome filtering and extracting device and an exosome extracting method.

Background

Exosomes are small membrane vesicles containing complex RNA and proteins, are currently regarded as specifically secreted membrane vesicles and participate in intercellular communication, and have 3 major components, namely proteins, nucleic acids and lipids, and play important roles in the physiological and pathological aspects of antigen presentation in immunity, tumor growth and migration, tissue injury repair and the like. Meanwhile, exosomes secreted by different cells have different components and functions, can be used as biomarkers for disease diagnosis, carry a large number of specific proteins (such as cytokines and growth factors) and functional bioactive substances such as mRNAs and miRNAs, participate in physiological processes such as cell communication, cell migration, angiogenesis promotion, anti-tumor immunity and the like in vivo, and are closely related to the occurrence and the progress of various diseases. Due to the special structure and function of the exosome, the exosome has potential application value, on one hand, the exosome can be used as a biological index for diagnosing various diseases, on the other hand, the exosome can also be used as a treatment means, the exosome can be used as a natural carrier of a medicament for clinical treatment in the future, the separation and purification of the exosome are always the concerns of scientific research workers, the obtaining of high-purity exosome is important for subsequent research, most of the existing filtration and extraction of exosome have complex technical means and high cost, and meanwhile, the purification purity of exosome is not high, and the extraction efficiency is low, so that an exosome filtration and extraction device and an exosome extraction method are provided for solving the technical problems.

Disclosure of Invention

Objects of the invention

In order to solve the technical problems in the background art, the invention provides an exosome filtration and extraction device and an exosome extraction method, wherein a large-aperture filter membrane and a small-aperture filter membrane are stably placed in the inner wall of an extraction reagent tube inner bin, cells are filtered out through the large-aperture filter membrane, impurities such as protein and the like are filtered out through the small-aperture filter membrane, preliminary filtration is realized, the rotation speed of a driving motor is adjusted to be high, an ultra-separation method is realized through a high-low speed matching mode, the low-speed centrifugation and high-speed centrifugation alternate mode is simple and convenient to operate, the fault tolerance rate is low, and after filtration and extraction are finished, a filtered exosome solution is filtered through a 0.1-micron porous membrane, so that the filtration concentration is improved, and the concentration of the exosome after filtration is ensured.

(II) technical scheme

The invention provides an exosome filtering and extracting device and an exosome extracting method, which comprise a driving motor cabin body, wherein the driving motor cabin body is hollow, a driving motor is fixedly installed on the inner wall of the bottom of the driving motor cabin body through four screws arranged in an annular array, the input end of the driving motor is fixedly connected with an external power supply, the output end of the driving motor is fixedly provided with a driving rotating shaft penetrating and extending to the top of the driving motor cabin body through a coupler, one end of the driving rotating shaft, which is positioned outside the top of the driving motor cabin body, is welded with a rotating wheel disc, six support columns arranged in an annular array are welded on the periphery of one side of the rotating wheel disc, which is far away from the driving rotating shaft, one end of the support columns, which is far away from the rotating wheel disc, is welded with a fixed lantern ring, and six placing holes arranged in an annular array are arranged on the periphery of the top of the fixed lantern ring, the inner clamping of the placing hole is provided with an extracting reagent tube shell.

As a preferable scheme of the invention, a groove with a gear-shaped cross section is formed in the extracting reagent tube shell, an extracting reagent tube inner bin is sleeved in the groove, and the cross section of the outer ring of the extracting reagent tube inner bin is matched with the cross section of the groove.

As a preferable scheme of the invention, a threaded sleeve is welded on one side of the inner bin of the extraction reagent pipe, which is far away from the outer shell of the extraction reagent pipe, and the outer ring of the threaded sleeve is provided with a plurality of circles of threads.

According to a preferable scheme of the invention, a sealing cover is sleeved on the outer ring of the inner bin of the extraction reagent tube, a groove is formed in the sealing cover, threads meshed with threads on the outer ring of the threaded sleeve are formed on the inner wall of the periphery of the groove, the threads in the sealing cover are aligned to the threads on the outer ring of the threaded sleeve, and then the sealing cover is screwed down, so that a sealing gasket in the sealing cover is in tight contact with the top edge of the threaded sleeve, and an effective sealing effect is further realized.

As a preferred scheme of the invention, a plurality of groups of through holes arranged in an annular array are formed in the top of the rotating wheel disc, the cross sections of the through holes are all in a wedge shape, and the plurality of wedge-shaped through holes formed in the rotating wheel disc aim to reduce the overall proportion of the device and realize light weight, and the fixing lantern ring can be better rotated through the supporting columns.

Compared with the prior art, the technical scheme of the invention has the following beneficial technical effects:

according to the invention, the large-aperture filter membrane and the small-aperture filter membrane are stably placed in the inner wall of the internal bin of the extraction reagent tube, cells are filtered out through the large-aperture filter membrane, impurities such as protein are filtered out through the small-aperture filter membrane, preliminary filtration is realized, the rotation speed of the driving motor is regulated to be high speed, the ultra-separation method is realized through a high-speed and low-speed matched mode, the alternate mode of low-speed centrifugation and high-speed centrifugation is adopted, the operation is simple and convenient, the fault tolerance rate is low, and after the filtration and extraction are finished, the filtered exosome solution is filtered through the 0.1-micron porous membrane, the filtration concentration is improved, and the concentration of the exosome after filtration is ensured.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, it is obvious that the drawings in the following description are only embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to the provided drawings without creative efforts.

Fig. 1 is a schematic structural diagram of an exosome filtering and extracting device and an exosome extracting method provided by the present invention.

Fig. 2 is a schematic view of a fixed sleeve structure of an exosome filtering and extracting device and an exosome extracting method provided by the present invention.

Fig. 3 is a schematic structural diagram of a driving motor cabin body of the exosome filtering and extracting device and the exosome extracting method provided by the invention.

Fig. 4 is a schematic structural diagram of a rotating wheel disc of the exosome filtering and extracting device and the exosome extracting method provided by the present invention.

Fig. 5 is a schematic structural diagram of an internal bin of an extraction reagent tube of the exosome filtration-extraction device and the exosome extraction method provided by the invention.

Fig. 6 is a schematic sectional structure view of an internal chamber of an extraction reagent tube of the exosome filtration-extraction device and the exosome extraction method provided by the invention.

Reference numerals: 1. a motor cabin body is driven; 2. rotating the wheel disc; 3. a support pillar; 4. a fixed collar; 5. extracting the reagent tube housing; 6. a drive motor; 7. driving the rotating shaft; 8. extracting a reagent tube inner bin; 9. a threaded bushing; 10. and (7) sealing the cover.

Detailed Description

The following description is merely exemplary in nature and is not intended to limit the present disclosure, application, or uses. It should be understood that throughout the drawings, identical or similar reference numerals indicate identical or similar parts and features. The drawings are only schematic representations of the concepts and principles of the embodiments of the disclosure, and do not necessarily show specific dimensions or proportions of the various embodiments of the disclosure. Certain features that are part of a particular figure may be exaggerated in order to illustrate relevant details or structures of embodiments of the present disclosure.

The first embodiment is as follows: referring to fig. 1-6, an exosome filtering and extracting device comprises a driving motor cabin 1, the interior of the driving motor cabin 1 is hollow, a driving motor 6 is fixedly installed on the inner wall of the bottom of the driving motor cabin 1 through four screws arranged in an annular array, the input end of the driving motor 6 is fixedly connected with an external power supply, the output end of the driving motor 6 is fixedly installed with a driving rotating shaft 7 penetrating and extending to the top of the driving motor cabin 1 through a coupler, a rotating wheel disc 2 is welded at one end of the driving rotating shaft 7 positioned outside the top of the driving motor cabin 1, six supporting columns 3 arranged in an annular array are welded around one side of the rotating wheel disc 2 away from the driving rotating shaft 7, a fixed lantern ring 4 is welded at one end of the supporting columns 3 away from the rotating wheel disc 2, six placing holes arranged in an annular array are arranged around the top of the fixed lantern ring 4, the inner clamping of the placing hole is provided with an extracting reagent tube shell 5.

Example two: referring to fig. 1-6, on the basis of embodiment one, an exosome filters extraction element, the inside of drawing reagent pipe shell 5 sets up the recess that transversal personally submits the gear form, the inside cover of recess is equipped with draws reagent intraductal storehouse 8, the outer lane cross section of drawing reagent intraductal storehouse 8 agrees with each other with the cross section of recess, draw one side welding that reagent intraductal storehouse 8 kept away from and draws reagent pipe shell 5 has threaded sleeve 9, several circles of screw thread have been seted up to the outer lane of threaded sleeve 9, the outer lane cover of drawing reagent intraductal storehouse 8 is equipped with sealed lid 10, the recess has been seted up to the inside of sealed lid 10, the screw thread with threaded sleeve 9 outer lane screw thread intermeshing has been seted up to the inner wall all around of recess, the top of rotating wheel dish 2 is seted up the multiunit and is the through-hole that annular array arranged, the cross section of through-hole all is the wedge.

Example three: referring to fig. 1-5, on the basis of the first and second embodiments, an exosome filtration extraction device and an exosome extraction method, first, a PEG reagent or a hydrophilic reagent is placed inside an extraction reagent tube inner bin, after slight shaking, a large-aperture filter membrane and a small-aperture filter membrane are sequentially and stably placed in the inner wall of the extraction reagent tube inner bin, and simultaneously, an exosome solution is introduced into the extraction reagent tube inner bin from top to bottom, after the exosome solution is introduced into the extraction reagent tube inner bin, the large-aperture filter membrane arranged inside the extraction reagent tube inner bin filters out cells, then, impurities such as proteins are filtered out by the small-aperture filter membrane, then, a sealing cover is tightened, an extraction reagent tube housing is inserted into a through hole where the top of a fixing lantern ring and the outer ring of the extraction reagent tube housing are firmly engaged with each other, and a gearbox inside a driving motor is adjusted to a low speed, make driving motor's output order about the rotation action that the drive pivot was done low-speed, the drive pivot rotates the rotary wheel dish with low-speed rotation simultaneously, the round point that the fixed lantern ring was placed on the fixed lantern ring will begin to use the fixed lantern ring rotates as the benchmark, rotate driving motor's slew rate at low-speed after a period and adjust to high-speed, realize super leaving the method through high-low speed complex mode, and adopt low-speed centrifugation, high-speed centrifugation alternate mode easy operation is convenient, the fault-tolerant rate is low, and after filtering and drawing, open sealed lid, will draw the inside big in reagent intraductal storehouse, the aperture filter membrane passes through tweezers and takes out, and the exosome solution after will filtering passes through.

The working principle and the concrete explanation of the invention are as follows:

firstly, PEG reagent or hydrophilic reagent is put into the inner chamber 8 of the extracting reagent tube, after slight shaking, a large-aperture filter membrane and a small-aperture filter membrane are stably placed in the inner wall of the extracting reagent tube inner chamber 8 in sequence, meanwhile, exosome solution is led into the inner chamber 8 of the extracting reagent tube from top to bottom, after exosome solution is led into the inner chamber 8 of the extracting reagent tube, the large-aperture filter membrane arranged in the inner chamber 8 of the extracting reagent tube filters cells, and then the small-aperture filter membrane filters out protein and other impurities, then, after a sealing pad is arranged in the inner wall of the top of the sealing cover 10, the screw threads in the sealing cover 10 are aligned to the screw thread of the outer ring of the threaded sleeve 9, then, the sealing cover 10 is screwed, so that the sealing pad in the sealing cover 10 is tightly contacted with the top edge of the threaded sleeve 9, and the outer ring points of the arranged extracting reagent tube inner chamber 8 are mutually fitted and protruded according to the through hole in the extracting reagent tube shell 5, and the extracting reagent tube shell 5 is inserted into a through hole which is firmly matched with the top of the fixed lantern ring 4 and the outer ring of the extracting reagent tube shell 5.

After the through holes on the fixed lantern ring 4 are all provided with the extracting reagent tube shells 5, the power supply gate of the driving motor 6 in the driving motor cabin body 1 is opened, and the gearbox in the driving motor 6 is adjusted to be at a low speed, so that the output end of the driving motor 6 drives the driving rotating shaft 7 to rotate at a low speed, and simultaneously the driving rotating shaft 7 drives the rotating wheel disc 2 to rotate at a low speed, and a plurality of wedge-shaped through holes are formed in the rotating wheel disc 2, aiming at reducing the overall proportion of the device and realizing light weight, the fixed lantern ring 4 can be better rotated through the supporting columns 3, after the fixed lantern ring 4 is correspondingly connected and driven by the supporting columns 3, the extracting reagent tube shells 5 placed on the fixed lantern ring 4 start to rotate by taking the fixed lantern ring 4 as a reference, the rotating speed of the driving motor 6 is adjusted to be at a high speed after rotating at a period of low speed, and the super separation method is realized in a high-low speed matching mode, and adopt low-speed centrifugation, high-speed centrifugation alternate mode easy operation convenient, the fault-tolerant rate is low to after filtering and drawing, open sealed lid 10, will draw the big, small aperture filter membrane in 8 insides in reagent pipe inner bin to take out through tweezers, and filter the exosome solution after will filtering and filter through 0.1 um's pore membrane, and then realize effective and simple and convenient exosome filtration and draw.

Finally, it should be noted that: the above examples are only intended to illustrate the technical solution of the present invention, but not to limit it; although the present invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some technical features may be equivalently replaced; and such modifications or substitutions do not depart from the spirit and scope of the corresponding technical solutions of the embodiments of the present invention.

Claims

1. The utility model provides an exosome filters extraction element, includes the driving motor storehouse body (1), its characterized in that, the inside of the driving motor storehouse body (1) is the cavity state, and the bottom inner wall of the driving motor storehouse body (1) has driving motor (6) through four screw fixed mounting that are the annular array and arrange, the input and external power supply fixed connection of driving motor (6), the output of driving motor (6) has drive pivot (7) that runs through and extend to driving motor storehouse body (1) top through shaft coupling fixed mounting, the one end welding that drive pivot (7) lie in driving motor storehouse body (1) top has rotating wheel dish (2), one side that rotating wheel dish (2) kept away from drive pivot (7) is all around welded six support columns (3) that are the annular array and arrange, the one end welding that rotating wheel dish (2) was kept away from to support column (3) has fixed lantern ring (4), six placing holes which are arranged in an annular array are formed in the periphery of the top of the fixed lantern ring (4), and an extraction reagent tube shell (5) is clamped inside the placing holes.

2. The exosome filtering and extracting device according to claim 1, characterized in that a groove with a gear-shaped cross section is formed in the extracting reagent tube housing (5), an extracting reagent tube inner bin (8) is sleeved in the groove, and the cross section of the outer ring of the extracting reagent tube inner bin (8) is matched with the cross section of the groove.

3. An exosome filtering and extracting device according to claim 2, characterized in that a threaded sleeve (9) is welded on one side of the extracting reagent tube inner bin (8) far away from the extracting reagent tube outer shell (5), and a plurality of turns of threads are arranged on the outer ring of the threaded sleeve (9).

4. A exosome filtering and extracting device according to claim 3, characterized in that a sealing cover (10) is sleeved on the outer ring of the extracting reagent tube inner bin (8), a groove is formed in the sealing cover (10), and screw threads meshed with the outer ring threads of the threaded sleeve (9) are formed on the peripheral inner wall of the groove.

5. An exosome-filtering-extracting device according to any one of claims 1-4, characterized by further comprising an exosome-extracting method:

firstly, PEG reagent or hydrophilic reagent is put into the inner part of an inner bin (8) of an extraction reagent tube, a large-aperture filter membrane and a small-aperture filter membrane are stably placed in the inner wall of the inner bin (8) of the extraction reagent tube after being slightly shaken, meanwhile, exosome solution is led into the inner part of the inner bin (8) of the extraction reagent tube from top to bottom, after the exosome solution is led into the inner bin (8) of the extraction reagent tube, the large-aperture filter membrane arranged in the inner bin (8) of the extraction reagent tube filters cells, impurities such as protein and the like are filtered by the small-aperture filter membrane, then, after a sealing gasket is installed in the inner wall of the top part of a sealing cover (10), the screw threads in the sealing cover (10) are aligned to the screw thread of the outer ring of a threaded sleeve (9), then, the sealing cover (10) is screwed tightly, so that the sealing gasket in the sealing cover (10) is in close contact with the top edge of the threaded sleeve (9), the protruding points of the outer circle of the set extraction reagent tube inner bin (8) are mutually fitted according to the through hole in the extraction reagent tube shell (5) and then are installed, and the extraction reagent tube shell (5) is inserted into the through hole which is formed by the top of the fixed lantern ring (4) and the outer circle of the extraction reagent tube shell (5) and is mutually firmly fitted;

after the extracting reagent tube shell (5) is installed on the through holes on the fixed lantern ring (4), a power supply gate of a driving motor (6) in the driving motor cabin body (1) is opened, a gearbox in the driving motor (6) is adjusted to be at a low speed, so that the output end of the driving motor (6) drives a driving rotating shaft (7) to rotate at a low speed, the rotating shaft (7) is driven to rotate a rotating wheel disc (2) at a low speed, a plurality of wedge-shaped through holes formed in the rotating wheel disc (2) aim at reducing the overall proportion of the device and realizing light weight, the fixed lantern ring (4) can be rotated better through a supporting column (3), after the fixed lantern ring (4) is correspondingly connected and driven by the supporting column (3), the extracting reagent tube shell (5) placed on the fixed lantern ring (4) starts to rotate by taking a round point of the fixed lantern ring (4) as a reference, rotate the slew rate of driving motor (6) after a period at low-speed and adjust to high-speed, realize super-separation method through high-low speed complex mode, and adopt low-speed centrifugation, high-speed centrifugation alternate mode easy operation is convenient, the fault-tolerant rate is low, and after filtering and drawing, open sealed lid (10), will draw the inside big, small aperture filter membrane in reagent intraductal storehouse (8) and take out through tweezers, and filter the exosome solution after will filtering and filter through 0.1 um's pore membrane.

6. The exosome filtering and extracting device according to claim 1, characterized in that a plurality of groups of through holes arranged in an annular array are arranged at the top of the rotating wheel disc (2), and the cross sections of the through holes are all wedge-shaped.