JPH0956813A - Red cell-removing system - Google Patents

Abstract

PROBLEM TO BE SOLVED: To make it possible to easily and rapidly make the operation to remove red cells from a raw material cell liquid at a low cost without using a centrifugal separator and special reagent and to make aseptic operation. SOLUTION: This red cell-removing system includes a recovered liquid storage section 4 which is respectively connected with a means 1 for storing the drawn raw material cell liquid, a filter means 2 for allowing the passage of the red cells and capturing the white cells and a means 3 for storing the red cell-contg. liquid flowing out of this filter means 3 respectively in series via first and second tubes and is connected to a third tube 8 branched from a second tube 7 downstream of the filter means and a white blood recovery section 5 which is connected to a fourth tube 9 branched from a first tube 6 upstream of the filter means. Further, the system has the means for passing the liquid only toward the filter means 2 from a recovered liquid storage section 4 at the branching point of the second and third tubes and the means for passing the liquid only toward a white cell recovery section 4 from the filter means 2 at the branching point of the first and fourth tubes.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a system for removing red blood cells from a cell population such as bone marrow or peripheral blood containing red blood cells and hematopoietic stem cells and / or hematopoietic progenitor cells.

[0002]

2. Description of the Related Art At present, a bone marrow transplantation method or a peripheral blood stem cell transplantation therapy using peripheral blood is performed for hematopoietic disorders, which are main side effects in chemotherapy of hematopoietic tumors such as leukemia and solid cancer. As is well known, when performing allogeneic transplantation in these transplantation therapies, the matching of HLA (white blood cell type) between the donor and the recipient (patient) is the most important. Has been resolved by removing red blood cells from donor bone marrow or peripheral blood. Conventionally, a method of separating a white blood cell fraction (more specifically, mononuclear cell fraction) containing hematopoietic stem cells and / or hematopoietic progenitor cells required for transplantation and a red blood cell fraction by a centrifuge as a means for removing red blood cells Is being done.
When it is desired to separate erythrocytes and mononuclear cells particularly effectively, a centrifugation method using a specific gravity liquid called density gradient centrifugation (for example, Ficoll manufactured by Pharmacia) is carried out. The operation process involves adding a cell suspension to the specific gravity liquid. It is complicated and requires skill such as never disturbing the liquid surface when stacking layers. Moreover, the cost is high because a specific gravity liquid is used. JP 61-8455
Although many attempts have been made to solve the complexity of operations in Japanese Patent Laid-Open No. 7-134564, Japanese Patent Laid-Open No. 2-134564, etc., there is still no technique for removing erythrocytes from bone marrow in a simple, low-cost and short time.

[0003]

SUMMARY OF THE INVENTION It is an object of the present invention to provide a red blood cell removal system which is simple, can be operated at low cost in a short time, and can be operated aseptically without using a centrifuge or a special reagent. It is in.

[0004]

FIG. 1 is a schematic diagram of the red blood cell removal system of the present invention. According to the present invention, the means 1 for storing the collected raw material cell fluid, the filter means 2 for passing the red blood cells and capturing the white blood cells, and the means 3 for storing the red blood cell-containing liquid flowing out from the filter means 2 are respectively the first. And connected in series via the second tube,
Recovery liquid reservoir 4 connected to a third tube 8 branched from the second tube 7 downstream of the filter means, and white blood cells connected to a fourth tube 9 branched from the first tube 6 upstream of the filter means. At the branch point of the second and third tubes, a means for circulating the collected liquid storage section 4 only in the direction of the filter means 2 is provided at the branch point of the first and fourth tubes. The point relates to an erythrocyte removal system characterized in that it has means for allowing the filter means 2 to flow only in the direction of the white blood cell recovery liquid part 5. Furthermore, it relates to a red blood cell removal system having means for rinsing the filter means before leukocyte recovery. Examples of the filter means used in the present invention include, but are not limited to, nylon wool, polyester non-woven fabric, hydroxyapatite beads, sponge-like polymer compounds, and the like. I wish I had it. However, if it is desired to remove platelets along with erythrocytes in order to improve the separation efficiency in the field of cell separation for the purpose of removing cells causing complications in allogeneic bone marrow transplantation or in removing cancer cells in autologous bone marrow transplantation, The surface charge is adjusted by a coating treatment or the like for passing platelets through the filter means of the invention. Examples of means for unidirectionally circulating the second and third tubes and the first and fourth tube branch points referred to in the present invention include three-way stopcocks, dual check valves, clamps, etc., but three-way stopcocks are used. Is desirable. Furthermore, in the collected solution storage section 4,
The filter means contains a recovery liquid for recovering the captured white blood cells. As the recovery liquid, physiological saline, HB
A buffer such as SS (Hanks's solution) or D-PBS (Dulbecco's phosphate buffer) to which a protein such as human serum albumin or an anticoagulant is added as needed is used. Further, the raw material cell liquid used in the present invention is a liquid containing red blood cells, hematopoietic stem cells and / or hematopoietic progenitor cells, and examples thereof include bone marrow, peripheral blood, cord blood, and those roughly separated by a centrifuge. It is considered that the presence / absence of erythrocyte removal will be further enhanced with the development of the technique of culturing and transplanting a small amount of hematopoietic stem cells using the stem cell expansion method in the future.

[0005]

Embodiment 1 One example of an embodiment of the present invention will be described with reference to the drawings. 1 is a container containing raw material cells (usually a blood bag)
And is connected to the first tube 6. Reference numeral 4 is a bag containing the recovery liquid, from which the recovery liquid is supplied to the leukocyte removal filter 2. The third tube 8 is a tube for supplying the recovery liquid to the filter means 2 and is connected to the second tube 7. A clamp 14 is provided in the middle of the third tube, and a three-way stopcock 12 is provided at a branch point between the third tube 8 and the second tube 7. The tube 10 is a tube for introducing the rinse liquid into the filter means 2, and is branched from the first tube 6 by a Y-shaped tube or the like. There is a clamp 13 in the middle of the tube 10. First
Clamp 16 on tube 6 and trap with mesh 1
Those with 5 are desirable. The fourth tube 9 is for introducing the recovered cell liquid into the white blood cell recovery unit 5. First
There is a three-way stopcock 11 at the branch point between the tube 6 and the fourth tube 9. The filter means 2 is a filter that passes red blood cells (and platelets when it is desired to remove platelets) but traps white blood cells. The bag used for each of the means 1, 3, 4, and 5 can be connected to the tube by a spike or an aseptic connector. Next, the usage method will be described. First, bone marrow, peripheral blood buffy coat, umbilical cord blood and the like collected by a conventional method are put in the blood bag 1 in advance. At this time, the clamp 13 is in the “closed” state, the three-way stopcock 11 is in the “closed” direction of the tube 9, and the three-way stopcock 12 is in the “closed” direction of the tube 8. The cell fluid in the blood bag 1 is
It is introduced into the leukocyte removal filter 2 through the tube 6. There, the white blood cells are captured, the red blood cells and the platelets pass, and are collected in the drainage bag 3 from the second tube 7. In addition, the clamp 13 is opened to wash away some red blood cells remaining in the filter 2, and the collection solution is collected from the collection bag 4 to the tube 10, the first tube 6, the leukocyte removal filter 2, and the second tube in this order. To the drainage bag 3. Next, the clamp 13 is closed, the direction of the second tube 7 (drainage bag side) of the three-way stopcock 12 is “closed”, and the direction of the first tube 6 of the three-way stopcock 11 (blood bag side) is changed. Make it "closed". Then, by strongly crushing the recovery solution bag 4, the recovery solution is introduced into the leukocyte removal filter 2, the leukocytes captured by the filter are pushed out, and the leukocyte recovery bag 5 is caused to flow through the fourth tube 9. The cell fluid collected in the leukocyte collection bag 5 is infused into the patient as it is or after the volume is reduced by a concentration operation, or is cryopreserved by performing necessary work such as addition of a frost damage protector and thawed after mass therapy. It is used to infuse patients.

[0006]

According to the present invention, it is possible to provide a red blood cell removal system which does not use a centrifuge or a special reagent, is simple, can be operated at low cost in a short time, and can be operated aseptically.

[Brief description of drawings]

FIG. 1 is a schematic view of a “red blood cell removal system” of the present invention.

[Explanation of symbols]

 1 Blood Bag 2 Leukocyte Removal Filter 3 Drainage Bag 4 Collection Fluid Bag 5 Leukocyte Collection Bag 6 First Tube 7 Second Tube 8 Third Tube 9 Fourth Tube 10 Filter Rinse Tube 11, 12 Three-way Stopcock 13 Clamp 14 Clamp 15 Mesh trap 16 Clamp

Claims (2)

[Claims] 1. A means 1 for storing the collected raw material cell fluid.
And filter means 2 for passing red blood cells and capturing white blood cells
And a means 3 for storing the erythrocyte-containing liquid flowing out from the filter means 2 are connected in series via first and second tubes, respectively, and branched from the second tube 7 downstream of the filter means. The recovered liquid storage part 4 connected to the third tube 8, and the first upstream of the filter means.
A leukocyte recovery part 5 connected to a fourth tube 9 branched from the second tube 6 and flowing only from the recovery liquid storage part 4 to the filter means 2 at the branch point of the second and third tubes. An erythrocyte removal system characterized in that it has means for urging it at a branch point between the first and fourth tubes so as to circulate it only from the filter means 2 toward the leukocyte recovery part 5. 2. The rinsing tube 1 of the filter means 2 which is connected to the first tube 6 from the recovered liquid storage section 4.
The red blood cell removal system according to claim 1, having 0.