JPH05337166A - How to wash frozen stored blood - Google Patents

Abstract

(57) [Summary] [Purpose] Frozen blood cells are washed by sufficiently contacting them with a washing solution without applying any stress to the blood cell components. [Composition] A washing chamber in which a washing solution is mixed with frozen stored blood after thawing, and the resulting mixture is provided with a washing solution receiving tank and a washing solution supply port provided in the lower portion of the washing solution receiving tank and having an opening at the upper side. And a cleaning liquid is supplied from a lower portion of the cleaning chamber, and a cleaning liquid is supplied from a lower portion of the cleaning chamber to raise the inside of the cleaning chamber from below. After contacting the cleaning liquid with the blood, the resulting mixture is allowed to flow from the upper end of the opening of the cleaning chamber and discharged from the cleaning liquid receiving tank, and the blood cells and the cleaning liquid are separated by a cleaning liquid separator. How to wash stored blood. [Effect] The antifreezing agent in blood cells and the surface of blood cells is thoroughly contacted with the washing solution and washed off without applying any stress to the blood cell components, and it becomes a closed system, so that the whole system is always positive pressure. Therefore, there is no risk of invading bacteria from the outside.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for washing cryopreserved blood. More specifically, the present invention relates to a method of washing frozen stored blood, which efficiently and continuously removes a protective solution added to blood to be frozen and stored.

[0002]

2. Description of the Related Art In recent years, blood transfusion is a method of transfusing blood containing all the components obtained by donating blood, so-called whole blood, from which only the blood components required by the patient are fractionated from the blood of another person to transfuse. Have moved to. Such component transfusion has less burden on the circulatory system compared to whole blood transfusion, and can avoid and reduce immunological side effects. Since only large amounts of necessary components can be transfused at one time, even a small percentage of whole blood Ingredients that can be expected to have sufficient effects, and ingredients that are not necessary for one patient can be effectively used for other patients [Tanaka Shimizu, Comprehensive Clinical Vol. 35, No. 11 (1986)], but this component transfusion is not sufficient from the viewpoint of avoiding infection and sensitization, which are the two major side effects associated with transfusion. Today, hepatitis due to transfusion, AIDS, Infection with ATLA and the like, or sensitization in children in particular, has been regarded as a problem, and preventive measures against these are strongly desired.

Recently, as a method capable of solving such a problem, an autologous blood transfusion method of transfusing a blood sampled in advance has attracted attention. If you use your own blood, problems such as virus infection and sensitization will not occur essentially. Moreover, even an extremely rare blood type person can be prepared for an emergency.

In such an autologous blood transfusion method, the collected own blood is separated into its components and then stored frozen. Currently, blood can be stored for up to 42 days if it remains liquid, but by storing it frozen in this way, semi-permanent storage is possible, and thus a long-term safe blood transfusion system will be established.

[0005]

By the way, if blood that has been separated into its components is to be stored frozen as it is, each component such as red blood cells, white blood cells, and platelets will be destroyed.
To store frozen, it is necessary to add a protective solution to each blood component. At present, this protective solution differs depending on the research period, etc., and, of course, the target blood component, but most of them contain glycerin, dimethyl sulfoxide, etc. When returning the stored blood to the body, it is necessary to prepare a deprotection solution by thawing and then diluting and washing the stored blood.

Some examples of the dilution / washing method currently used are, for example, a concentrated red blood cell solution, a Huggins solution (glycerin 79%, butter sugar 8%, fructose 1%, EDTA-2 sodium salt). 0.3%) and slowly frozen, 50% glucose solution + 5% fructose solution first
Washed twice, washed with 5% fructose solution for the second time, further washed with 5% fructose solution for the third time, washed with physiological saline for the fourth time, and then resuspended in saline [Boston. Massachusetts General Hospital (Hagging), National Fukuoka Central Hospital (Sumida)], as well as red blood cell concentrate and oral fluid (glycerin-2).
8%, mannitol 3%, sodium chloride 0.65%)
In addition, in the case of rapid freezing, the first centrifugation was performed as it was, and the supernatant was discarded. The second was 0.45% mannitol-added 0.45.
% NaCl solution is added, and the supernatant is discarded after centrifugation. In the third and fourth times, 0.9% NaCl solution is added and the supernatant is discarded after centrifugation, and finally resuspended in physiological saline. Etc. are taken [New York Blood Center (ROE
W)].

Thus, at present, the dilution and washing of stored blood is
All of them are carried out in a batch system and also require centrifugation or centrifugation, which is not only very complicated in operation, but also the recovery rate of blood components is not sufficient. Met.

Therefore, it is an object of the present invention to provide a novel method and apparatus for cleaning frozen stored blood.

It is another object of the present invention to provide a method for washing frozen stored blood, which efficiently and continuously removes a protective solution added to blood to be frozen stored.

A further object of the present invention is to provide a method of washing frozen preserved blood which enables an aseptic washing operation.

A further object of the present invention is to provide a method for washing frozen preserved blood with a high blood component recovery rate.

[0012]

[Means for Solving the Problems] The above-mentioned various purposes are to mix a washing solution with frozen stored blood after thawing, and then mix the resulting mixture with a washing solution receiving tank and a washing solution supply port at a lower portion provided in the washing solution receiving tank. And a cleaning chamber that is open at the top,
A cleaning liquid that is supplied from a blood supply pipe of a blood cleaning device that is formed with a blood supply pipe that is opened below the cleaning chamber, and that is supplied with a cleaning liquid from a lower portion of the cleaning chamber and that rises in the cleaning chamber from below After contacting with the blood, the obtained mixture is allowed to flow from the upper end of the opening of the washing chamber, discharged from the washing liquid receiving tank, and the blood and the washing liquid are separated by a washing liquid separator. It is achieved by the cleaning method of.

The above-mentioned various purposes are to wash frozen frozen blood after thawing with a washing solution, and then supply the washing solution separator to separate blood cells and the washing solution in the washing solution separator via a tube to the washing solution separator. This can also be achieved by a method for cleaning frozen stored blood, which is characterized in that the internal pressure of the cleaning liquid separator is adjusted by changing the position above the separating bag which is in communication with the position below.

[0014]

First, the apparatus for cleaning frozen stored blood according to the present invention will be described.

As shown in FIG. 1, a cylindrical cleaning liquid receiving tank 1 is provided with a cylindrical cleaning chamber 4 having a cleaning liquid supply port 2 at a lower portion and an opening 3 at an upper portion. The supply port 2 communicates with a cleaning liquid supply pipe 5. In addition, a filter 6 such as a class filter, a glass mat, a porous membrane, or a filter paper is provided in the lower portion of the cleaning chamber 4 as required.

A blood supply pipe 8 having an opening 7 is provided below the cleaning chamber 4 in the cleaning chamber 4. When the filter 6 is a porous membrane, its pore size is preferably 0.01 to 6.5 μm, particularly preferably 0.1 to 6 μm. Further, a cleaning discharge port 9 is provided at the bottom of the cleaning liquid receiving tank 1, and a filter medium 10 such as a non-woven fabric, a woven fabric or a knitted fabric may be placed if necessary. Examples of the material of the filter medium include natural or synthetic fibers such as cotton, polyamide, and polyester, and preferably polyamide and polyamide in which cotton is mixed.

A lid 11 is attached to the upper end of the cleaning liquid receiving tank 1, and the blood supply pipe 8 is usually attached to the lid 11.

Next, a method for washing frozen stored blood using the washing apparatus according to the present invention will be described.

That is, as shown in FIG. 2, after thawing the blood in the blood cell bag 21 containing the frozen preserved blood, the clamp 23 is opened and the first washing solution is added from the first bag 22 to Shake gently for about 5 minutes to mix well. Next, open the clamp 25, add the second washing solution from the second bag 24, mix by the same operation for about 5 to 7 minutes, and wash the blood after thawing to wash glycerin, ethylene glycol, dimethyl sulfoxide, etc. The intracellular antifreezing agent (hereinafter, referred to as antifreezing agent) is washed to some extent.

Furthermore, in order to accelerate the elution of the antifreezing agent in the blood (blood cell component) and the antifreezing inhibitor on the surface of the blood cell component, the blood cell component liquid that has been subjected to the above treatment is opened by opening the clamp 26 and the cleaning device 27. Send to.

In the cleaning device 27, the clamp 28
As shown in FIG. 1, the third cleaning liquid bag 29 is supplied from the cleaning liquid supply pipe 5 through the supply port 2 and the filter 6 to the third cleaning liquid 12 from below in the cleaning chamber 4 as shown in FIG. On the other hand, the mixed liquid 13 obtained by washing the blood bag 21 to some extent is supplied from the lower end opening 7 of the blood supply pipe 8 to the vicinity of the bottom of the washing chamber 4.

The blood cell component-containing mixed liquid supplied into the third cleaning liquid is mixed and raised while coming into contact with the rising cleaning liquid in the cleaning chamber 4, and is sufficiently cleaned during that period. When it reaches the upper end of the cleaning chamber 4, it overflows from the peripheral wall portion 14 to reach the cleaning liquid receiving tank 1, is filtered by a filter medium 10 provided as necessary, and is then discharged from the cleaning liquid discharge port 9,
It is sent to the cleaning liquid separator 32.

In the washing liquid separator 32, blood cell components are separated into washing liquid, antifreezing agent and the like. Cleaning liquid separator 32
There are various types, but for example, JP-A-1-121,055 and JP-A-1
No. 121,056, a large number of porous hollow fiber membranes are arranged in a housing, and the inner space of the hollow fiber membranes is communicated with a first fluid inlet and outlet provided in the housing. And a second fluid inlet and outlet provided in the housing communicate with the space surrounded by the inner peripheral surface of the housing and the outer surface of the porous hollow fiber membrane. The first fluid circulation space is partitioned (in this example, is partitioned by a partition wall formed of a potting agent) to form a second fluid circulation space, and one or a plurality of them are used. ..

Of course, a plasma separator type washing liquid separator can also be used.

When the hollow fiber membrane type washing liquid separator as described above is used, after the second fluid inlet is closed, the blood component-containing washing liquid mixed liquid is introduced from the first fluid inlet and Cleaning liquid separator 32 tube 36
It is desirable to dispose the lower end of the separation bag 33 connected by the above-mentioned structure at substantially the same level as the upper end of the washing separator 32, as shown in FIG.

In this case, since the internal pressure in the hollow fiber membrane is high,
The blood component (blood cell component) remains in the hollow fiber membrane, and the cleaning liquid containing the antifreezing agent such as glycerin is filtered and discharged from the tube 37 and passed through the clamp 34 to the waste liquid bag 3
Recovered in 5. In the meantime, the internal pressure is adjusted by gradually lowering the position of the separation bag 33, and finally, the blood cell component in the hollow fiber membrane is collected in the separation bag 33 by moving it below the washing separator 32. ..

When the separation of the waste liquid is completed, the clamp 30 is opened, the fourth cleaning liquid is supplied from the fourth cleaning liquid bag 31 to the cleaning device 27, and the blood components remaining in the system are washed away through the tube 36. The blood is supplied to the separation bag 33 and the concentration of the blood component in the separation bag 33 is adjusted. In this case, since the internal pressure in the hollow membrane is low, the fourth cleaning liquid mainly moves to the separation bag 33 side.

In the case of using the hollow fiber membrane type washing separator as described above, it goes without saying that blood solution may be passed through the second fluid inlet to separate blood components.

The blood to be washed by the washing apparatus according to the present invention is not only whole blood but also blood cell components such as red blood cells, white blood cells and platelets, preferably blood cell components,
Especially red blood cells.

[0030]

EXAMPLES Next, the present invention will be described in more detail with reference to examples.

Example 1 Using the apparatus shown in FIGS. 1 and 2, the frozen stored red blood cells after thawing were washed.

First, a hemme bag 23 is opened in a blood cell bag 21 containing 400 ml of frozen erythrocytes using glycerin as an antifreezing agent and thawed. 100-150 ml of a first washing liquid consisting of 88 ml of an aqueous solution was added, and the mixture was shaken slowly for about 2-3 minutes and mixed well. Then, open the clamp 25 and add 5% glycerin aqueous solution (glycerin 5 ml and 5%
Second aqueous solution of trisodium citrate (95 ml)
Further, 400 ml of the washing solution of was added and mixed by the same operation for about 3 to 5 minutes to wash red blood cells.

Further, in order to promote the elution of glycerin in the red blood cells and the degreserin on the surface of the red blood cells, the red blood cell-containing liquid treated as described above was sent to the cleaning device 27 by opening the clamp 26.

In the cleaning device 27, the clamp 2
8 is opened, and the third cleaning solution bag 29 is passed through the cleaning solution supply port 5 through the cleaning solution supply port 2 and the glass filter 6 as shown in FIG. 1 to pass 1000 ml of the third cleaning solution (sodium chloride 0.8 % And glucose 0.2%) 12 was fed to the washing chamber 4 from below.

On the other hand, the mixed solution 13 obtained by cleaning the blood bag 21 to some extent was supplied from the lower end opening 7 of the blood supply tube 8 to the vicinity of the bottom of the cleaning chamber 4.

The red blood cell-containing mixed liquid supplied into the third cleaning liquid was mixed and raised while coming into contact with the rising cleaning liquid in the cleaning chamber 4, and was thoroughly cleaned during that period. When it reaches the upper end of the washing chamber 4, it overflows from the peripheral wall portion 14 to reach the washing liquid receiving tank 1 and is filtered by a filter medium 10 made of cotton, and when the disrupted substances or red blood cells are washed, residual leukocytes are removed and then the washing liquid is discharged. It was discharged from the outlet 9 and sent to the hollow fiber membrane type cleaning liquid separator 32.

In this cleaning liquid separator 32, after closing the second fluid inlet, the erythrocyte-containing cleaning liquid mixture is introduced from the first fluid inlet and the separation bag 3
While adjusting the position of 3, the internal pressure in the hollow fiber membrane is adjusted to separate the red blood cells and the glycerin-containing washing liquid, the red blood cells are collected in the separation bag 33, while the waste liquid is the waste liquid bag 3
Recovered to 5.

Then, the clemen 30 is opened, and the fourth cleaning solution (0.9% saline solution) is supplied from the fourth cleaning solution bag 31.
Was supplied to the cleaning device 27 to wash away the red blood cells remaining in the system, and the red blood cell concentration in the separation bag 33 was adjusted to 85 hematocrit (%).

The results at this time are shown in Table 1.

Example 2 The same procedure as in Example 1 was carried out except that 8% saline solution was used as the first washing solution and 1.6% saline solution was used as the second washing solution. The results shown in Table 1 were obtained.

[0041]

[Table 1]

[0042]

As described above, according to the present invention, after the thawed frozen stored blood is mixed with the washing liquid, the resulting mixture is supplied to the washing liquid receiving tank and the lower portion provided in the washing liquid receiving tank. A washing chamber with a mouth and an open top,
A cleaning liquid that is supplied from a blood supply pipe of a blood cleaning device that is formed with a blood supply pipe that is opened below the cleaning chamber, and that is supplied with a cleaning liquid from a lower portion of the cleaning chamber and that rises in the cleaning chamber from below After contacting with the blood, the resulting mixture is allowed to flow from the upper end of the opening of the washing chamber, discharged from the washing liquid receiving tank, and the blood and the washing liquid are separated by a washing liquid separator. Therefore, the antifreezing agent in blood cells and the antifreezing agent present on the surface of blood cells are sufficiently brought into contact with the washing solution and washed away without any stress on blood, particularly blood cell components.

Further, according to the present invention, a cleaning liquid receiving tank, a cleaning chamber provided with a cleaning liquid supply port in a lower portion provided in the cleaning liquid receiving tank and having an upper opening, and a lower opening of the cleaning chamber. Since the cryopreserved blood cleaning device including the blood supply tube is used, as described above, the blood cells and the cleaning solution are sufficiently contacted.

Further, the frozen stored blood after thawing is washed with a washing liquid and then supplied to a washing liquid separator, and the separation of blood cells and washing liquid in the washing liquid separator is communicated to the washing liquid separator via a tube. Since it is a method of washing frozen stored blood, which is characterized in that it is performed by adjusting the internal pressure of the washing liquid separator by changing the position above the separating bag to the position below, a closed system is not only possible. Since there is always positive pressure in the entire system (bag and circuit), there is no risk of invasion of germs from the outside.

[Brief description of drawings]

FIG. 1 is a cross-sectional view showing an embodiment of a device for cleaning frozen stored blood used in the method of the present invention.

FIG. 2 is a flow sheet showing an embodiment of a method for cleaning frozen stored blood according to the present invention.

[Explanation of symbols]

DESCRIPTION OF SYMBOLS 1 ... Cleaning liquid receiving tank, 2 ... Cleaning liquid supply port, 4 ... Cleaning chamber, 5 ... Cleaning liquid supply pipe, 6 ... Filter, 7 ... Opening part, 8 ... Blood supply pipe, 9 ... Cleaning liquid discharge port, 22, 24, 29, 31 … Cleaning solution bag, 2
7 ... Washing device, 32 ... Washing liquid separator, 33 ... Separation bag.

Claims (7)

[Claims] 1. A mixture of a frozen storage blood after thawing and a washing solution, and the resulting mixture is provided with a washing solution receiving tank, and a washing solution supply port provided in a lower portion provided in the washing solution receiving tank and having an upper opening. The cleaning chamber is supplied from a blood supply pipe of a blood cleaning apparatus including a cleaning chamber and a blood supply pipe opened below the cleaning chamber, and a cleaning liquid is supplied from a lower portion of the cleaning chamber, and the cleaning chamber is supplied from below. After contacting the rising washing liquid with the blood, the resulting mixture is allowed to flow further from the upper end of the opening of the washing chamber, discharged from the washing liquid receiving tank, and the blood cells and the washing liquid are separated by a washing liquid separator. Method for washing frozen stored blood. 2. The method for washing frozen stored blood according to claim 1, wherein the mixture overflowing the upper end of the opening of the washing chamber passes through the filter medium and is discharged and collected. 3. The method for cleaning frozen stored blood according to claim 1, wherein the stored blood is a blood cell component. 4. The method for washing frozen stored blood according to claim 3, wherein the blood cell component is red blood cells. 5. The frozen stored blood after thawing is washed with a washing solution and then supplied to a washing solution separator, and the separation of blood cells and washing solution in the washing solution separator is communicated with the washing solution separator via a tube. A method for washing frozen preserved blood, which is performed by adjusting the internal pressure of the washing liquid separator by changing the position above the existing separation bag to the position below. 6. The method for washing frozen stored blood according to claim 5, wherein the washing liquid separator is a hollow fiber membrane type separator. 7. The cleaning is performed by a cleaning liquid receiving tank, a cleaning chamber provided in the cleaning liquid receiving tank at a lower portion with a cleaning liquid supply port and having an upper opening, and a lower opening of the cleaning chamber. 7. A cleaning device comprising a blood supply pipe, wherein blood is supplied from the blood supply pipe and a cleaning liquid is supplied from a lower portion of the cleaning chamber to bring blood cells into contact with the cleaning liquid. Method for washing frozen stored blood.