KR102073031B1 - Apparatus for artificial insemination - Google Patents

Abstract

The present invention relates to an artificial insemination device and an artificial insemination method using the same to separate active sperm and inactive sperm out of the sperm and to modify and incubate any one of the active sperm.
The present invention is a sperm injection port is injected sperm; A fertilization port connected to the sperm injection port so that an egg is disposed and an active sperm that recognizes an attractant generated in the ovum among sperm injected into the sperm injection port is moved to be fertilized with the ovum; A dummy port which is formed to be connected to the sperm injection port and moves and collects inactive sperm which does not recognize a attractant generated in the egg among sperm injected into the sperm injection port; And a microchannel connecting the sperm injection port, the fertilization port, and the dummy port, respectively, and receiving a sperm transfer fluid for movement of the sperm. do.

Description

Apparatus for artificial insemination

The present invention relates to an artificial insemination device, and more particularly, to an artificial insemination device for separating inactive sperm and inactive sperm out of sperm and incubating by modifying any one of the sperm and the egg.

Globally, the number of infertile couples is increasing due to environmental hormones, stress, irregular eating habits and the aging of childbirth.

Recently, male infertility is on the rise. These male infertility patients can not have a baby because of in vitro fertilization (IVF) can not normally be fertilized. In vitro fertilization is divided into in vitro fertilization and intracytoplasmic sperm injection (ICSI).

In vitro fertilization is a procedure in which sperm and eggs are collected in vitro to induce spontaneous fertilization. In vitro fertilization is generally more successful in female infertility than in male infertility. Therefore, intracellular sperm injection has been developed as a complementary procedure. The intracellular sperm injection method is a procedure in which the sperm of the testicle or testicular body is collected by a surgical method and artificially directly injected into the egg cell to be fertilized. Such a procedure is used when there are very few sperm in semen or is low in activity, so that it is difficult to fertilize normally, or when there is no semen in the semen or the sperm is blocked due to a vas deferment surgery or the like, and thus the semen does not exist in semen.

On the other hand, dead sperm or cell debris acts as an inhibitor of artificial insemination when in vitro fertilization or intracellular sperm injection is performed. Therefore, the sperm processing (sperm processing) for the selection of the movement sperm is necessary for this procedure.

Conventional sperm screening method includes a process for separating sperm from other components contained in semen by a centrifuge. This can cause sperm damage. In addition, the methods currently in use have low recovery rate of motility sperm, and thus are applicable only to general in vitro fertilization and require a long time and a lot of labor. In addition, conventionally, before performing the intracellular sperm injection method, the doctor selects the visible sperm directly between the dead sperm and the cell debris while visually selecting a lot of time and subjective judgment.

Recently, a technique for separating active sperm with a simple reaction time, a low reaction sample, and a short reaction time has been developed. However, these techniques use laminar flow to separate motile sperm.

In other words, by using a fluid stream having two or more thin layers, the fluid streams flow parallel to each other without causing turbulent mixing at the interface between the streams. However, such laminar flow has the disadvantage of requiring an expensive pumping system, such as a syringe pump or a gravity flow pump, because it must maintain a continuous flow of fluid. In addition, the sample is consumed a relatively large amount, decisively there is a problem that the flow of the sample rather hinders the movement of sperm to specifically separate only the straight motility sperm.

Therefore, there is an urgent need to develop a new technology that can very easily and efficiently separate active and inactive sperm from semen samples of male infertility.

There is also a need for a technique that can separate active sperm and then fertilize the active sperm and eggs to culture.

Korean Registered Patent Publication No. 10-1088984 (2011.12.01 registered notification, the name of the invention: active sperm separation device)

An object of the present invention is to solve the conventional problems, the present invention is to use a fertilization port and a dummy port to separate the active sperm and inactive sperm artificial fertilization device to be able to culture by modifying the active sperm and egg To provide.

In order to achieve the above object of the present invention, the artificial insemination device according to the present invention, the sperm injection port is injected sperm; A fertilization port connected to the sperm injection port so that an egg is disposed and an active sperm that recognizes an attractant generated in the ovum among sperm injected into the sperm injection port is moved to be fertilized with the ovum; A dummy port which is formed to be connected to the sperm injection port and is collected by moving inactive sperm which does not recognize attractants generated in the ovum among sperm injected into the sperm injection port; And a microchannel connecting the sperm injection port, the fertilization port, and the dummy port, respectively, and receiving a sperm transfer fluid for movement of the sperm.

An artificial insemination device according to the present invention, comprising: a culture medium supply port formed to be connected to the fertilization port and supplying a culture solution to the fertilization port for culturing eggs (egg) with active sperm fertilized; And a culture solution discharge port formed to be connected to the fertilization port and discharging the culture solution in the fertilization port to the outside.

In the artificial insemination device according to the present invention, in order to prevent the culture medium supplied to the fertilization port is moved to the sperm injection port by the flow path resistance of the microchannel, the first plug for selectively opening and closing the sperm injection port ; And a second stopper for selectively opening and closing the dummy port in order to prevent the culture solution supplied to the fertilization port from being moved to the dummy port by the flow resistance of the microchannel.

In the artificial insemination device according to the present invention, it is disposed on the upper side of the fertilization port, the lens to observe that the egg is cultured; may further include a.

The artificial insemination method of the artificial insemination device according to the present invention uses the above-described artificial insemination device, the sperm transfer fluid supply step of supplying the sperm transfer fluid for the movement of sperm to the microchannel; A sperm injection step of injecting sperm into the sperm injection port; And a step of confirming whether active sperm recognizing attractants generated from the ovum among the sperm injected into the sperm injection port is moved to the fertilization port and confirmed that fertilized with an egg disposed in the fertilization port. .

In the artificial insemination method of the artificial insemination apparatus according to the present invention, after the identification step, the culture medium supply step of supplying a culture solution for culturing the eggs fertilized active sperm and egg to the fertilization port; may further include.

The artificial insemination device and the artificial insemination method using the same have an effect of accurately separating active and inactive sperm by using a fertilized port in which an egg is disposed and a dummy port in which an egg is not disposed.

In addition, the artificial insemination device of the present invention and the artificial insemination method using the same, there is an effect that can be cultured eggs by supplying the culture solution to the eggs fertilized active sperm and eggs through the fertilization port.

In addition, the artificial insemination apparatus of the present invention and the artificial insemination method using the same, by closing the sperm port and the dummy port using a stopper in the process of supplying the culture solution, it is possible to prevent the culture medium is moved to the sperm port and the dummy port It has an effect.

In addition, the artificial insemination apparatus of the present invention and the artificial insemination method using the same, the lens is provided on the upper side, there is an effect that can be observed that the egg is cultured.

1 is a perspective view schematically showing the structure of an artificial insemination device according to an embodiment of the present invention.
2 is a plan view showing a state in which active sperm and inactive sperm are moved through an artificial insemination device according to an embodiment of the present invention.
3 is a schematic diagram showing a state in which the active sperm is moved to the fertilization port according to an embodiment of the present invention.
Figure 4 is a schematic diagram showing a state in which the inactive sperm collected by moving to the dummy port according to an embodiment of the present invention.
Figure 5 is a schematic diagram showing the process of supplying the culture solution to the fertilization port according to an embodiment of the present invention.
6 is a perspective view showing a stopper and a lens is installed in the artificial insemination device according to an embodiment of the present invention.

Hereinafter, with reference to the accompanying drawings, preferred embodiments of the present invention in which the above-described problem to be solved may be specifically realized. In describing the present embodiments, the same name and the same reference numerals are used for the same configuration and additional description thereof will be omitted below.

1 is a perspective view schematically showing the structure of an artificial insemination device according to an embodiment of the present invention, Figure 2 is a state in which active sperm and inactive sperm is moved through the artificial insemination device according to an embodiment of the present invention Figure 3 is a plan view, Figure 3 is a schematic diagram showing a state in which the active sperm is moved to the correction port according to an embodiment of the present invention.

In addition, Figure 4 is a schematic diagram showing a state in which the inactive sperm collected by moving to the dummy port according to an embodiment of the present invention, Figure 5 is a process for supplying the culture solution to the fertilization port according to an embodiment of the present invention 6 is a perspective view illustrating a stopper and a lens installed in an artificial insemination device according to an embodiment of the present invention.

1 to 6, the artificial insemination device according to an embodiment of the present invention, while separating the active sperm (S1) and inactive sperm (S2) of the sperm active sperm (S1) and egg (O) is fertilized As a device capable of culturing, the artificial insemination device includes a main body 10, a sperm injection port 20, a fertilization port 30, a culture solution supply port 50, a culture solution discharge port 60, The microchannel, the first stopper 81, the second stopper 82, and the lens 90 may be included.

The main body 10 may include a first plate 11 and a second plate 12 stacked on the bottom surface of the first plate 11.

The first plate member 11 is preferably made of a transparent material so that the inside can be visually inspected and the cells can be observed under a microscope when necessary.

Although not particularly limited, a transparent polymer or glass is more preferable among the transparent materials. And, among the transparent polymers, PDMS (polydimethylsiloxane) polymer may be very suitable for the transparent plate of the present invention.

As shown in FIGS. 1 to 3, the sperm injection port 20 may be formed to penetrate the inside of the first plate 11 so that sperm may be injected into the first plate 11. In this case, the sperm may be injected into the sperm injection port through an injection device such as a syringe pump (P).

More specifically, the sperm injection port 20 may pass through one surface of the first plate 11 and pass through the other surface. In this case, the sperm injection port 20 may be connected to one side of the first microchannel 71 recessed in the lower surface of the first plate 11 to be described later.

Therefore, the sperm injected into the sperm injection port 20 can be moved through the first microchannel (71).

The fertilization port 30 may be formed to penetrate the inside of the first plate 11 so that the egg O may be disposed inside the first plank. At this time, the diameter of the crystal port 30 is preferably formed larger than the diameter of the sperm injection port (20). This is to make it possible to place an egg (O) larger than the sperm.

At this time, the fertilization port 30 may be connected to the sperm injection port 20, the active sperm (S1) to recognize the attractant generated from the ovum (O) of the sperm injected into the sperm injection port 20 is moved Can be fertilized with an egg (O)

More specifically, the fertilization port 30 may be formed to pass through from one side of the first plate 11 to the other side in the same manner as the sperm injection port 20. In this case, the correction port 30 may be connected to the other side of the first microchannel 71.

Accordingly, active sperm S1 of the sperm injected into the sperm injection port 20 may be moved to the fertilization port 30 through the first microchannel 71.

The dummy port 40 may be formed to penetrate the inside of the first plate 11, but may be connected to the sperm injection port 20.

The dummy port 40 may be collected by moving inactive sperm S2 which does not recognize the attractant generated from the egg O among the sperm injected into the sperm injection port 20.

More specifically, the dummy port 40 may be formed to penetrate from one side of the first plate 11 to the other side like the other ports. In this case, the dummy port 40 may be connected to one side of the second microchannel 72 branched from the first microchannel 71 to be described later.

Accordingly, inactive sperm S2 of the sperm injected into the sperm injection port 20 may be moved through the first microchannel 71 and then moved to the dummy port 40 through the second microchannel 72. .

The microchannel may be a flow path through which the sperm injected into the sperm injection port 20 is formed recessed in the lower surface of the first plate 11.

The microchannel may include a first microchannel 71 and a second microchannel 72.

One side of the first microchannel 71 may be connected to the sperm injection port 20, and the other side thereof may be connected to the crystal port 30.

The second microchannel 72 may be branched from the first microchannel 71 and connected to the dummy port 40.

That is, the active sperm S1 injected through the sperm injection port 20 is moved to the fertilization port 30 along the first microchannel 71, and the inactive sperm S2 is the first microchannel ( 71 is moved along the second microchannel 72 branched from the first microchannel 71 to the dummy port 40.

Meanwhile, the sperm transport fluid L for moving the sperm may be accommodated in the first microchannel 71 and the second microchannel 72.

Sperm transfer fluid (L) can be used without limitation as long as it is well known to those skilled in the art, for example, human tubal fluid medium or human serum albumin HEA similar in composition to the body fluids of human fallopian tubes (human serum albumin) can be used.

That is, as shown in FIGS. 2 and 3, when sperm is injected into the sperm injection port 20, the sperm is moved along the first microchannel 71. At this time, the active sperm S1 recognizing the attractant generated from the egg O disposed in the fertilization port 30 is moved along the first microchannel 71 to enter the fertilization port 30.

One of the active sperm (S1) of the active sperm (S1) entering the fertilization port 30 is to be fertilized with the egg (O) disposed in the fertilization port (30).

On the other hand, as shown in Figure 2 and 4, inactive sperm (S2) that does not recognize the attractant generated in the egg O of the sperm moving along the first microchannel 71 is the second microchannel It moves along 72 and is collected in the dummy port 40.

As a result, since the fertilization port 30 and the dummy port 40 are provided, the active sperm S1 and the inactive sperm S2 can be separated from the sperm injected into the sperm injection port 20.

As shown in FIG. 5, the culture medium supply port 50 is formed to penetrate the inside of the first plate 11, but is connected to the fertilization port 30, so that the active sperm S1 and the egg O are formed. Culture medium (M) may be supplied to the fertilization port (30) to culture the fertilized egg (E, egg).

At this time, the culture medium (M) can be used without limitation as long as it is well known to those skilled in the art, for example, Tyrode-albumin-lactate-pyruvate culture (TALP medium), serum-free embryo culture (Serum-free embryo culture medium) and the like can be used.

Here, the culture medium (M) may be injected into the culture solution injection port through an injection device such as a syringe pump (P).

More specifically, the culture solution supply port 50 may be formed to pass through one surface of the first plate member 11 to the other surface in the same manner as the sperm injection port 20.

Meanwhile, the microchannel may further include a third microchannel 73 having one side connected to the culture solution supply port 50 and the other side connected to the fertilization port 30.

That is, the culture solution M injected into the culture solution supply port 50 is supplied to the fertilization port 30 through the third microchannel 73.

The culture medium discharge port 60 is formed so as to penetrate the inside of the first plate 11 is connected to the crystal port 30, it can discharge the culture medium (M) in the correction port 30 to the outside.

More specifically, the culture medium discharge port 60 may be formed to pass through from one side of the first plate 11 to the other side in the same manner as the sperm injection port 20.

Meanwhile, the microchannel may further include a fourth microchannel 74 having one side connected to the fertilization port 30 and the other side connected to the culture medium discharge port 60.

That is, the culture solution M, which has supplied the nutrients to the egg E in the fertilization port 30, is discharged to the culture solution discharge port 60 through the fourth microchannel 74.

On the other hand, the second plate 12 is laminated on the lower surface of the first plate 11, the injection port, the correction port 30, the dummy port 40, the culture medium supply port 50 and the culture medium discharged One end of the port 60 is blocked by the second plate 12 except for a passage connected to one side of each microchannel.

As shown in Figure 6, the first stopper 81 is to prevent the culture medium (M) supplied to the fertilization port (30) by the flow resistance of the microchannel to move to the sperm injection port (20), Sperm injection port 20 can be selectively opened and closed.

That is, when the first stopper 81 closes the sperm injection port 20 in the process of supplying the culture medium M to the fertilization port 30, the microchannel connected to the sperm injection port 20 and the fertilization port 30. This is to prevent the culture medium (M) supplied to the fertilization port (30) by the flow resistance of the movement to the sperm injection port (20).

The first stopper 81 selectively opens and closes the dummy port 40 to prevent the culture medium M supplied to the fertilization port 30 from being moved to the dummy port 40 by the flow resistance of the microchannel. can do.

That is, when the second medium closes the dummy port 40 in the process of supplying the culture medium (M) to the fertilization port (30), the flow path resistance of the microchannel connected to the dummy port (40) and the fertilization port (30). This is to prevent the culture medium (M) supplied to the fertilization port 30 is moved to the dummy port (40).

On the other hand, in order to prevent movement to the sperm injection port 20 and the dummy port 40 of the culture medium (M) supplied to the fertilization port 30 by using the first stopper 81 and the second stopper 82 It may be desirable to close the sperm injection port 20 and the dummy port 40 at the same time to further increase the flow path resistance of the microchannel connected to the crystal port 30.

The lens 90 is disposed above the fertilization port 30 to observe that the egg E is incubated.

That is, it is possible to observe that the egg (E) is incubated using the lens 90, and if a problem occurs in the cultured egg (E), the egg (E) is removed from the fertilization port (30). The advantage is that the action can be performed immediately.

Now, an artificial insemination method using an artificial insemination apparatus according to an embodiment of the present invention will be described.

Artificial insemination apparatus according to an embodiment of the present invention may include a transport fluid supply step, sperm injection step, confirmation step and the culture medium supply step.

In the transport fluid supply step, a process of supplying a transport fluid for moving sperm to the first microchannel 71 and the second microchannel 72 is performed.

At this time, the sperm transfer fluid (L) may be supplied through the sperm injection port 20 using the syringe pump (P). Of course, other ports may be used to supply the sperm transfer fluid L.

In the sperm injection step, a process of injecting sperm into the sperm injection port 20 is performed.

At this time, the sperm may be injected into the sperm injection port 20 using the syringe pump (P).

In the checking step, the active sperm (S1) to recognize the attractant generated from the ovum (O) of the sperm injected into the sperm injection port 20 is moved to the fertilization port 30 and the egg is disposed in the fertilization port 30 (O) and the process of checking whether it has been modified is performed.

On the other hand, in the confirming step, the inactive sperm (S2) that does not recognize the attractant generated in the ovum O of the sperm injected into the sperm injection port 20 may check the state collected in the dummy port (40).

In the culture medium (M) supply step is a process for supplying the culture medium (M) for culturing the egg (E) fertilized active sperm (S1) and egg (O) to the fertilization port (30).

At this time, the culture medium (M) may be supplied to the fertilization port (30) through the culture medium (M) injection port using a syringe pump (P).

 On the other hand, the culture medium (M) supply step may also be performed to periodically exchange the culture medium (M) supplied to the fertilization port (30).

That is, when the egg (E) receives the nutrients of the culture (M) for a predetermined time in the process of culturing the egg (E), it is necessary to supply a new culture (M). Therefore, the culture medium (M) deprived of nutrients is discharged through the culture medium discharge port 60 and the process of supplying a new culture medium (M) through the culture medium supply port (50) is performed.

On the other hand, while the egg (E) is incubated, it is possible to observe the incubation process of the egg (E) using the lens (90).

Through this process, the sperm injected into the sperm injection port 20 can be separated into active sperm (S1) and inactive sperm (S2), active sperm (S1) and the egg (O) can be cultured while fertilizing. Will be.

As described above, the artificial insemination device and the artificial insemination method using the same can accurately separate active sperm and inactive sperm by using a fertilized port in which an egg is disposed and a dummy port in which an egg is not disposed. It has an effect.

In addition, the artificial insemination device of the present invention and the artificial insemination method using the same, there is an effect that can be cultured eggs by supplying the culture solution to the eggs fertilized active sperm and eggs through the fertilization port.

In addition, the artificial insemination apparatus of the present invention and the artificial insemination method using the same, by closing the sperm port and the dummy port using a stopper in the process of supplying the culture solution, it is possible to prevent the culture medium is moved to the sperm port and the dummy port It has an effect.

In addition, the artificial insemination apparatus of the present invention and the artificial insemination method using the same, the lens is provided on the upper side, there is an effect that can be observed that the egg is cultured.

While the preferred embodiments of the present invention have been described with reference to the drawings as described above, those skilled in the art will appreciate that the present invention may be modified in various ways without departing from the spirit and scope of the invention as set forth in the claims below. Can be modified or changed.

20: sperm injection port 30: fertilization port
40: dummy port 50: culture medium supply port
60: culture medium discharge port 71: first microchannel
72: second fine channel 73: third fine channel
74: fourth microchannel
S1: Inactive Sperm S2: Inactive Sperm
L: Sperm transfer fluid

Claims (6)

A main body including a first plate made of a transparent material and a second plate stacked on a lower surface of the first plate;
A sperm injection port formed through the first plate and into which sperm are injected;
Is formed through the first plate, the egg is disposed, the active sperm to recognize the attractant generated in the egg of the sperm injected into the sperm injection port is connected to the sperm injection port to be modified with the egg Correction port;
The dummy port is formed through the first plate and connected to the sperm injection port, and the inactive sperm which does not recognize the attractant generated in the ovum among the sperm injected into the sperm injection port is moved and collected. ; And
A microchannel formed in the bottom surface of the first plate and connecting the sperm injection port, the fertilization port, and the dummy port, respectively, and containing a sperm transfer fluid for movement of the sperm; Fix device. The method of claim 1,
A culture medium supply port formed to be connected to the fertilization port and supplying a culture solution to the fertilization port for culturing active eggs and eggs fertilized with eggs (egg); And
It is formed to be connected to the fertilization port, the artificial insemination device further comprises; culture medium discharge port for discharging the culture solution in the fertilization port to the outside. The method of claim 2,
A first stopper for selectively opening and closing the sperm injection port to prevent the culture solution supplied to the fertilization port from being moved to the sperm injection port by the flow resistance of the microchannel; And
And a second stopper for selectively opening and closing the dummy port to prevent the culture solution supplied to the fertilization port from being moved to the dummy port by the flow resistance of the microchannel. . The method of claim 2,
An artificial insemination device further comprising: a lens disposed on the upper side of the fertilization port, the lens to observe that the egg is incubated. delete delete

Images