CN116337826A

CN116337826A - Biological material separator

Info

Publication number: CN116337826A
Application number: CN202111604470.5A
Authority: CN
Inventors: 吴钍荣; 徐震宇; 林晓东; 陈瑛娜
Original assignee: Guangzhou Zhaorui Medical Biotechnology Co ltd
Current assignee: Guangzhou Zhaorui Medical Biotechnology Co ltd
Priority date: 2021-12-24
Filing date: 2021-12-24
Publication date: 2023-06-27
Also published as: WO2023115734A1

Abstract

The application belongs to the technical field of biological material separation. The utility model specifically discloses a biological material separator, including the bottom plate, the bottom plate includes at least one extraction mechanism, extraction mechanism includes extraction element, extraction element includes first cavity, connecting portion and second cavity, first cavity pass through connecting portion connect in the second cavity, the internal diameter of connecting portion is less than first cavity with the second cavity. The application has at least one of the following beneficial effects: the biological material separation device provided by the application is characterized in that the connecting part is arranged between the two chambers, so that the separation interface is reduced, and the target object is separated more conveniently.

Description

Biological material separator

Technical Field

The present application relates to the field of biological material separation technology, and more particularly, to a biological material separation device.

Background

In the fields of chemistry, biology, medicine and the like, high-efficiency extraction and separation of trace and trace solid phases in a liquid phase matrix are frequently faced. In the biomedical field, when separating plasma from whole blood, the contact surface between the plasma and blood cells is large, so that the extraction position is far away from the interface between the plasma and the blood cells as far as possible when extracting the plasma, and the plasma is prevented from being extracted, more plasma will remain, and part of plasma is wasted. In addition, since the interface between blood cells and plasma is large, the interface is unstable, and even a slight shaking during the aspiration of plasma, the blood cells and plasma are mixed with each other, and further, the plasma is lost.

Disclosure of Invention

In view of this, in order to reduce the loss of the target as much as possible, the present application provides a biological material separation apparatus capable of effectively reducing the separation interface, thereby reducing the loss of the target due to the separation interface.

The application is realized by the following scheme:

the application provides a biological material separation device, biological material separation device includes the bottom plate, the bottom plate includes at least one extraction mechanism, extraction mechanism includes extraction element, extraction element includes first cavity, connecting portion and second cavity, first cavity pass through connecting portion connect in the second cavity, connecting portion's internal diameter is less than first cavity with the second cavity.

In the application, through the design of the connecting part (namely necking) between the first chamber and the second chamber, the connecting interface after separation is reduced, the mixing degree of the target object and the separation substance is reduced, and more target objects can be taken out.

In a specific embodiment of the present application, the first chamber transitions to the connection portion where a step is provided, or the first chamber transitions to the connection portion through a slope having a certain gradient, so that the impurity entering the second chamber can be prevented from reentering the first chamber.

In a specific embodiment of the present application, the extraction mechanism further includes a sample introduction unit, the sample introduction unit is used for inputting and storing a sample to be separated, the extraction unit is used for extracting a target object in the sample to be separated, the sample introduction unit is disposed at an upstream of the extraction unit, and the sample introduction unit is connected to the first chamber through a first channel.

In the application, the sample to be separated can be firstly added into the sample injection unit for storage, and then subsequent treatment is carried out.

In one specific embodiment of the present application, the number of the extraction mechanisms 10 is 2 or more, and the sample injection units of the different extraction mechanisms are connected or not connected.

In one specific embodiment of the present application, the sample injection units of the different extraction mechanisms are connected through a first capillary tube.

The utility model discloses a can set up a plurality of extraction mechanism in this application, can pass through capillary interconnect between the sampling unit of different extraction mechanism, so, when the application of sample, only need carry out the application of sample to a sampling unit, then through capillary action, the sample can flow in other sampling units. If the sample injection units of the plurality of different extraction mechanisms are not connected, the different extraction mechanisms can be used for separating different samples at the same time, and the same sample can be separated by using the different extraction mechanisms at the same time, so that the processing capacity of each time can be increased, the processing efficiency is improved, and the time is saved.

In one embodiment of the present application, the biomaterial separation device further includes a rotation center disposed at a central portion of the biomaterial separation device.

In a specific embodiment of the present application, the number of the extraction mechanisms is 1, the sample introduction unit and the extraction unit are disposed on two opposite sides of the rotation center, and the first channel is in an arc shape surrounding the rotation center.

In this application, the biomaterial separation device may be used for centrifugation. The rotation center can be used as the rotation center in centrifugal operation, and the sample feeding unit and the extraction unit are arranged on two opposite sides of the rotation center, so that the whole biological material separation device tends to be balanced.

In a specific embodiment of the present application, the number of the extraction mechanisms is 3 or more, and the extraction mechanisms are uniformly distributed around the rotation center.

In a specific embodiment of the present application, the number of the extraction mechanisms is more than 3, the extraction mechanisms are uniformly distributed around the rotation center, and the centers of the sample injection units of the extraction mechanisms are on the same circle.

In a specific embodiment of the present application, the number of the extraction mechanisms is more than 3, the extraction mechanisms are uniformly distributed around the rotation center, and the central connection lines of the sample injection units of the extraction mechanisms are not completely on the same circle. For example, the extraction mechanisms are 2n (n is a positive number and n is greater than 1), the center lines of the sample introduction units of the 1 st, 3, 5 th..2n-1 th extraction mechanism are on a first circle, and the center lines of the sample introduction units of the 2 nd, 4 th, 6 th..2n extraction mechanisms are on a second circle, the first circle being different from the second circle. For another example, the extraction mechanisms are 3n (n is a positive number, and n is greater than 1), and the central connecting line of the sample injection units of the 1 st, 4 th and 7 th extraction mechanisms and the 3n-2 th extraction mechanisms are on the first circle; the center connecting line of the sample injection units of the 2 nd, 5 th and 8 th extraction mechanisms and the 3n-1 th extraction mechanisms are arranged on a second circle; the centers of the sample introduction units of the 3 rd, 6 th and 9 th extraction mechanisms are connected on a third circle, and the first circle, the second circle and the third circle can be the same or different from each other.

In a specific embodiment of the present application, the number of the extraction mechanisms is more than 3, the extraction mechanisms are sequentially arranged around the rotation center, and the sample introduction units of the extraction mechanisms are sequentially away from the rotation center.

In one embodiment of the present application, the biomaterial separation device further comprises a third chamber connected to the extracting unit through a second channel.

In one embodiment of the present application, the biomaterial separation device further comprises a cover plate, wherein the cover plate is matched with the base plate, i.e. when the base plate has no rotation center, the cover plate has no rotation center. When the rotation center is arranged on the bottom plate, the rotation center is also arranged on the corresponding cover plate. In this application, apron cooperatees with the bottom plate, can prevent the liquid spill in the bottom plate.

In a specific embodiment of the present application, a first through hole is provided on the cover plate of the sample introduction unit. In the application, the sample to be separated can be added to the sample introduction unit through the first through hole.

In one embodiment of the present application, the sample introduction unit includes a fourth chamber connected to the first chamber through a first channel.

In a specific embodiment of the present application, the shape of the fourth chamber may be a "triangle" shape, a "circle" shape, a "quadrilateral" shape, a "sector" shape, an "oval" shape, a "diamond" shape, or the like, but is not limited to the above-mentioned shapes.

In one embodiment of the present application, the first through hole is on a side of the fourth chamber near the center of rotation.

In one embodiment of the present application, the first channel is connected to a side of the fourth chamber remote from the rotation center.

In a specific embodiment of the present application, the sample injection unit may further include a first sample injection hole, where the first sample injection hole is connected to the fourth chamber, and is used for inputting a sample to be separated into the fourth chamber, and the fourth chamber is used for storing the sample to be separated.

In one specific embodiment of the present application, the first sample application hole is disposed on a side of the fourth chamber near the rotation center.

In one embodiment of the present application, the first through hole is opposite to the first loading hole.

In one embodiment of the present application, the biomaterial separation device further comprises a first filter unit connected to the connection part of the extracting mechanism through a second capillary tube.

In one embodiment of the present application, the first filter unit comprises a first membrane for intercepting a portion of the substance flowing from the second capillary tube.

In one embodiment of the present application, the first filter unit includes a first groove, and the first membrane is embedded in the first groove.

In one embodiment of the present application, the first filter unit further comprises a fifth chamber connected to the first groove through a third channel 1135.

In a specific embodiment of the present application, a connection point where the first groove transitions to the third channel is provided with a step.

In one embodiment of the present application, the first groove transitions to the third channel through a ramp having a slope.

In one specific embodiment of the present application, a sixth chamber is disposed on a side of the first groove near the rotation center, and the second capillary is connected to the first groove through the sixth chamber.

In one embodiment of the present application, a seventh chamber is provided on a side of the first groove remote from the rotation center, and the third channel is connected to the first groove through the seventh chamber.

In a specific embodiment of the present application, the depth of the sixth chamber and the seventh chamber is smaller than the depth of the first groove.

In one embodiment of the present application, the second capillary tube is in the shape of an inverted "V" in front view, the tip of the "V" being positioned closer to the center of rotation relative to the first chamber upstream thereof.

In one embodiment of the present application, the first filter unit further comprises a seventeenth chamber connected to a side of the fifth chamber near the rotation center through a fourth channel.

In a specific embodiment of the present application, the bottom of the seventeenth chamber is provided with a first supporting table and a first boss, which can support the first container placed in the seventeenth chamber.

In one embodiment of the present application, a second concave wall is provided on the cavity wall of the seventeenth cavity toward the direction of the cavity for fixing the first container placed in the seventeenth cavity.

In one specific embodiment of the present application, a first flow channel is provided at the top of the first boss, for draining the liquid in the first container.

In a specific embodiment of the present application, the top of the first boss is equal to the table top of the first supporting table with respect to the bottom plate surface.

In a specific embodiment of the present application, the top of the first boss is higher than the table top of the first supporting table based on the bottom plate surface.

In the present application, the first support stand and the first boss are used for supporting the first container. When the first container is pressed downwards under the force, the first container takes the first supporting table as a fulcrum to perform leverage, and the first boss can crush the first container, so that the reagent in the first container enters the seventeenth chamber. The first flow passage facilitates drainage of the reagent in the first container into the seventeenth chamber.

In a specific embodiment of the present application, the first container may be a container that is breakable by applying a certain pressure.

In one embodiment of the present application, the first container includes a second cavity and a first release hole disposed at a bottom surface of the second cavity. A first sealing film is arranged on one side of the first release hole.

In a specific embodiment of the present application, the first container may be integrally formed, or a top cover may be disposed on the second cavity, so that the second cavity and the top cover cooperate to form a container for sealing the reagent disposed in the second cavity.

In a specific embodiment of the present application, the first release hole is opposite to the first boss when the first container is placed in the seventeenth chamber.

In a specific embodiment of the present application, the side of the second cavity is further provided with a first concave wall. The first concave wall is matched with the second concave wall and used for fixing the first container, so that the first container is prevented from generating large rotary displacement on the bottom plate, the first release hole is always located on the first boss, and the first container can be effectively broken when being extruded.

In a specific embodiment of the present application, the first recess wall and the second recess wall may be interchangeably disposed, i.e. in that the first recess wall is disposed in the seventeenth chamber and the second recess wall is disposed in the second chamber.

In this application, the first recess wall and the second recess wall cooperate to limit rotational displacement of the first container on the base plate. Thus, other ways of limiting the rotational displacement of the first container on the base plate are also suitable for the present application. For example, the concave wall is designed as a convex wall or designed as a snap-in manner, etc.

In a specific embodiment of the present application, if the first sealing film is disposed on the inner side of the second cavity, and the first release hole is deeper (i.e., the bottom surface of the second cavity is thicker), the top of the first boss extends into the first release hole, but the top of the first boss does not contact the first sealing film. Alternatively, the top of the first boss is slightly contacted with the first sealing film, but the first sealing film has a certain supporting function, and the first sealing film is not broken under the condition of no external pressure.

In a specific embodiment of the present application, if the first sealing film is disposed on the outer side of the second cavity, that is, the first sealing film is disposed on a side of the first release hole near the first boss, the top of the first boss is slightly higher than the table top of the first supporting table, but the top of the first boss is not contacted with the sealing film. Alternatively, the top of the first boss 7 is in slight contact with the first sealing film, but the first sealing film has a certain supporting effect, and the first sealing film is not crushed in the absence of externally applied pressure.

In one embodiment of the present application, the biomaterial separation device further comprises a second filter unit connected to the first filter unit through a fifth channel. Preferably, the first filter unit is connected with the fifth channel through a third capillary tube.

In a specific embodiment of the present application, in a front view, the third capillary tube has an inverted "V" shape, and a tip portion of the "V" shape is positioned closer to a center of rotation with respect to a fifth chamber upstream thereof.

In one embodiment of the present application, the second filter unit comprises a second membrane for intercepting a portion of the substance flowing from the fifth channel.

In one embodiment of the present application, the second filter unit includes a second groove, and the second membrane is embedded in the second groove.

In one embodiment of the present application, the second filter unit further comprises an eighth chamber connected to the second recess through a sixth channel.

In a specific embodiment of the present application, a step is provided at the connection of the sixth channel of the second groove.

In one embodiment of the present application, the second groove transitions to the sixth channel through a ramp having a slope.

In one embodiment of the present application, the sixth channel is curved. The curved arrangement on the one hand contributes to an increased flow path length; on the other hand, the degree of mixing of the mixture in the sixth passage can be reduced.

In a specific embodiment of the present application, the sixth channel includes a first branch channel, a second branch channel and a third branch channel, the first branch channel and the third branch channel are straight channels, the second branch channel is a curved channel, the first branch channel is connected to the second groove, the third branch channel is connected to the eighth chamber, and the number of bends of the middle second branch channel is greater than or equal to 1.

In a specific embodiment of the present application, a connection point where the second groove transitions to the first branch passage is provided with a step.

In a specific embodiment of the present application, the second groove transitions into the first branch channel through a slope having a slope.

In a specific embodiment of the present application, the second filter unit further comprises a ninth chamber. The ninth chamber is connected to the first branch passage through a seventh passage.

In a specific embodiment of the present application, the seventh channel is inclined in an upstream direction of the sixth channel, and the inclination angle may be any angle between greater than 0 ° and less than 90 °. In one embodiment, the angle of inclination is 30-90. For example, the angle of inclination is 30 °,35 °,40 °,45 °,50 °,55 °,60 °,65 °,70 °,75 °,80 °, 85 °, or the like.

In a specific embodiment of the present application, a nineteenth chamber is provided on a side of the second groove remote from the rotation center, and the first branch passage is connected to the second groove through the nineteenth chamber.

In one specific embodiment of the present application, the second filter unit further includes a twentieth chamber disposed at a side of the second groove near the rotation center and connected to the second groove through an eleventh passage.

In a specific embodiment of the present application, an eighteenth chamber is disposed between the second recess and the eleventh channel, and the fourth channel is connected to the second recess through the eighteenth chamber.

In a specific embodiment of the present application, the twentieth chamber may be the same as the seventeenth chamber. I.e. the bottom of the twentieth chamber is provided with a second support table and a second boss for supporting a second container placed in the twentieth chamber.

In a specific embodiment of the present application, a third concave wall is provided on the cavity wall of the twentieth chamber toward the direction of the cavity for fixing the second container placed in the twentieth chamber.

In one specific embodiment of the present application, a second flow channel is provided at the top of the second boss, for draining the liquid in the second container.

In a specific embodiment of the present application, the top of the second boss is equal to the table top of the second supporting table or the top of the second boss is higher than the table top of the second supporting table based on the bottom plate surface.

In a specific embodiment of the present application, the second supporting platform and the second boss are used for supporting the second container, when the second container is pressed downwards by force, the second container takes the second supporting platform as a pivot, leverage occurs, and the second boss will crush the second container, so that the reagent in the second container enters the twentieth chamber. The second flow passage facilitates drainage of reagent in the second container into the twentieth chamber.

In a specific embodiment of the present application, the second container is the same as the first container, that is, the second container includes a second cavity and a second release hole disposed on a bottom surface of the second cavity, and a second sealing film is disposed on one side of the second release hole. The second release aperture is aligned with the second boss when the second container is placed in the twentieth chamber. The second container may further comprise a fourth recess wall, the fourth recess wall cooperating with the third recess wall.

In one embodiment of the present application, the twentieth chamber and the seventeenth chamber may be used for directly placing a reagent, or may be used for placing a container for holding a reagent.

In one embodiment of the present application, the biomaterial separation device further comprises a third filter unit comprising a third groove connected to the seventh channel. In this application, the reagent for detection may be placed in the third recess, which serves as a reaction detection chamber.

In one embodiment of the present application, the third filter unit further comprises an eleventh chamber connected to the third recess by a passage.

In one embodiment of the present application, a third film is embedded in the third groove. The substance to be detected passes through the third membrane and then enters the eleventh chamber for detection.

In one embodiment of the present application, the third groove is connected to the seventh passage through a twelfth chamber on a side near the rotation center.

In one embodiment of the present application, the third filter unit further includes a fourteenth chamber connected to a side of the twelfth chamber near the rotation center through an eighth passage.

In a specific embodiment of the present application, the third filter unit further comprises a thirteenth chamber connected to a side of the third recess remote from the rotation center through a ninth channel. The eleventh chamber is connected to the ninth channel through a tenth channel.

In a specific embodiment of the present application, the ninth channel may be a straight channel or may include a curved channel, where the number of bends in the curved channel of the ninth channel is smaller than the number of bends in the sixth channel.

In a specific embodiment of the present application, the ninth channel is a straight channel, the ninth channel includes a fourth branch channel and a fifth branch channel, in a front view, the fourth branch channel and the fifth branch channel are not on the same straight line, a certain included angle is formed between the fourth branch channel and the fifth branch channel, the fourth branch channel is connected to the thirteenth chamber, the fifth branch channel is connected to the third groove, and the tenth channel is connected to the fifth branch channel.

In one embodiment of the present application, a fifteenth chamber is provided on a side of the third groove remote from the rotation center, and the fifth branch passage is connected to the third groove through the fifteenth chamber.

In a specific embodiment of the present application, the inlet direction of the tenth channel is inclined to the upstream direction of the ninth channel, and the inclination angle may be any angle between greater than 0 ° and less than 90 °. In one embodiment, the angle of inclination is 30-90. For example, the angle of inclination is 30 °,35 °,40 °,45 °,50 °,55 °,60 °,65 °,70 °,75 °,80 °, 85 °, or the like.

In one embodiment of the present application, the third filter unit further comprises a sixteenth chamber connected to a side of the eleventh chamber near the rotation center through a passage.

In a specific embodiment of the present application, the sixth chamber, seventh chamber, eleventh chamber, twelfth chamber, fifteenth chamber, seventeenth chamber, eighteenth chamber and nineteenth chamber may be used as a container directly for holding a reagent, or as a reaction chamber or the like.

In a specific embodiment of the present application, the design of the sixth chamber, seventh chamber, eleventh chamber, twelfth chamber, fifteenth chamber, eighteenth chamber and nineteenth chamber may be the same as the design of the chamber of the seventeenth chamber, for placing a container for holding a reagent, such as the first container described above, if necessary.

In one embodiment of the present application, the first film, the second film and the third film may be adhered to the hard substrate and then embedded in the groove together with the hard substrate, thereby overcoming the problem of easy deformation of the thin film. The operation is simple, and the membrane can be effectively and firmly placed in the flow path.

Taking the first film as an example, in a specific embodiment of the present application, a window is formed in the middle of the hard substrate, and the first film is adhered to one side of the window. Preferably, when the hard substrate is placed in the first groove, the side of the window to which the first film is attached is close to the rotation center.

In one embodiment of the present application, a first film is adhered to one side of the window for sealing the window. The attaching here may be that the first film is bonded on the first substrate. The bonding mode comprises adhesive bonding, thermocompression bonding or ultrasonic welding. Preferably, the first film is thermocompression bonded on the first substrate.

In a specific embodiment of the present application, the depth of the first groove is greater than that of the adjacent sixth chamber and seventh chamber, so that the hard substrate can be ensured to be more stable when placed in the first groove (16 filter membrane 1 placement groove).

In one embodiment of the present application, the first film is placed in the first recess perpendicular to the floor surface to ensure effective utilization of the film.

Taking the second film as an example, in a specific embodiment of the present application, the hard substrate includes a first cavity and a sleeve, where the sleeve is detachably disposed in the first cavity and is used in cooperation with the first cavity. The bottom of the first cavity is provided with a first outflow hole. The second membrane is arranged at one end of the sleeve. When in use, one end of the sleeve provided with the second film is arranged at the bottom of the first cavity, and then the whole hard substrate is arranged in the second groove. When placed, the second film is distanced from the rotation center.

In one specific embodiment of the present application, the bottom of the first cavity is provided with a plurality of (2 or more) first outflow holes, so that the liquid in the sleeve passes through the second membrane and then rapidly enters the downstream channel.

In a specific embodiment of the present application, the bottom of the first cavity is provided with a fourth groove, and the first outflow hole is all arranged in the fourth groove, so that the fourth groove can be used as a channel for liquid circulation, and the liquid to be treated can be further accelerated to pass through the second membrane and enter the downstream channel. When the first outflow holes are multiple, a film supporting table is arranged between the adjacent first outflow holes and used for supporting the second film to prevent the second film from being broken due to overlarge stress. The number of the membrane support tables may be set as needed, for example, when the number of the first outflow holes is large, the membrane support tables may be set in a gap.

In a specific embodiment of the present application, the second film and the sleeve may be joined by bonding, for example, adhesive bonding, thermocompression bonding, ultrasonic welding, or the like. Preferably, the second film is thermocompression bonded to one end of the sleeve. In this application, through the cooperation of sleeve and first cavity use, make the leakproofness of stereoplasm base plate stronger.

In a specific embodiment of the present application, a first top pinhole is disposed on the cover plate, and the first top pinhole is communicated with the seventeenth chamber.

In a specific embodiment of the present application, the first top pin hole is aligned with the first boss, so that when pressure is applied to the first container disposed in the seventeenth chamber through the first top pin hole, the first boss is facilitated to burst the first container.

In a specific embodiment of the present application, a second top pinhole is disposed on the cover plate, and the second top pinhole is communicated with the twentieth chamber.

In a specific embodiment of the present application, a fourth through hole is disposed on the cover plate, and the second through hole is communicated with the sixteenth chamber.

In a specific embodiment of the present application, a third through hole is disposed on the cover plate, and the third through hole is communicated with the fourteenth chamber.

In the application, the cover plate and the bottom plate can be bonded by double-sided adhesive or hot-press bonding.

In one embodiment of the present application, the cover plate and the base plate may be made of the same material. For example, the cover plate and the bottom plate are made of polymethyl methacrylate (PMMA), polycarbonate (PC) or polyvinyl chloride (PVC) and other polymer materials.

In one embodiment of the present application, the cover plate and the base plate may be made of different materials. For example, PMMA is selected as the cover plate, and PC is selected as the bottom plate.

In one embodiment of the present application, the cover plate and the base plate are made of Polycarbonate (PC).

In one embodiment of the present application, the inner surface of the channel is provided with a hydrophilic material or a hydrophobic material. The affinity of the channels for the circulating liquid is made different by subjecting the channels to hydrophilic or hydrophobic treatments.

In a specific embodiment of the present application, the bottom plate is provided with a balancing chamber for keeping the entire bottom plate in a certain equilibrium state.

In one embodiment of the present application, the channels, capillaries, chambers, grooves, etc. are formed all over the bottom plate by recessing down on the same surface of the bottom plate.

In a specific embodiment of the present application, the shape of the entire bottom plate may be related to the type of centrifuge in the prior art, for example, designed into a shape of an 8-well plate, a 96-well plate or a circle, etc., and the inner surfaces of the chamber and the groove may be designed into an arc surface, a rectangular surface, a square surface, a triangular surface or a diamond surface, etc., as required. The channels may be designed as straight channels, curved channels, channels with bends, etc.

In this application, the first membrane 1132, the second membrane 11410, the third membrane 2012, the adsorption membrane 11311, the first sealing membrane 1203, the second sealing membrane, and the reagents in the first and second containers may be replaced according to the separated or detected substances. The same substance may be repeatedly separated or detected by the whole biomaterial separation device by replacing the adsorbing film 11311, the first sealing film 1203, the second sealing film, and the reagents in the first and second containers, or the whole biomaterial separation device may be repeatedly used after directly replacing the hard substrate, the first and second containers.

The biological material separation device provided in the application can be applied to the separation of trace liquid in various fields.

In one embodiment of the present application, the biomaterial separation device is used to separate exosomes.

In one embodiment of the present application, the biomaterial separation device is used to separate and detect exosomes.

The biological material separation device provided by the application has at least one of the following beneficial effects:

the biological material separation device provided by the application is characterized in that the connecting part is arranged between the two chambers, so that the separation interface is reduced, and the target object is separated more conveniently.

Drawings

Fig. 1 is a front view of a biomaterial separating device with only 1 extracting mechanism provided in the present embodiment.

Fig. 2 is a front view of the biomaterial separating device having 2 extracting mechanisms provided in the present embodiment.

Fig. 3 is a front view of the biomaterial separating device having a plurality of extracting mechanisms provided in the present embodiment.

Fig. 4 is a front view of the biomaterial separation device including the rotation center provided in the embodiment.

Fig. 5 is a front view of the biomaterial separation device including the rotation center provided in the embodiment.

Fig. 6 is a front view of the biomaterial separation device including the plurality of extracting mechanisms provided in the present embodiment.

Fig. 7 is a front view of the biomaterial separation device including a plurality of extracting mechanisms provided in the present embodiment.

Fig. 8 is a front view of the biomaterial separation device including the first filter unit provided in the embodiment.

Fig. 9 is a front view of the biomaterial separation device including the first filter unit provided in the embodiment.

Fig. 10 is a front view of the biomaterial separation device including the second filter unit provided in the embodiment.

Fig. 11 is a front view of the biomaterial separation device including the third filter unit provided in the embodiment.

Fig. 12 is an enlarged view of G provided in the present embodiment.

Fig. 13 is an exploded view of the first container provided in the present embodiment.

Fig. 14 is an exploded view of the hard substrate provided in the present embodiment.

Fig. 15 is an exploded view of the hard substrate provided in the present embodiment.

Fig. 16 is a perspective view of the cover plate provided in the present embodiment.

Fig. 17 is a statistical chart of the particle diameters of the extracted exosomes provided in this example.

Fig. 18 is an electron microscope image of the extracted exosomes provided in this example.

Fig. 19 is an electron microscope image of the extracted exosomes provided in this example.

Detailed Description

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs.

The following description of the embodiments of the present application will clearly and fully describe the technical solutions of the present application, and it is apparent that the described embodiments are only some embodiments of the present application, but not all embodiments. All other embodiments, which can be made by one of ordinary skill in the art based on the embodiments herein without making any inventive effort, are intended to be within the scope of the present application. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.

The capillary tube is a thin tube, the inner diameter of the capillary tube is smaller, when liquid is in the tube, the liquid can rise by overcoming the gravity due to the difference of cohesion and adhesion, a capillary phenomenon is generated, and the inner diameter of a channel or a runner is larger, so that no obvious capillary phenomenon exists.

The target in the application refers to a substance or a solution or suspension thereof to be separated, the impurity refers to a part of the substance or the solution or the suspension except the target, or the whole substance or the solution or the suspension, but a layering interface can be formed between the target and the impurity through operation means such as standing or centrifugation.

The biological material separation device can be used for separating substances and detecting the substances while separating the substances.

The biological material separation device in the application is not only suitable for various liquid samples such as whole blood, plasma, serum, saliva, urine, milk, cerebrospinal fluid, semen, pleural effusion and the like in the biological field, but also suitable for the separation and detection of trace components in other fields.

In the present application, the processing modes of the biomaterial separation device include, but are not limited to, CNC, laser engraving, soft lithography, 3D printing, injection molding, and the like.

Example apparatus for separation

As shown in fig. 1, in one embodiment of the present application, a biological material separation device is provided that includes a base plate. The base plate comprises an extraction mechanism 10, the extraction mechanism comprises an extraction unit 112, the extraction unit 112 comprises a first chamber 1121, a connecting portion 1122 and a second chamber 1123, the first chamber 1121 is connected to the second chamber 1123 through the connecting portion 1122, and the inner diameter of the connecting portion 1122 is smaller than the first chamber 1121 and the second chamber 1122. In the biomaterial separation device provided in this embodiment, the first chamber is used for storing the target object, the second chamber is used for storing the impurity, and the mixing interface of the target object and the impurity is located at the connecting portion. The diameter of the connecting part is obviously smaller than that of the first chamber and the second chamber, so that the mixing interface of the target object and the impurities is obviously reduced, and the extraction of the target object is facilitated.

In this embodiment, a step is provided where the first chamber 1121 transitions to the connection 1122, or the first chamber 1121 transitions to the connection 1122 through a slope having a certain gradient to prevent impurities entering the second chamber 1123 from re-entering the first chamber 1121.

As shown in fig. 1, the extraction mechanism 10 in this embodiment further includes a sample introduction unit, where the sample introduction unit includes a fourth chamber 1111, and the fourth chamber 1111 is connected to the first chamber 1121 through a first channel 1124. The sample feeding unit is used for inputting and storing samples to be separated.

The biomaterial separation device in this embodiment may be used for separation of liquid, for example, when separating whole blood, a whole blood sample is added to the fourth chamber 1111, and the biomaterial separation device is placed on a centrifuge for centrifugation. During centrifugation, the rotational speed is controlled so that the centrifuged blood cells are collected in the second chamber 1123, and the plasma is collected in the first chamber 1121, and the connection 1122 shows the interface between the blood cells and the plasma. After centrifugation, the plasma in the first chamber 1121 can be conveniently removed for later use. The rotating speed range can be 2500-5500 rpm, and the time T1 can be controlled between 120s and 480s.

As shown in fig. 2 and 3, the biomaterial separation device in one embodiment of the application may include a plurality of extracting mechanisms 10, and the extracting mechanisms 10 may or may not be connected to each other. The fourth chamber 1111 between the two extraction mechanisms 10 in fig. 2 is connected by the first capillary tube 1113, and in fig. 3, the fourth chamber 1111 between part of the two extraction mechanisms is connected by the first capillary tube 1113, and the fourth chamber 1111 between part of the two extraction mechanisms is not connected.

In this application, if there is no connection between the plurality of extraction mechanisms, different extraction mechanisms 10 may be used to separate different samples to be separated, such as whole blood samples (blood) derived from different patients, and the like. If the sample injection units among the plurality of extraction mechanisms are connected through capillary tubes, the extraction mechanisms which are connected with each other can be used for separating the same sample to be separated, so that the once treatment capacity is improved, and the treatment efficiency is improved.

In this application, the number of the extraction mechanisms 10 may be set according to actual needs. For example 1, 2, 4, 6, 8, 12, 24, 48, 96 or 384, etc. For example, the bottom plate is designed into the shape of a 96-well plate, and then the bottom plate can be directly placed on a 96-well plate centrifuge for centrifugation.

As shown in fig. 4, in another embodiment of the present application, the biological material separation device further includes a rotation center 101, the rotation center 101 is disposed at a center position of the biological material separation device, the sample introduction unit and the extraction unit 112 are disposed at opposite sides of the rotation center 101, and the first channel 1124 has an arc shape surrounding the rotation center 101.

In this application, the rotation center can be regarded as the rotation center when centrifugal operation, with sampling unit and extraction element setting in the both sides that rotation center is relative, can make whole biological material separator tend to balance. In order to balance the whole biological material separation device during centrifugation, a balancing chamber 102 is also provided on the bottom plate.

As shown in fig. 5, 6 and 7, the number of extraction mechanisms 10 is 2 or more, and the extraction mechanisms 10 are uniformly distributed around the rotation center 101.

As shown in fig. 6, the number of the extraction mechanisms 10 is 3, and the centers of the fourth chambers 1111 of the 3 extraction mechanisms are on the same circle a.

As shown in fig. 7, the extraction mechanisms 10 are 6, which are distributed uniformly around the rotation center 101 in turn, wherein the centers of the sample introduction units of the 1 st, 3 th and 5 th extraction mechanisms are on the same circle B, and the centers of the sample introduction units of the 2 nd, 4 th and 6 th extraction mechanisms are on another circle C.

In fig. 5, the sample injection units of 2 extraction mechanisms are connected by a first capillary 1113. The sample introduction unit further includes a first sample introduction hole 1112, where the first sample introduction hole 1112 is disposed on a side of the fourth chamber 1111 near the rotation center 101, for inputting a sample to be separated into the fourth chamber 1111, and the fourth chamber 1111 is used for storing the sample to be separated. The first channel 1124 is connected to a side of the fourth chamber 1111 remote from the center of rotation 101. Thus, upon centrifugation, the sample to be separated which enters the fourth chamber 1111 enters the first passage 1124 along the side wall of the fourth chamber 1111 under the action of centrifugal force to prevent the sputtering of the sample to be separated.

For example, when separating a whole blood sample, the whole blood sample to be separated is introduced into the fourth chamber 1111 on the right side through the first introduction hole 1112, and the whole blood sample introduced into the fourth chamber 1111 is introduced into the fourth chamber 1111 on the left side through the first capillary tube 1113 by capillary action. When the biomaterial separation device is placed on a centrifuge for centrifugation, plasma is introduced into the first chamber 1121 and blood cells are introduced into the second chamber 1123, respectively. In this way, more samples can be processed at a time to increase the processing efficiency.

In the present embodiment, the shape of the fourth chamber 1111 may be a "triangle" shape, a "circle" shape, a "quadrangle" shape, a "fan" shape, an "ellipse" shape, a "diamond" shape, or the like, but is not limited to the above-described shape. Preferably, the shape of the fourth chamber 1111 may be a "triangle" shape, a "quadrangle" shape, or a "diamond" shape, etc. with "corners", and the first wells 1112 are disposed at positions adjacent to the "corners" of the "triangle" shape, the "quadrangle" shape, or the "diamond" shape.

As shown in fig. 8, in one embodiment of the present application, the biomaterial separation device includes 2 extracting mechanisms, and the 2 extracting mechanisms are symmetrical about the rotation center 101. One of the separation mechanisms will be described below as an example.

The biomaterial separation device in the embodiment further includes a first filtering unit 113 for filtering, and the first filtering unit 113 is connected to the connecting part 1122 through a second capillary tube 1131. The first filter unit 113 includes a first membrane 1132 and a fifth chamber 1133, and a sample to be separated may be stored in the fifth chamber 1133 after passing through the first membrane 1132.

The second capillary 1131 in this embodiment is in the shape of an inverted "V" whose tip is positioned closer to the rotation center 101 with respect to the first chamber 1121 upstream thereof.

In fig. 8, a first membrane 1132 (not shown) may be embedded in the first groove 1134, and the fifth chamber 1133 is connected to the first groove 1134 through a third channel 1135.

In this embodiment, the junction where the first groove 1134 transitions to the third channel 1135 is provided with a step for preventing the liquid entering the fifth chamber 1133 from flowing back into the first groove 1134. The steps in this embodiment may be replaced with slopes having a slope.

In this embodiment, a sixth chamber 1136 is disposed between the first groove 1134 and the second capillary 1131 and adjacent to the first groove 1134 for buffering the liquid flowing in from the second capillary 1131. A seventh chamber 1137 is provided between the first groove 1134 and the third channel 1135 and adjacent to the first groove 1134 for collecting liquid passing through the first membrane 1132.

In fig. 8, the biomaterial separation device in the embodiment further includes a third chamber 1125, and the third chamber 1125 is connected to the first chamber 1121 through a second channel. The design of the third chamber 1124 in this application may enable dosing of the first chamber 1121. For example, when whole blood is separated, excess whole blood is added to the fourth chamber 1111 and after centrifugation, the excess blood will enter the third chamber 1125.

When proteins having a molecular weight of 100 ten thousand or less are separated by the biomaterial separation device in fig. 8, the first membrane 1132 may be an ultrafiltration membrane of 100 ten thousand Da. The specific separation process is as follows:

the whole blood sample to be separated is added to the fourth chamber 1111:

s1: the rotational speed of the biomaterial separation device is controlled, the first chamber 1121 is filled with plasma after the completion of the rotation, blood cells enter the second chamber 1123, and the third chamber 1125 accommodates excessive blood. The rotating speed range can be 2500-5500 rpm, and the time T1 can be 120-480 s.

S2: stopping standing for a period of time. The second capillary 1131 fills with plasma due to capillary effect. The standing time T2 may be 5s to 50s.

S3: the biomaterial separation device is controlled to perform high-speed rotation, at this time, the second capillary tube 1131, which has been filled, has a siphon effect, and the plasma in the first chamber 1121 flows out through the second capillary tube 1131 into the sixth chamber 1136, and then is stored in the fifth chamber 1133 after passing through the first die 1132 disposed in the first groove 1134. The rotation speed range can be 2000-4500 revolutions/min, and the time T3 can be 15-120 s.

As shown in fig. 9, in another embodiment of the present application, compared to the biological material separation device in fig. 8, in the biological material separation device in fig. 9, the first filtering unit further includes a seventeenth chamber 1138, a fifth groove 11312 and a tenth chamber 11310, the seventeenth chamber 1138 is connected to a side of the fifth chamber 1133, which is close to the rotation center 101, through a fourth channel 1139, the fifth groove 11312 is disposed at a side of the fifth chamber 1133, which is far from the rotation center 101, and is connected to the fifth chamber 1133 through a channel, and the tenth chamber 11310 is disposed at a side of the fifth groove 11312, which is far from the rotation center 101, and is connected to the fifth groove 11312 through a channel. The fifth groove 11312 is embedded with an adsorption film 11311 (not shown). Seventeenth chamber 1138 may contain a reagent, which may be an eluent, a reagent that binds to the target, or the like. The fifth groove is provided with a chamber on both the side close to the rotation center 101 and the side far from the rotation center 101. The depth of the fifth groove is greater than the depth of the chambers on both sides.

The specific procedure for separating samples, e.g. fibrinogen (clotting factor, about 34 Da) in blood, using the biomaterial separation device in fig. 9 is as follows (see labels in fig. 8 and 9):

a blood sample to be separated is added to the fourth chamber 1111:

S2: stopping standing for a period of time. The second capillary 1131 fills with plasma due to capillary effect. The standing time T2 may be 5s to 50s. During resting, a solution containing fibrinogen antibodies may be added to seventeenth chamber 1138.

S3: the biological material separation device is controlled to perform high-speed rotation, at this time, the second capillary tube 1131 which is filled in the biological material separation device has a siphon effect, the plasma in the first chamber 1121 flows out through the second capillary tube 1131 and enters the sixth chamber 1136, and then the plasma is stored in the fifth chamber 1133 after passing through the first membrane 1132 arranged in the first groove 1134, and the first membrane 1132 can filter out impurities with molecular weight of more than 100 ten thousand Da. The solution in the seventeenth chamber 1138 also enters the fifth chamber 1133 through the fourth channel 1139. In the fifth chamber 1133, the target fibrinogen binds to fibrinogen antibodies to form a complex. The rotation speed range can be 2000-4500 revolutions/min, and the time T3 can be 15-120 s.

S4: standing for a period of time promotes accelerated mixing of the two solutions in the fifth chamber 1133. The standing time T4 may be 120s to 480s.

S5: the rotation speed of the biological material separation device is controlled, after the rotation is completed, fibrinogen and fibrinogen antibody complex is adsorbed on the adsorption film 11311, and other solution is introduced into the tenth chamber 11310. And taking out the adsorption film 11311 to elute to obtain fibrinogen. The rotation speed range can be 2500-5500 rpm, and the time T5 can be 15-120 s.

The adsorption film 11311 and tenth chamber 11310 in fig. 9 may be replaced with similar first chamber 1121, connection 1122 and second chamber 1123, and the fibrinogen and fibrinogen antibody complex is precipitated into the second chamber 1123 by centrifugation in step S5, and the other liquid is introduced into the first chamber 1121.

In fig. 9, the seventeenth chamber 1138 may be omitted, and the fibrinogen antibody contained in the seventeenth chamber 1138 may be directly immobilized on the adsorption film 11311, and when the centrifugation in S5 is performed, the fibrinogen stored in the fifth chamber 1133 may be directly bound to the fibrinogen antibody immobilized on the adsorption film 11311.

Another embodiment of the present application is shown in fig. 10. The biological material separating apparatus of this embodiment includes not only one extraction mechanism and the first filter unit in fig. 9, but also a second filter unit connected to the first filter unit through a fifth passage 1141. The second filter unit includes a second membrane 11410 for intercepting a portion of the material flowing from the fifth channel 1141.

The second filter unit includes a second recess 1144, and a second membrane 11410 (not shown) is embedded in the second recess 1144. The side of the second recess 1144 remote from the rotation center 101 is provided with an eighth chamber 1143, and the eighth chamber 1143 is connected to the second recess 1144 through a sixth channel 1145.

In this embodiment, the sixth channel 1145 is curved and includes a first branch channel, a second branch channel and a third branch channel, where the first branch channel and the third branch channel are straight channels, and the second branch channel is a curved channel. The first branch channel and the third branch channel are not on the same straight line, and a certain included angle is formed between the first branch channel and the third branch channel. In this embodiment, the first branch channel is connected to the second recess 1144, the third branch channel is connected to the eighth chamber 1143, and the number of bends of the middle second branch channel is greater than or equal to 1. The curved channel on the one hand contributes to an increased flow path length; on the other hand, the degree of mixing of the mixture in the sixth passage 1145 can be reduced.

In this embodiment, the ninth chamber 1149 is connected to the first branch passage through a seventh passage 1147. Preferably, the seventh channel 1147 is inclined in the upstream direction of the sixth channel 1145 by an angle of more than 0 ° and less than 90 °. The angle of inclination in this embodiment is 60 °. The seventh passage 1147 is inclined in the upstream direction of the sixth passage 1145, so that the liquid can be prevented from entering the ninth chamber 1149 when controlling the centrifugation of the liquid into the eighth chamber 1143. Preferably, the ninth chamber 1149 is disposed at a side of the second recess 1144 remote from the rotation center 101.

In this embodiment, an eighteenth chamber 1146 and a nineteenth chamber 1149 are disposed next to two sides of the second recess 1144, respectively, the fourth chamber 1141 is connected to the second recess 1144 through the eighteenth chamber 1146, and the first branch channel is connected to the second recess 1144 through the nineteenth chamber. Preferably, the ninth chamber 1149 is provided at a side of the nineteenth chamber remote from the rotation center 101.

In this embodiment, the junction where the second recess 1144 transitions to the first branch passage is provided with a step to prevent the liquid entering the eighth chamber 1143 from flowing into the second recess 1144 again. In this embodiment, the steps may be replaced with slopes having a certain gradient.

In the present embodiment, the twentieth chamber 1148 is connected to the eighteenth chamber 1146 through an eleventh passage, and the twentieth chamber 1148 is disposed on a side of the eighteenth chamber 1146 near the rotation center 101.

In this embodiment, a third capillary tube 1142 may be provided between the fifth channel 1141 and the second filter unit.

The biological material separation device of fig. 10 is used to separate samples, such as fibrinogen from blood samples, as follows:

a blood sample to be separated is added to the fourth chamber 1111:

S1: the rotational speed of the biological material separating apparatus is controlled, and after the rotational speed is completed, the first chamber 1121 is filled with plasma, blood cells enter the second chamber 1123, and excessive blood enters the third chamber 1125. The rotating speed range can be 2500-5500 rpm, and the time T1 can be 120-480 s.

S3: the biomaterial separation device is controlled to perform high-speed rotation, and at this time, the second capillary 1131 filled with the biomaterial separation device has a siphon effect, and the plasma in the first chamber 1121 flows out through the second capillary 1131, passes through the first membrane 1132 disposed in the first groove 1134, and is stored in the fifth chamber 1133. The solution in the seventeenth chamber 1138 also enters the fifth chamber 1133 through the channels. In the fifth chamber 1133, the target fibrinogen binds to fibrinogen antibodies to form a complex. The rotation speed range can be 2000-4500 revolutions/min, and the time T3 can be 15-120 s.

S4: the biological material separation device is controlled to circularly rotate at high-low-high speed to promote the accelerated mixing of the two solutions in the fifth chamber 1133, the high speed in alternation can be 2500-4000 rpm, and the time T4 can be 3-20 s; the low speed can be 300-2000 rpm, the time T5 can be 3-20 s, the directions of the two speeds can be the same direction or reverse direction, and the number of times of alternate running can be 3-15 times.

S5: the third capillary 1142 is stopped standing for a while, and is filled with the mixed liquid by capillary action. The standing time T6 may be 5s to 50s.

S6: the biomaterial separation device is controlled to rotate at a high speed, and at this time, the third capillary 1142 filled with the mixed solution in S5 has a siphon effect, the mixed solution in the fifth chamber 1133 passes through the fifth channel 1141 and is transferred to the eighth chamber 1143 through the sixth channel 1145 after passing through the second membrane 11410 embedded in the second recess 1144, and the fibrinogen and fibrinogen antibody complex is retained on the second membrane 11410. The speed can be 2000-5000 rpm, and the time T7 can be 20-120 s.

S7: stopping standing for a period of time; eluent is added to the twentieth chamber 1148 during rest. The standing time T8 may be 5s to 40s.

S8: the biomaterial separation device is controlled to rotate at a high speed, and the eluent stored in the twentieth chamber 1148 passes through the channels to elute fibrinogen and fibrinogen antibody complex retained on the second membrane 11410. After passing through the second membrane 11410, the eluted complex solution fills the void space of the sixth channel 1145 in a small portion and is transferred from the seventh channel 1147 to the ninth chamber 1149 for collection in a large portion. The speed can be 5500-8000 rpm, and the time T9 is 7-20 min.

In another embodiment of the present application, as shown in fig. 11, the biomaterial separation device further includes a third recess 2015, the third recess 2015 being connected to the seventh channel 1147. A third film 2012 may be embedded in the third recess 2015. The thirteenth chamber 2011 is connected to a side of the third recess 2015 remote from the center of rotation through a ninth channel 2018. In this embodiment, the ninth channel 2018 may be a straight channel or may include a curved channel, but the number of bends of the curved channel of the ninth channel is smaller than the number of bends of the fifth channel. As shown in fig. 10, the ninth channel 2018 is a straight channel, and includes a fourth branch channel and a fifth branch channel, wherein the fourth branch channel is connected to the thirteenth chamber 2011, and the fifth branch channel is connected to the third recess 2015.

In this embodiment, a twelfth chamber 2017 and a fifteenth chamber are respectively disposed at two sides of the third recess 2015, the fifth branch channel is connected to the third recess 2015 through the fifteenth chamber, and the fourteenth chamber 2016 is connected to one side of the twelfth chamber 2017 near the rotation center through the eighth channel.

In this embodiment, the eleventh chamber 2013 is connected to the fifth branch passage through a tenth passage 2019. Preferably, the inlet direction of the tenth channel 2019 is inclined to the upstream direction of the ninth channel by an angle of more than 0 ° and less than any angle between 90 °. In this embodiment, the angle of inclination is 60 °.

In this embodiment, the sixteenth chamber 2014 is connected to a side of the eleventh chamber 2013 near the rotation center through a passage.

As shown in fig. 11 and 12, the bottom of the seventeenth chamber 1138 is provided with a first support table 11381 and a first boss 11382, and the first support table 11381 and the first boss 11382 cooperate with each other to support the first container placed in the seventeenth chamber 1138. A second concave wall 11383 is provided on a cavity wall of the seventeenth chamber 1138 toward the direction of the inside of the cavity for fixing the first container placed in the seventeenth chamber 1138. The top of the first boss 11382 is provided with a first flow channel 11384 for draining the liquid disposed in the first container.

As shown in fig. 12, in a front view of the biological material separating apparatus, the top of the first boss 11382 is at the same height as the table top of the first support table 11381, or the top of the first boss 11382 is higher than the table top of the first support table 11381.

The first support table 11381 and the first boss 11382 are used for supporting the first container, when the first container is pressed downwards by force, the first container takes the first support table 11381 as a pivot, leverage is generated on the first container, and the first boss 11382 will crush the first container, so that the reagent in the first container enters the seventeenth chamber 1138. The first flow channel 11384 facilitates drainage of the reagent in the first container into the seventeenth chamber 1138.

In this embodiment, the first container is used to store the reagent, and a certain pressure is applied to the first container to squeeze the reagent out of the first container, so as to release the reagent therein. For example, as shown in fig. 13, the first container includes a second cavity 1201 and a first release hole 1202 provided at a bottom surface of the second cavity 1201, and a first sealing film 1203 is provided at one side of the first release hole 1202 for sealing the first release hole 1202.

Preferably, the first release hole 1202 is aligned with the first boss 11382 when the first container is placed in the seventeenth chamber 1138.

Preferably, a first concave wall 1204 is further disposed on the side of the second cavity 1201, and the first concave wall 1204 cooperates with the second concave wall 11383 to fix the first container, and prevent the first container from generating large rotational displacement on the bottom plate, so as to ensure that the first release hole 1202 sealed by the first sealing film 1203 is always located above the first boss 11382, and can effectively rupture the first container when the first container is pressed.

In this embodiment, the twentieth chamber 1148 may be identical to the seventeenth chamber 1138. I.e., the bottom of the twentieth chamber 1148, is provided with a second support table and a second boss, which can support a second container placed in the twentieth chamber 1148. A third recess wall is provided in the wall of the twentieth chamber in the direction of the chamber for securing a second container disposed in the twentieth chamber 1148. The top of the second boss may be provided with a second flow channel for draining liquid in the second container.

In this embodiment, the second container may be the same as the first container, that is, the second container includes a second cavity and a second release hole disposed on a bottom surface of the second cavity, and a second sealing film is disposed on one side of the second release hole. The second release aperture is opposite the second boss when the second container is placed in the twentieth chamber 1148. The second container may further comprise a fourth recess wall, the fourth recess wall cooperating with the third recess wall.

In this embodiment, the first sealing film 1203 and the second sealing film may be aluminum plastic films.

As shown in fig. 11, in order to maintain the balance of the entire biological material separating apparatus while the biological material separating apparatus is rotated around the rotation center, a plurality of balance chambers, such as a balance chamber 1021, a balance chamber 1022, and a balance chamber 1023, are provided on the bottom plate, and ventilation holes, such as the balance chamber 1021 and the balance chamber 1022, are provided in part of the balance chamber 102, and

ventilation holes

10211 and 10221, respectively, are provided in the balance chamber 1021 and the balance chamber 1022, so that the air pressure in the entire biological material separating apparatus is the same as the outside, to ensure the flow of the liquid in the biological material separating apparatus.

In this embodiment, the first container may be integrally formed, or as shown in fig. 13, the first container further includes a top cover 1205, and the second cavity 1201 cooperates with the top cover 1205 to form a container for sealing the reagent disposed in the second cavity.

In this embodiment, the first concave wall 1204 and the second concave wall 11383 cooperate to limit the rotational displacement of the first container on the bottom plate, and thus other ways of limiting the rotational displacement of the first container on the bottom plate are also suitable for the present application. For example by means of a snap-fit or by designing the first and second recess walls as protruding walls protruding outwards of the chamber, etc.

In this embodiment, the first sealing film 1203 may be disposed inside the second cavity 1201 or may be disposed outside the second cavity 1201.

If the first sealing film 1203 is disposed inside the second cavity 1201 and the first release hole 1202 is deeper (i.e., the bottom surface of the second cavity 1201 is thicker), the top of the first boss 11382 goes deep into the first release hole 1202, but the top of the first boss 11382 does not contact the first sealing film 1203, or the top of the first boss 11382 slightly contacts the first sealing film 1203, but the first sealing film has a certain supporting function, and the first sealing film is not crushed under the condition that no external pressure is applied.

If the first sealing film 1203 is disposed on the outer side of the second cavity 1201, that is, the first sealing film 1203 is disposed on the side of the first release hole 1202 close to the first boss 11382, the top of the first boss 11382 is slightly higher than the table surface of the first support table 11381, but the top of the first boss 11382 does not contact the first sealing film 1203, or the top of the first boss 11382 slightly contacts the first sealing film 1203, but the first sealing film has a certain supporting effect, and the first sealing film cannot be broken by being squeezed under the condition that no external pressure is applied.

In the above embodiment, the first film 1132, the second film 11410, the third film 2012, and the adsorption film 11311 may be adhered on the hard substrate and then respectively embedded in the respective grooves together with the hard substrate, thereby effectively firmly placing the films in the flow paths. Taking the first film 1132 as an example, the hard substrate is provided as shown in fig. 14. In fig. 14, the hard substrate includes a first substrate 4011, a first film 1132 is adhered to one side of the first substrate 4011, a window 4012 is formed in the first substrate 4011, and the window 4012 penetrates through one surface of the first substrate facing the first film 1132 and one surface facing away from the first film 1132. In this embodiment, the first film 1132 is attached to the first substrate 4011 by bonding. The bonding mode can be specifically adhesive bonding, thermocompression bonding, ultrasonic welding or the like. Preferably, the first film 1132 is thermocompression bonded on the first substrate 4011. In the present embodiment, the depth of the first groove 1134 for placing the first film 1132 is larger than the depths of the sixth and

seventh chambers

1136 and 1137 adjacent to the first groove 1134 for better fixing the first film 1132. For example, the depth of the first groove 1134 is 0.2mm to 7.8mm, the width is 0.2mm to 5mm, the depths of the sixth cavity 1136 and the seventh cavity 1137 are 0.1mm to 6.8mm, and the like.

In this embodiment, the first film 1132 is integrated on the bottom plate through the hard substrate, so that the film can be firmly integrated on the bottom plate, and the first film 1132 can be placed in the first groove 1134 perpendicular to the bottom plate surface, so that the effective utilization rate of the film can be ensured by placing the first film 1132 in the first groove 1134 parallel to the bottom plate surface.

In another embodiment of the present application, as shown in fig. 15, the hard substrate includes a first cavity 4111 and a sleeve 4114, where the sleeve 4114 is detachably disposed in the first cavity 4111 and is used with the first cavity 4111. The bottom of the first cavity 4111 is provided with a first outflow hole 4112. The first membrane 1132 is disposed at one end of the sleeve 4114. In use, the sleeve 4114 is placed in the first cavity 4111. In this embodiment, a plurality (2 or more) of first outflow holes 4112 may be disposed at the bottom of the first cavity 4111, so that the liquid in the sleeve 4114 can quickly enter the downstream channel. Preferably, the bottom of the first cavity 4111 is provided with a fourth groove 4113, and the first outflow hole 4112 is disposed in the fourth groove 4113, so that the fourth groove 4113 may be used as a channel for liquid circulation, and further drain the liquid into the downstream channel rapidly. When the number of the first outflow holes 4112 is plural, a membrane supporting table 4115 is disposed between the adjacent first outflow holes 4112, for supporting the first membrane 1132, so as to prevent the first membrane 1132 from being broken due to excessive stress. The number of the membrane support tables 4115 may be set as needed, for example, when the number of the first outflow holes is large, the gap may be set.

In this embodiment, the first membrane 1132 and the sleeve 4114 may be connected by bonding, for example, adhesive bonding, thermocompression bonding, ultrasonic welding, or the like. Preferably, the first membrane 1132 is thermocompression bonded to one end of the sleeve 4114.

When separating a sample using the biological material separating apparatus shown in fig. 11, for example, fibrinogen in blood, a first container is placed in the seventeenth chamber 1138, a second container is placed in the twentieth chamber 1148, a hard substrate containing the first film 1132 is placed in the first groove 1134, a hard substrate containing the second film 11410 is placed in the second groove 114, and a hard substrate containing the third film 2012 is placed in the third groove 2015. The hard substrate may be selected from the hard substrates shown in fig. 14 or 15. The following will describe a hard substrate in fig. 14 as an example.

The first container contains fibrinogen antibody solution, the second container contains eluent for eluting fibrinogen, and the fibrinogen separation comprises the following specific steps:

a blood sample to be separated is added to the fourth chamber 1111:

s1: the biological material separation device is controlled to rotate at a high speed, and after the rotation is completed, the first chamber 1121 is filled with plasma, blood cells enter the second chamber 1123, and excessive blood enters the third chamber 1125. The rotating speed range can be 2500-5500 rpm, and the time T1 can be 120-480 s.

S2: stopping standing for a period of time, and filling the second capillary 1131 with plasma due to capillary effect; during resting, pressure is applied to the first container, under pressure, the first boss 11382 squeezes the first sealing membrane 1203 apart, causing the first container to release the stored fibrinogen antibody solution from the first release hole 1202 into the seventeenth chamber 1138. The standing time T2 may be 5s to 50s.

S3: controlling the biological material separating device to perform high-speed rotation, wherein the filled second capillary tube 1131 has a siphon effect, and the plasma in the first chamber 1121 flows out through the second capillary tube 1131, passes through the first membrane 1132 embedded in the first groove 1124 and is transferred into the fifth chamber 1133; the fibrinogen antibody solution stored in the first container is also transported through the channel into the fifth chamber 1133. In the fifth chamber 1133, the target fibrinogen binds to fibrinogen antibodies to form a complex. The rotation speed range can be 2000-4500 revolutions/min, and the time T3 can be 15-120 s.

S5: stopping standing for a period of time, and filling the third capillary 1142 with the mixed liquid; the standing time T6 may be 5s to 50s.

S6: the biomaterial separation device is controlled to rotate at a high speed, and at this time, the third capillary 1142 filled with the mixed fluid in S5 has a siphon effect, and the mixed fluid in the fifth chamber 1133 passes through the second membrane 11410 embedded in the second recess 1144 and is transferred to the eighth chamber 1143 through the sixth channel 1145, and the fibrinogen and fibrinogen antibody complex is retained on the second membrane 11410. The speed can be 2000-5000 rpm, and the time T7 can be 20-120 s.

S7: stopping standing for a period of time; during resting, pressure is applied to the second container such that the second boss squeezes the second container, and the second boss squeezes and ruptures the second sealing membrane on the second container under pressure such that the second container can release the stored eluent from the second release aperture and into the twentieth chamber 1148. The standing time T8 may be 5s to 40s.

S8: the biological material separation device is controlled to rotate at a high speed, the eluent stored in the twentieth chamber 1148 passes through the channels, the fibrinogen and fibrinogen antibody complex retained on the second membrane 11410 is eluted, and after the eluted complex solution passes through the second membrane 11410, a small part of the solution fills the gaps of the sixth channel 1145, and most of the solution is transferred from the seventh channel 1147 to the ninth chamber 1149 for collection. After the ninth chamber 1149 is filled, a portion of the eluted complex solution enters the third recess 2015, the fibrinogen and fibrinogen complex is intercepted on the third mould 2012, and other liquid enters the thirteenth chamber 2011 through the ninth channel 2018. The speed can be 5500-8000 rpm, and the time T9 is 7-20 min.

S9: stopping standing for a period of time. Fibrinogen and fibrinogen antibody complex is intercepted on the third membrane 2012; to the fourteenth chamber 2016, a dissociation liquid (e.g., the dissociation liquid includes 100 uL of sodium chloride solution (0.15M) at ph 11) is added); to the sixteenth chamber 2014 is added a detection solution (e.g., the detection solution includes 10uL sodium sulfite (0.6M), 10uL nano-silver colloid (1 ml/L) and 10uL 2 amino-2 methyl-1 propanol (AMP) buffer (80 mM)). The standing time T10 was 30s.

S10: the biological material separation device is controlled to circularly rotate at high-low speed, the high speed in alternation can be 2500-3000 rpm, and the time can be 3-20 s; the low speed can be 300-1000 rpm, the time can be 3-20 s, the directions of the two speeds can be the same direction or reverse direction, and the number of times of alternate running can be 3-15 times. In this step, fibrinogen dissociates from the fibrinogen antibody complex, freeing fibrinogen, and the liquid in the sixteenth chamber 2014 flows into the eleventh chamber 2013. The phase time T11 may be 20-40min.

S11: the biological material separation device is controlled to rotate at a high speed so that fibrinogen flows into the tenth channel 2019 through the third membrane 2012 into the eleventh chamber 2013 to react with the detection liquid already present in the eleventh chamber 2013 for 25 minutes. The speed can be 5500-8000 rpm, and the total time T12 can be 30min. And finally, directly detecting at the detection hole, wherein the detection wavelength is 630nm.

In another embodiment of the present application, the biomaterial separation device further comprises a cap plate adapted to the base plate. The present embodiment is described by taking a cover plate adapted to the bottom plate in fig. 11 as an example. As shown in fig. 16, a rotation center 201 may be provided on the cover plate as well, corresponding to the rotation center 101 of the base plate. Just as for the position of the fourth chamber 1111 near the rotation center, the cover plate is provided with a first through hole 202. If the bottom plate is provided with the first sample loading hole 1112, the first through hole 202 is opposite to the first sample loading hole 1112. For the seventeenth chamber 1138, a first top pinhole 203 is provided on the cover plate; for the twentieth chamber 1148, the cover plate is provided with a second top pinhole 204; for the fourteenth chamber 2016, a third through hole 205 is provided in the cover plate; a second through hole 206 is provided in the cover plate for the sixteenth chamber 2014;

air holes

207 and 208 are provided opposite the

trim chambers

1021 and 1022, respectively.

The biological material separation device provided by the application can be used for separating and detecting exosomes which are research hot spots in the current biological field.

Exosomes are released outside the cell in the form of exocrine after fusion of the cell membrane with the intracellular multivesicular bodies (multivesicular bodies), approximately 40-100 nm in diameter. Carrying various proteins, lipids, DNA, mRNA, miRNA and the like related to cell sources, and participating in the processes of intercellular communication, cell migration, angiogenesis, immunoregulation and the like. The rise of exosomes is found in diabetes, cardiovascular diseases, aids, chronic inflammatory diseases and cancer, which are highly likely to be diagnostic markers of such diseases, and therefore, accurate qualitative and quantitative studies of exosomes are particularly important.

The existing exosome extraction method mainly comprises differential centrifugation, density gradient centrifugation, polymer precipitation (PEG-base precipitation), ultrafiltration, magnetic bead immunization, kit method, etc. Differential centrifugation is the most commonly used means for purifying extracellular vesicles, and low-speed centrifugation and high-speed centrifugation are alternately performed to separate vesicle particles with similar sizes. Density gradient centrifugation is a method of enriching extracellular vesicles by forming a density hierarchy using ultracentrifugation. PEG-base precipitation, which exploits the co-precipitation properties of polyethylene glycol (PEG) in combination with hydrophobic proteins and lipid molecules, was previously applied to the collection of viruses from serum and the like, is now also used to precipitate exosomes, the principle of which may be related to the competitive binding of free water molecules. Ultrafiltration centrifugation is a selective separation using ultrafiltration membranes of different cut-off relative molecular masses. The magnetic bead immune method utilizes the specific markers (such as CD63 and CD9 protein) on the surface of the exosome, and the exosome can be adsorbed and separated by incubating the exosome with the magnetic beads coated with the anti-marker antibody and then combining. The above methods are all based on routine procedures in the laboratory to achieve separation of exosomes. Under the operation of a conventional laboratory, the method for extracting the exosomes generally has the defects of long time, complex operation, large-scale instrument requirement, high cost and the like. The biological material separation device provided by the application is utilized to separate the exosome, multiple sample adding and sample changing are not needed, the operation of the extraction process is simple and convenient, the time required by the extraction process is short, and the efficiency is high.

In the detection of exosomes, the size and morphology of exosomes are mainly analyzed by an electron microscope, cell surface markers are analyzed by a flow cytometer, proteins are analyzed by Western blot, ELISA and other methods, or RNA is analyzed by qPCR and New Generation Sequencing (NGS). The utilization of the above method requires not only expensive equipment such as transmission electron microscope equipment, flow cytometry, etc., but also a lot of time for performing the exosome detection analysis and complicated operations such as performing Western blot and ELISA experiments, etc.

The biological material separation device can separate and extract exosomes, and can detect exosomes. Specifically, the biological material separation device shown in fig. 11 is combined with a cover plate to form a separated biological material separation device, and a quantitative separation of a whole blood sample is taken as an example.

The biomaterial separation device and the cover in fig. 11 are made of Polycarbonate (PC) material, and the channels are hydrophilized, for example, by using a silicone surfactant or ethanol. The channels are treated with a silicone surfactant in this example. The first groove 1134 places the hard substrate in fig. 14, and the second groove 1144 and the third groove 2015 place the hard substrate in fig. 15.

The first container is preloaded with 120 μl of the loading solution and placed in the seventeenth chamber 1138.

The second container is preloaded with 200 μl of eluent and placed in the twentieth chamber 1148.

A hard substrate including a first film 1132 is placed in the first groove 1134. Preferably, when placed, the side of the window 4012 to which the first film 1132 is adhered is close to the center of rotation 101.

The rigid substrate containing the second film 11410 is placed in the second recess 1144. One end of the sleeve 4114 provided with the second membrane 11410 is placed at the bottom of the first cavity 4111, so that the side of the sleeve 4114 containing the second membrane 11410 is tightly bonded to the bottom of the first cavity 4111. The sleeve 4114 and the first cavity 4111 may be designed with an interference fit to ensure the tightness of the second membrane 11410, that is, ensure that all the liquid can reach the first cavity 4111 only after passing through the second membrane 11410, or a small amount of silica gel, that is, other types of glue, outside the sleeve 4114 to seal the gap more effectively. The entire rigid substrate is then placed in the second recess 1144, preferably with the second membrane 11410 adjacent the center of rotation 101.

Similar to the rigid substrate containing the second film 11410, the rigid substrate containing the third film 2012 is placed in the third recess 2015. Preferably, the third membrane 2012 is placed away from the center of rotation 101.

The quantitative separation and detection process of exosomes is specifically as follows:

through the first through hole 202 of the cover plate, 500 μl (may be 250uL to 550 uL) of blood sample to be separated is added to the fourth chamber 1111:

s1: the biomaterial separation device is controlled to rotate at a high speed so that the blood sample flows into the extraction unit from the fourth chamber 1111. The first chamber 1121 is filled with plasma, blood cells enter the second chamber 1123, and excess blood enters the third chamber 1125. The rotation speed range may be 2500 rpm to 5500 rpm, the time T1 may be 120s to 480s, the rotation speed in this embodiment is 4500 rpm, and T1 is 150s.

S2: and (5) standing. The second capillary 1131 fills with plasma due to capillary effect; during the standing period, the first container is pressed downwards by the thimble through the first thimble hole 203 of the cover plate, under the action of pressure, the first boss 11382 takes the first supporting table 11381 as a fulcrum to perform leverage, and the first boss 11382 presses and breaks the first sealing film 1203, so that the first container releases the loading liquid from the first release hole 1202 and enters the seventeenth chamber 1138. The standing time T2 may be 5s to 50s. In this embodiment, T2 is 30s.

S3: controlling the rotating motion of the biological material separating device, wherein the second capillary 1131 filled with the biological material separating device has a siphon effect, and the plasma in the first chamber 1121 flows out through the second capillary 1131, passes through the first membrane 1132 embedded in the first groove 1124 and is transferred into the fifth chamber 1133; the loading fluid in the seventeenth chamber 1138 also flows into the fifth chamber 1133 through the channel to mix with the plasma. The rotation speed range can be 2000-4500 revolutions/min, and the time T3 can be 15-120 s. In this example, the rotation speed was 2500 rpm and the time was 20s.

S4: the biological material separation device is controlled to perform high-low-high speed circulation rotation to promote the accelerated mixing of the two solutions in the fifth chamber 1133. The high speed in the alternation can be 2500-4000 rpm, and the time T4 can be 3-20 s; the low speed can be 300-2000 rpm, the time T5 can be 3-20 s, the directions of the two speeds can be the same direction or reverse direction, and the number of times of alternate running can be 3-15 times. In the embodiment, the high speed is 3000 rpm, 5s; the low speed is 500 revolutions per minute, 5s; the number of times was 10.

S5: standing for a period of time. At this time, the third capillary 1142 is filled with the mixed liquid due to the capillary effect; the standing time T6 may be 5s to 50s. In this embodiment, T6 is 30s.

S6: the biological material separating apparatus is controlled to rotate, and at this time, the third capillary 1142 filled with the mixed liquid has a siphon effect, and the mixed liquid in the fifth chamber 1133 passes through the second membrane 11410 embedded in the second groove 1144 and is transferred to the eighth chamber 1143 through the sixth channel 1145. The speed can be 2000-5000 rpm, and the time T7 can be 20-120 s. In this example, the speed was 3000 rpm and T7 was 60s.

S7: stopping, and standing for a period of time. During the standing period, pressure is applied to the second container through the second top pinhole 204 of the cover plate, so that the second container takes the second supporting table as a supporting point, leverage is generated, the second boss presses the second container, and under the action of the pressure, the second boss presses and breaks the second sealing film on the second container, so that eluent in the second container can be released from the second release hole and enter the twentieth chamber 1148. The standing time T8 may be 5s to 40s. In this embodiment, T8 is 30s.

S8: after the eluting solution stored in the twentieth chamber 1148 passes through the second membrane 11410 and a small part of the eluting solution fills the gap of the sixth channel 1145, most of the eluting solution is transferred from the seventh channel 1147 to the ninth chamber 1149 for collection, i.e. the eluted exosomes, for later study.

After the ninth chamber 1149 is filled, a portion of the eluent enters the third recess 2015 for desalting. The speed can be 5500-8000 rpm, and the time T9 is 7-20 min. In this example, the rotation speed is 5500 rpm, and T9 is 10min.

S9: stopping, and standing for a period of time. 100ul of bead-aptamer-Probe-HRP solution is added to the fourteenth chamber 2016 through the third through hole 205 in the cover plate; 10uL H is added to the sixteenth chamber 2014 through the second through hole 206 in the cover plate ₂ O ₂ (1 mM) and 10uL of Amplex Red (10 uM) (fluorescent Red dye, protected from light). The standing time T10 was 30s.

S10: the biological material separation device is controlled to circularly rotate at high-low speed, the high speed in alternation can be 2500-3000 rpm, and the time can be 3-20 s; the low speed can be 300-1000 rpm, the time can be 3-20 s, the directions of the two speeds can be the same direction or reverse direction, and the number of times of alternate running can be 3-15 times. The high speed in this example is 3000 rpm, 15s; the low speed was 500 rpm, 15s. At this point, the solution in the fourteenth chamber 2016 enters the fifteenth chamber 2015 through a channel, wherein MB-Aptamer binds to exosomes, freeing Probe-HRP; the solution in the sixteenth chamber 2014 passes through the channel into the eleventh chamber 2013.

S11: controlling the biological material separation device to rotate at a high speed, after the free Probe-HRP passes through the third membrane 2012, the free Probe-HRP enters the eleventh chamber 2013 through the tenth channel 2019 and the H existing in the eleventh chamber 2013 ₂ O ₂ And an duplex Red solution. The speed can be 5500-8000 rpm, and the time T12 can be 25-40min. In this example, the speed was 5500 rpm and the time was T12 30min. Finally, fluorescence detection is directly carried out at the detection hole, the excitation wavelength is 535nm, and the emission wavelength is 584nm.

Experimental results:

the ninth chamber 1149 was collected with a total of 95. Mu.L of exosome solution, and the exosome was detected and tracked by NTA (Nanoparticle Tracking Analysis), and the experimental results are shown in FIG. 17. The exosomes were observed using transmission electron microscopy and the experimental results are shown in fig. 18-19.

As can be seen from FIG. 17, the average particle diameter of the exosomes was about 93.7nm, and the exosomes concentration was about 4.1X10% ⁶ And each mL.

As can be seen from fig. 18 and 19, the collected exosomes are intact.

When utilizing the biomaterial separator separation that this application provided to detect the exosome, not only easy operation, required time is short moreover, improvement detection efficiency that can be by a wide margin.

The technical features of the above embodiments may be arbitrarily combined, and all possible combinations of the technical features in the above embodiments are not described for brevity of description, however, as long as there is no contradiction between the combinations of the technical features, they should be considered as the scope of the description.

The foregoing examples illustrate only a few embodiments of the invention, which are described in detail and are not to be construed as limiting the scope of the invention. It should be noted that it will be apparent to those skilled in the art that several variations and modifications can be made without departing from the spirit of the invention, which are all within the scope of the invention. Accordingly, the scope of protection of the present invention is to be determined by the appended claims.

Claims

1. The biological material separating device is characterized by comprising a bottom plate, wherein the bottom plate comprises at least one extraction mechanism, the extraction mechanism comprises an extraction unit, the extraction unit comprises a first chamber, a connecting part and a second chamber, the first chamber is connected with the second chamber through the connecting part, and the inner diameter of the connecting part is smaller than that of the first chamber and the second chamber.

2. The biological material separation device according to claim 1, wherein the extraction mechanism further comprises a sample introduction unit for inputting and storing a sample to be separated, the extraction unit is used for extracting a target in the sample to be separated, the sample introduction unit is disposed upstream of the extraction unit, and the sample introduction unit is connected to the first chamber through a first channel.

3. The biomaterial separation device as claimed in claim 2, wherein the number of extraction mechanisms is 2 or more, and the sample introduction units of the different extraction mechanisms are connected or disconnected.

4. A biomaterial separation device as claimed in claim 3 wherein the sample injection units of the different extraction mechanisms are connected by a first capillary tube.

5. The biological material separation device of claim 2, further comprising a center of rotation disposed at a middle portion of the biological material separation device.

6. The biological material separation device according to claim 5, wherein the number of the extraction mechanisms is 1, the sample introduction unit and the extraction unit are disposed on opposite sides of the rotation center, and the first channel is arc-shaped around the rotation center.

7. The biomaterial separation device of claim 5, wherein the extraction mechanisms are more than 3, the extraction mechanisms being evenly distributed about the center of rotation.

8. The biomaterial separation device as claimed in claim 7, wherein the extracting means are sequentially arranged around the rotation center in order, and centers of the sampling units of the extracting means are on the same circle.

9. The biological material separation device according to any one of claims 1-8, further comprising a third chamber connected to the extraction unit by a second channel.

10. The biological material separation device of claim 9, further comprising a cover plate that mates with the base plate.