JP2010166859A - Apparatus for treating biological tissue - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To obtain sufficient stirring, and to simply separate a sufficient number of cells from a biological tissue without increasing the size of the apparatus. <P>SOLUTION: The apparatus 1 for treating a biological tissue includes: a tissue introduction part 2 for introducing a liquid A containing the biological tissue; an enzyme introduction part 4 for introducing a digestive enzyme B; and a stirring flow channel 5 for making a mixed liquid C containing the liquid A introduced from the tissue introduction part 2, and containing the biological tissue, and the digestive enzyme B introduced from the enzyme introduction part 4 and for flowing it while stirring. The stirring flow channel 5 includes: a main pipe 9 for making the liquid A introduced from the tissue introduction part 2, and containing the biological tissue flow; and at least one auxiliary pipe 10 connected to the wall surface of the main pipe 9 and for jetting the digestive enzyme B introduced from the enzyme introduction part 4 in the direction crossing the flow direction of the liquid A containing the biological tissue in the main pipe 9. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

  The present invention relates to a biological tissue processing apparatus.

  Conventionally, in order to separate cells from living tissue, a method is known in which living tissue is suspended in a biocompatible salt solution, digestive enzymes are added, and the living tissue is digested to separate cells ( For example, see Patent Document 1.) In this method, a mixed solution of a saline solution containing a living tissue and a digestive enzyme is placed in a container and stirred to digest the living tissue. Leading is done.

  In addition, a method of digesting a liquid mixture containing a biological tissue and a digestive enzyme while flowing in a pipe connected to a cell concentrator without using a digestion container (for example, patents) is also known. Reference 2).

International Publication No. 2005/012480 Pamphlet International Publication No. 2007/009036 Pamphlet

However, the method of Patent Document 1 has a disadvantage that it takes a considerable amount of time to digest in the container, and the processing is prolonged.
On the other hand, in the method of Patent Document 2, since the digestion is performed while flowing in the pipe, waste of time can be eliminated. However, in order to sufficiently stir the mixed liquid in the pipe, it is made to flow in a turbulent state. Must flow at high speed. And in order to ensure time for an enzyme to digest a biological tissue, making a liquid mixture flow at high speed, it is necessary to lengthen piping sufficiently, and there exists a problem that an apparatus enlarges. Conversely, if the piping is shortened, there is a disadvantage that the number of cells to be separated decreases without sufficient digestion.

  The present invention has been made in view of the circumstances described above, and is a biological tissue treatment that can obtain sufficient agitation and easily separate a sufficient number of cells from the biological tissue without increasing the size of the apparatus. The object is to provide a device.

In order to achieve the above object, the present invention provides the following means.
The present invention includes a tissue introduction part for introducing a liquid containing a biological tissue, an enzyme introduction part for introducing a digestive enzyme, a liquid containing a biological tissue introduced from the tissue introduction part, and a digestion introduced from the enzyme introduction part An agitation channel that causes the mixed liquid containing the enzyme to flow while being agitated, and the agitation channel flows the liquid containing the biological tissue introduced from the tissue introduction part; and A biological tissue processing apparatus comprising: at least one auxiliary pipe connected to a wall surface and ejecting the digestive enzyme introduced from the enzyme introduction part into a main pipe in a direction intersecting a flow direction of the liquid containing the biological tissue I will provide a.

  According to the present invention, when the liquid containing the biological tissue introduced from the tissue introduction part flows in the main pipe constituting the stirring channel, the digestive enzyme introduced from the enzyme introduction part is connected to the main pipe. Since the secondary pipe is ejected into the main pipe in a direction crossing the liquid flow direction, a complicated flow is formed in the main pipe constituting the stirring flow path, and the mixed liquid flows while being stirred. Is done. Thereby, stirring time is ensured by making it flow at a comparatively low flow rate which becomes a laminar flow state without lengthening the main piping, and sufficient digestion can be performed.

That is, a sufficient number of cells can be easily separated from the living tissue without increasing the size of the apparatus.
In addition, since it is not necessary to flow the liquid mixture at a high flow rate that causes a turbulent state, damage to cells contained in the liquid mixture can be reduced and cells in a healthy state can be obtained.

In the said invention, the said subpipe is good also as ejecting the said digestive enzyme toward diagonally forward with respect to the flow direction of the liquid containing the said biological tissue.
In this way, the digestive enzyme jet flowed into the main pipe is prevented from flowing counter to liquid containing biological tissue flowing through the main pipe, causing excessive damage to the cells. You do n’t have to.

Moreover, in the said invention, the said auxiliary | assistant piping may be provided with two or more at intervals in the flow direction of the liquid containing the said biological tissue.
By doing in this way, a liquid mixture is stirred more effectively by the jet stream of the digestive enzyme ejected from several places in main piping.

Moreover, in the said invention, the said subpipe is good also as ejecting the said digestive enzyme in the direction in alignment with the tangent of the cross section of the said main piping in the plane which cross | intersects the longitudinal axis of the said main piping.
By doing in this way, the swirl | vortex flow swirling spirally generate | occur | produces in the main piping which comprises a stirring flow path, and a liquid mixture is stirred more effectively.

Moreover, in the said invention, you may provide the cell concentration part which is connected downstream of the said stirring flow path and concentrates the cell in the liquid mixture which passed this stirring flow path.
In this way, the cell suspension obtained by sufficiently digesting the biological tissue in the stirring channel is supplied to the cell concentration unit connected downstream of the stirring channel and concentrated there. Thus, it can be recovered as a cell group having high purity. The stirring channel can also be used as a pipeline for supplying the cell suspension to the cell concentration unit.

In the above invention, the cell concentrating unit may be a centrifuge that includes a container that contains the mixed liquid that has passed through the stirring channel, and rotates the bottom of the container outward in the radial direction. Good.
By doing in this way, by rotating the container containing the mixed solution so that the bottom part is directed outward in the radial direction, it is possible to separate the cells and the supernatant in order of the specific gravity from the bottom side, Cells can be concentrated easily.

Moreover, in the said invention, the heater which heats the liquid mixture which flows through the inside of this stirring flow path may be provided in the said stirring flow path.
In this way, the mixed liquid flowing through the stirring channel can be heated to a temperature suitable for digestion by operating the heater, promoting digestion, and sufficient digestion and processing time of living tissue. Can be shortened.

  According to the present invention, there is an effect that a sufficient agitation can be obtained and a sufficient number of cells can be easily separated from a living tissue without increasing the size of the apparatus.

1A is an overall configuration diagram showing a biological tissue processing apparatus according to an embodiment of the present invention, FIG. 1B is a longitudinal sectional view of a stirring channel, and FIG. 2C is a transverse sectional view of a main pipe along a cut surface D. It is a whole block diagram explaining the concentration process by the cell concentration part of the biological tissue processing apparatus of FIG. It is a longitudinal cross-sectional view which shows the 1st modification of the stirring flow path of the biological tissue processing apparatus of FIG. It is a cross-sectional view showing a second modification of the stirring channel of the biological tissue processing apparatus of FIG. It is a longitudinal cross-sectional view which shows the 3rd modification of the stirring flow path of the biological tissue processing apparatus of FIG.

A biological tissue processing apparatus 1 according to an embodiment of the present invention will be described below with reference to the drawings.
As shown in FIG. 1 (a), the biological tissue processing apparatus 1 according to the present embodiment includes a liquid A (biocompatible) containing a biological tissue (eg, adipose tissue) collected from a living body (eg, a human body). An introduction channel 3 having a tissue introduction port (tissue introduction part) 2 for introducing a soluble salt solution) at one end, an enzyme container (enzyme introduction part) 4 containing digestive enzyme B, an introduction channel 3 and A stirring channel 5 connected to the enzyme container 4 and a cell concentrating device 6 connected downstream of the stirring channel 5 are provided.

As shown in FIG. 1, the introduction flow path 3 is provided with a pump 7 that sends the liquid A containing biological tissue in one direction.
The stirring flow path 5 includes a main pipe 9 connected to the introduction flow path 3 via a check valve 8 and disposed horizontally, and a plurality of sub pipes 10 connected to the enzyme container 4.

A filter 11 that collects biological tissue and a valve 12 that opens and closes the main pipe 9 are disposed on the main pipe 9 on the downstream side.
As shown in FIG. 1A, the sub-pipe 10 is provided with a pump 13 that sends the digestive enzyme solution B supplied from the enzyme container 4 in one direction, and a valve 14 that opens and closes the sub-pipe 10. ing.

As shown in FIG. 1B, the sub-pipe 10 has a discharge port 10 a that branches into a plurality of pipes, each of which penetrates the wall surface of the main pipe 9 and opens into the main pipe 9. .
In this embodiment, as shown in FIGS. 1B and 1C, the sub-pipe 10 is obliquely forward along the inner wall surface of the main pipe 9, that is, a predetermined crossing with the longitudinal axis of the main pipe 9. Each discharge port 10a is arranged so that the digestive enzyme solution B is ejected in the tangential direction in the cross section of the cut surface D.

  As a result, the digestive enzyme solution B sent through the sub-pipe 10 by the operation of the pump 13 is ejected toward the liquid A containing the biological tissue flowing in the main pipe 9 in a direction crossing the flow direction. The liquid A containing the biological tissue and the digestive enzyme solution B are agitated and mixed. As shown in FIG. 1C, the mixed liquid C forms a swirling flow while being flown through the main pipe 9, and is stirred more effectively.

A plurality of discharge ports 10 a are provided at intervals in the longitudinal axis direction of the main pipe 9.
Thereby, the digestive enzyme liquid B can be ejected several times, while the liquid A containing a biological tissue is made to flow through the main pipe 9.
The cell concentrating device 6 includes a centrifuge container 15 and a centrifuge 16 that rotates the bottom of the centrifuge container 15 outward in the radial direction.

The operation of the biological tissue processing apparatus 1 according to this embodiment configured as described above will be described.
In order to separate and concentrate cells from a living tissue using the living tissue processing apparatus 1 according to the present embodiment, first, the liquid A containing the living tissue collected from the living body is transferred from the tissue introduction port 2 to the introduction channel 3. By introducing the pump 7 and operating the pump 7, the pump 7 is caused to flow into the stirring flow path 5 through the check valve 8. Moreover, the digestive enzyme solution B accommodated in the enzyme container 4 is ejected into the stirring flow path 5 by opening the valve 14 of the sub-pipe 10 and operating the pump 13.

  Since the discharge port 10a of the sub-pipe 10 is arranged so as to eject the digestive enzyme solution B obliquely forward along the inner wall surface of the main pipe 9, the digestive enzyme solution B is a flow of the liquid A containing biological tissue. The liquid A is ejected and mixed in the liquid A containing the living tissue without going against the above. Thereby, the digestive enzyme liquid B can be uniformly mixed in the liquid A containing the living tissue without causing undue damage to the cells constituting the living tissue.

  In this case, according to the biological tissue processing apparatus 1 according to the present embodiment, the stirring flow path 5 is not stirred at a high flow rate that causes a turbulent flow state, but is stirred while flowing at a sufficiently low flow rate. Can do. Therefore, the mixed liquid C flows slowly over time in the stirring channel 5, and sufficient digestion can be performed even with a short channel length. Therefore, the entire apparatus can be configured compactly.

  Moreover, in this embodiment, since the discharge port 10a of the sub piping 10 formed in the main piping 9 is arranged in multiple numbers at intervals in the longitudinal direction of the main piping 9, the liquid A containing a biological tissue is A new digestive enzyme solution B is supplied each time when the digestive enzyme solution B is sprayed a plurality of times while flowing in the main pipe 9. Thereby, a biological tissue is digested efficiently.

  Furthermore, in this embodiment, since the discharge port 10a is arrange | positioned so that the digestive enzyme liquid B may be ejected toward the tangential direction of the cross section of the main piping 9, the digestive enzyme liquid ejected from the discharge port 10a B flows along the inner surface of the main pipe 9, and a swirling flow is formed in the main pipe 9. When the biological tissue has a specific gravity smaller than water, such as adipose tissue, the biological tissue tends to flow in the upper part of the flow cross section and the liquid component in the lower part of the flow cross section in the main pipe 9, but forms a swirl flow. By doing so, these are mutually wound up and down, and stirring is fully performed.

  Then, the mixed solution C of the liquid A containing the living tissue and the digestive enzyme solution B is stirred, whereby the living tissue is digested and the cells are separated. As a result, undigested living tissue and cells separated from the living tissue float in the mixed solution C.

  When the mixed liquid C flows to the downstream side of the stirring channel 5, the living tissue is sufficiently digested and can be led to the cell concentrator 6 connected downstream. Since the filter 11 is provided on the downstream side of the stirring channel 5, undigested biological tissue contained in the mixed solution C is collected by the filter 11, and cells and liquid components separated from the biological tissue are filtered. 11 is introduced into the cell concentrator 6.

  The mixed solution C guided to the cell concentrating device 6 is stored in the centrifuge container 15. Therefore, as shown in FIG. 2, the centrifuge 16 is operated and the centrifuge container 15 is rotated so that the bottom thereof is directed outward in the radial direction. The liquid component is separated from the cell group on the side away from the bottom. That is, a concentrated cell group can be obtained by sucking and removing the liquid component in the centrifuge container 15. The valves 8 and 14 shown in black in the figure indicate that they are closed.

  Thus, according to the biological tissue processing apparatus 1 which concerns on this embodiment, in the stirring flow path 5 arrange | positioned in the middle from the tissue inlet 2 to the cell concentration apparatus 6, the liquid A containing a biological tissue and digestive enzyme The living tissue is digested while the mixed liquid C with the liquid B flows at a low speed. Accordingly, the length of the main pipe 9 is shortened to make the device compact, and the digestion can be promoted by sufficiently stirring the mixed liquid C, and the living tissue remaining undigested can be reduced efficiently. Cells can be collected.

  Moreover, according to the biological tissue processing apparatus 1 which concerns on this embodiment, since it is not necessary to give a cell the flow rate which should generate | occur | produce a turbulent flow in the main piping 9, it is only necessary to flow the liquid mixture C at low speed, There is an advantage that a cell group in a healthy state can be collected by reducing damage to the cells.

In the present embodiment, a plurality of auxiliary pipes 10 are provided and their discharge ports 10a are arranged at intervals in the longitudinal direction of the main pipe 9. The digestive enzyme solution B may be ejected into the main pipe 9 on the upstream side of the path 5.
Moreover, although the discharge port 10a was arrange | positioned so that the digestive enzyme liquid B may be jetted toward diagonally forward, it replaces with this and it is turned toward the direction orthogonal to the longitudinal axis of the main piping 9, as FIG. You may arrange | position so that the digestive enzyme liquid B may be ejected. By doing in this way, since the digestive enzyme liquid B is ejected in the direction which inhibits a flow more with respect to the flow of the liquid A containing a biological tissue, the liquid mixture C can be stirred favorably.

  In addition, the digestive enzyme solution B is ejected in the tangential direction of the cross section of the main pipe 9, but instead, it is ejected in the radial direction toward the center of the main pipe 9, as shown in FIG. You may decide to make it. By doing in this way, although a swirling flow does not generate | occur | produce in the main piping 9, the flow in the main piping 9 can be disturbed and it can stir favorably.

In the present embodiment, the main pipe 9 constituting the stirring channel 5 is arranged substantially horizontally. However, the present invention is not limited to this, and the main pipe 9 is inclined in the vertical direction or the horizontal direction. You may decide to arrange.
In addition, the liquid A containing biological tissue is caused to flow through the main pipe 9 and the digestive enzyme solution B is ejected into the main pipe 9 from the discharge port 10a of the sub pipe 10. The digestive enzyme solution may be flowed, and the liquid A containing the biological tissue may be ejected from the discharge port of the auxiliary pipe 10.

  Further, as shown in FIG. 5, the main pipe 9 constituting the stirring flow path 5 is provided with a heater 17 for heating the mixed liquid C flowing in the main pipe 9 along the main pipe 9. Is preferred. By doing in this way, there exists an advantage that digestion can be accelerated | stimulated by heating the liquid mixture C to the temperature suitable for digestion.

A Liquid containing biological tissue B Digestive enzyme solution (digestive enzyme)
C mixture 1 biological tissue processing device 2 tissue introduction port (tissue introduction part)
4 Enzyme container (enzyme introduction part)
5 Stirring channel 6 Cell concentrator (cell concentrator)
9 Main piping 10 Sub piping 15 Centrifugal container (container)
16 Centrifuge (centrifuge)

Claims (7)

A tissue introduction part for introducing a liquid containing biological tissue;
An enzyme introduction part for introducing a digestive enzyme;
A stirring flow path for flowing while stirring the liquid mixture containing the biological tissue introduced from the tissue introduction part and the digestive enzyme introduced from the enzyme introduction part,
The stirring channel is connected to the main pipe for flowing the liquid containing the biological tissue introduced from the tissue introduction part, and the digestive enzyme introduced from the enzyme introduction part is connected to the wall surface of the main pipe. A biological tissue processing apparatus comprising: at least one auxiliary pipe that ejects the liquid in a direction intersecting a flow direction of the liquid containing the biological tissue.   The biological tissue processing apparatus according to claim 1, wherein the sub-pipe ejects the digestive enzyme obliquely forward with respect to the flow direction of the liquid containing the biological tissue.   The biological tissue processing apparatus according to claim 1, wherein a plurality of the auxiliary pipes are provided at intervals in the flow direction of the liquid containing the biological tissue.   The biological tissue processing apparatus according to claim 1 or 2, wherein the sub-pipe ejects the digestive enzyme in a direction along a tangent to a cross section of the main pipe in a plane intersecting a longitudinal axis of the main pipe.   The biological tissue processing apparatus according to claim 1, further comprising a cell concentration unit that is connected downstream of the stirring channel and concentrates cells in the mixed solution that has passed through the stirring channel.   The biological tissue treatment according to claim 5, wherein the cell concentrating unit is a centrifugal separation device that includes a container that contains the mixed solution that has passed through the stirring channel, and rotates the bottom of the container outward in the radial direction. apparatus.   The biological tissue processing apparatus according to claim 1, wherein the stirring channel is provided with a heater for heating a mixed liquid flowing in the stirring channel.

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