JP2001194365A - Container and method for measuring cell function - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a cell function measuring container to easily and efficiently prepare a control sample and a sample to be measured from a blood sample in one single container. SOLUTION: In the cell function measuring container, a second tubular container 3 is disposed inside a first tubular container 2, upper end openings 2a, 3a of the first and second tubular containers 2, 3 are closed by a plug 4, a communication mechanism is constituted by the second tubular container 3 and the plug body 4, the first and second tubular containers 2, 3 are communicated with each other making use of the communication mechanism after the blood sample 8 is introduced in the second tubular container 3, and inverted in the condition. A part of the blood sample flows into a space between the first and second tubular containers 2, 3 from the second tubular container 3, and a stimulant substance 5 to induce the production of the physiologic active substance in the blood sample reacts with the blood sample in the space.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a cell function measuring container capable of performing steps from the collection of a blood sample to the measurement of the result obtained by reacting the blood sample with a reagent or the like, and to the use of the cell function measuring container. Cell function measurement method.

[0002]

2. Description of the Related Art Leukocytes such as granulocytes, monocytes, macrophages, and lymphocytes play various roles in various biological defense reactions such as an inflammatory response and an immune response in blood or in each organ or organ. These cells play an important role in various disease states such as infectious diseases; inflammatory diseases such as hepatitis and nephritis; immunological and allergic diseases such as rheumatoid arthritis and asthma; and cancers. Is known to suppress or enhance the function of cells.

[0003] In addition, anti-inflammatory drugs,
Various drugs such as immunosuppressants, immunopotentiators, and anticancer drugs have been used, and it is also known that the function of these cells is suppressed or enhanced in such cases. Therefore, it is important to study the functions of these cells to determine the treatment guidelines and determine the dosage and timing of drugs by understanding the pathology of various diseases and the effects or side effects of drugs. is there.

Conventionally, in an examination room or an examination center of a hospital,
For the reasons described above, a granulocyte phagocytic function test, a granulocyte bactericidal ability (active oxygen-producing ability) test, a lymphocyte blastogenesis test and the like have been performed to measure such cell functions.
In recent years, surface antigen tests using a flow cytometer and a fluorescently labeled monoclonal antibody against various immunocompetent cell surface antigens have been performed. However, the conventional test methods require special techniques such as cell separation, cell culture, and microscopic measurement, and the measurement requires a long time, and requires expensive equipment such as an RI facility and a flow cytometer.

[0005] In addition, monocytes in blood and macrophages which have been differentiated and matured from monocytes are involved in elimination of foreign substances through phagocytosis, establishment of immunity through antigen presentation, and cytokines. By secreting various physiologically active substances such as prostaglandins, they have a wide variety of functions such as regulating inflammatory and immune responses. Therefore, monocytes and macrophages, like granulocytes and lymphocytes, play an important role in various disease states,
It is very important to examine the function of these cells.
In particular, in infectious diseases and the like, monocytes and macrophages differ from granulocytes and lymphocytes in that the number of cells does not fluctuate so much, and their function is mainly amplified, and it is more important to measure changes in cell functions ( "Macrophage", Toru Tokunaga, Kodansha Scientific 1986, first edition).

Tumor necrosis factor α (hereinafter, referred to as TNF-α), interleukin-1β (hereinafter, referred to as IL-1β), interleukin-6 (hereinafter, referred to as IL-6)
Is called a monokine, which is produced by blood cells mainly by leukocytes including monocytes and macrophages, and is a cytokine involved in various inflammatory and immune responses.

Various methods have been reported so far for examining the above-mentioned cytokine production function from blood or leukocytes separated from blood. For example, JP-A-2-196951 and JP-A-3-285692 describe that T is produced by reacting blood with lipopolysaccharide (LPS) or lectin.
Methods for measuring cytokines such as NF-α and IL-1β have been disclosed. Also, JP-A-6-209998 and JP-A-7-67955 disclose that a polymer material having a specific surface roughness or a polymer material having a specific chemical structure is brought into contact with blood to obtain TNF-α. A method for inducing the production of is disclosed. Further, Japanese Unexamined Patent Publication No. 7-299732
Japanese Patent Application Laid-Open No. 7-151752 discloses that a polymer material having a specific surface roughness is brought into contact with blood,
A biological reaction test method for measuring the production amount of -α or IL-1β is disclosed. Also, Japanese Patent Application Laid-Open No. 7-500905 discloses TNF-α and IL-1 from human peripheral blood leukocytes.
There is disclosed a method for measuring the immunoreactivity of a test substance by measuring the amount of induction of production of a cytokine such as β.

However, the following problems have been encountered in the cell function measuring method conventionally used in hospital examination rooms and examination centers and the method disclosed in the above-mentioned patent publication. In other words, in all of these tests, after collecting blood from a subject with a syringe, blood is transferred to various reaction vessels by means of pipetting or the like, cell separation for separating leukocytes, etc., and function Special operations such as cell culture for measurement were required. Therefore, there is a risk that the test worker may come into contact with blood and be infected with various infectious diseases such as hepatitis and AIDS. Also, during these operations, various germs and dust may be mixed into the blood sample, and cells in the blood are unnecessarily stimulated by these contaminants or physical stimulation by the operation, and the measurement results Could be adversely affected.

In a conventional method of collecting blood and measuring the production of cytokines, particularly TNF-α and IL-1β from leukocytes, in a specific reaction vessel, a blood sampling device such as a syringe or a reaction device is used. Originally, the container sometimes contained endotoxin such as LPS derived from Gram-negative bacteria. Endotoxin is found in trace amounts from leukocytes
To induce the production of NF-α and IL-1β, for example, when a small amount of endotoxin is mixed into the blood collection device or the reaction container due to contamination of a small amount of dust in a manufacturing process or contamination from washing water used. It was impossible to obtain reliable measurement results.

[0010] Because of the above-mentioned problems, there is a demand for a method of measuring cell functions that is simpler, less dangerous, and more accurate than before. One example of a cell function measurement container for collecting a blood sample and measuring cell function is described in WO98 / 07031.
No., published in Japanese Unexamined Patent Publication No. Here, biologically active substances produced by blood cells, such as cytokines, TNF-α and I
A kit for measuring cell function for measuring L-1β or the like is disclosed. In this kit for cell measurement, a material for inducing the production of the above-mentioned physiologically active substance, for example, a control first cell function measuring container in which the endotoxin content is an amount that does not produce a physiologically active substance from blood cells, A second cell function measurement container accommodated therein.

Therefore, blood must be collected, reacted, and measured in both the first cell function measurement container and the second cell function measurement container. Therefore,
The operation was complicated.

Conventionally, when analyzing a blood sample,
Generally, a blood collection tube is used.A blood sample is collected by a blood collection tube, and after obtaining serum or plasma, a blood sample such as serum or plasma is dispensed into a reaction container for measurement and analyzed. A method of analyzing in an apparatus or the like is generally used. Therefore, a blood collection tube is used for collecting a blood sample, and a tubular container or the like for measurement is used to supply the blood sample to the analyzer. Alternatively, as described above, when a blood sample is reacted with another substance and the result of the reaction is measured, the plasma or serum is taken out from the blood collection tube, and the serum or plasma is placed in a reaction container containing the above-mentioned reaction substance. And the like had to be dispensed and reacted.

[0013]

SUMMARY OF THE INVENTION It is an object of the present invention to overcome the above-mentioned disadvantages of the prior art, to prepare control and measurement samples in one container, and to obtain a reaction result from the collection of a blood sample. It is an object of the present invention to provide a cell function measurement container and a cell function measurement method that enable a series of steps up to a single step to be performed in a single container.

[0014]

According to a broad aspect of the present invention, there is provided a cell function measurement container for measuring a cell function of a blood sample, wherein the first container has an opening at an upper end and a bottom. A tubular container inserted into the first tubular container, having an opening at the upper end and having a bottom, and a second container for collecting blood.
Tubular container, a plug closing the first and second tubular containers, and a gap between the first and second tubular containers, and reacts with the blood sample to produce a physiologically active substance. A stimulating substance that induces production, and further comprising a communication mechanism that enables dispensing a blood sample guided into the second tubular container into a gap between the first and second tubular containers. A cell function measurement container is provided.

According to another broad aspect of the present invention, there is provided a cell function measuring method using the cell function measuring container according to the present invention. The method for measuring cell function comprises the steps of collecting a blood sample in a second tubular container, and using at least a part of the blood sample collected in the second tubular container by using the communication mechanism. The blood sample into the first tubular container and the second tubular container by flowing into the gap between the two tubular containers, and the blood sample in the gap between the first tubular container and the second tubular container. With a stimulating substance.

[0016] In a specific aspect of the cell function measurement container according to the present invention, the communication mechanism is formed so as to extend downward from an upper open end on the inner surface of the plug and the second tubular container. A large-diameter insertion portion, which is formed of a groove, wherein the plug body liquid-tightly closes an upper end opening of the first tubular container;
A small-diameter insertion portion connected to the lower end of the large-diameter insertion portion and for closing the upper end opening of the second tubular container in a liquid-tight manner, and extending vertically in the outer peripheral surface of the small-diameter insertion portion. A second groove is formed, and a third groove is formed on a lower surface of the large-diameter insertion portion so as to communicate a gap between the first and second tubular containers and the second groove. The lower end of the first groove is positioned lower than the lower end of the small-diameter insertion portion when the small-diameter insertion portion is inserted into the second tubular container.

In a specific aspect of the cell function measuring method according to the present invention, the communication mechanism includes a plug and a first groove formed on the inner surface of the second tubular container so as to extend downward from the upper open end. A large-diameter insertion portion that closes an upper end opening of the first tubular container in a liquid-tight manner, and a lower end of the large-diameter insertion portion; A small-diameter insertion portion for closing an upper end opening of the tubular container in a liquid-tight manner, a second groove extending vertically in an outer peripheral surface of the small-diameter insertion portion is formed, and an upper end of the second groove is formed. Is connected to a third groove formed on the lower surface of the large-diameter insertion portion so as to communicate the gap between the first and second tubular containers and the second groove, and the lower end of the first groove is When the small-diameter insertion portion is inserted into the second tubular container, the small-diameter insertion portion is located below the lower end of the small-diameter insertion portion.
Then, a part of the blood sample is transferred to the second using the communication mechanism.
Flowing into the gap between the first and second tubular containers from the inside of the tubular container, rotating the plug, and inserting the second groove of the plug into the second groove.
Overlapping with the first groove of the tubular container, thereby communicating between the first and second tubular containers, and turning the first and second tubular containers closed by the stopper upside down. Flowing the blood sample from the second tubular container into the first tubular container; and inverting the cell function measuring container upside down again to the first tubular container and the second tubular container. Separating the sample and reacting the blood sample with the stimulant in the gap between the first tubular container and the second tubular container.

In a specific aspect of the cell function measuring container according to the present invention, the communication mechanism is constituted by the plug and a cutout formed at an upper opening end of the second tubular container. The plug body is connected to a large-diameter insertion portion that closes an upper end opening of the first tubular container in a liquid-tight manner, and is connected to a lower end of the large-diameter insertion portion, and the upper end opening of the second tubular container is A small-diameter insertion portion for closing in a liquid-tight manner, and a groove extending vertically and continuous downward is formed on the outer peripheral surface of the small-diameter insertion portion.

In a specific aspect of the cell function measuring method according to the present invention, the communication mechanism includes a stopper and a notch formed at an upper open end of the second tubular container. The body is connected to a large-diameter insertion portion that closes the upper end opening of the first tubular container in a liquid-tight manner, and is connected to a lower end of the large-diameter insertion portion.
And a small-diameter insertion portion for closing the upper end opening of the second tubular container in a liquid-tight manner, and a groove extending vertically and continuing downward is formed on the outer peripheral surface of the small-diameter insertion portion. . Then, the step of causing a part of the blood sample to flow into the gap between the first and second tubular containers from inside the second tubular container by using the communication mechanism includes rotating the stopper and closing the groove of the stopper. A step of overlapping with the notch of the second tubular container, thereby communicating between the first and second tubular containers, and inverting the first and second tubular containers closed by the plug body. Flowing the blood sample from the second tubular container into the first tubular container; and inverting the inverted cell function measuring container upside down again into the first tubular container and the second tubular container. Separating the blood sample and reacting the blood sample with a stimulant in a gap between the first tubular container and the second tubular container.

In a specific aspect of the cell function measurement container according to the present invention, endotoxin is used as the stimulating substance. In a further specific aspect, the inner surface of the first tubular container and / or the outer surface of the second tubular container are provided. Whether the stimulant is physically or chemically immobilized,
The stimulating substance is physically or chemically immobilized on a carrier having a size that cannot move between the tubular container and the second tubular container.

[0021]

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, an embodiment of a cell function measuring container and a cell function measuring method according to the present invention will be described with reference to the drawings.

FIG. 1A is a longitudinal sectional view showing a cell function measuring container according to one embodiment of the present invention, and FIG. 3 is a perspective view showing an appearance thereof. FIG. 4A is a perspective view showing the plug body 4 and the second tubular container 3 separately, and FIG.
Is a perspective view of the plug body viewed from below, and FIG. 5 is a perspective view showing the first tubular container 2.

The cell function measuring container 1 includes a first tubular container 2
And a second tubular container 3 inserted into the first tubular container 2 and a plug 4 made of an elastic material. The first tubular container 2 has an opening 2a at the upper end. The first tubular container 2 has a bottom and a substantially cylindrical shape. The first tubular container 2 can be made of an appropriate material such as a synthetic resin or glass. However, in consideration of the upside down operation described below, the first tubular container 2 is made of a synthetic resin in order to prevent breakage. Is preferred. In addition, the first tubular container 2 is desirably transparent so that the reaction result can be visually confirmed from the outside or measured using a spectrophotometer or the like. Preferably, polypropylene having excellent transparency and excellent water vapor barrier properties is used.

The second tubular container 3 is formed of a substantially cylindrical bottomed tubular body having a smaller diameter than the first tubular container 2. The second tubular container 3 has an opening 3a at the upper end. The second tubular container 3 can be made of an appropriate material such as glass or a synthetic resin, but is preferably made of a synthetic resin in order to prevent breakage. More preferably, polyethylene terephthalate having excellent gas barrier properties is used.

The second tubular container 3 is desirably transparent so that the internal state can be observed from the outside. The plug 4 is made of an elastic material such as synthetic rubber or natural rubber, and has a handle 4a at the upper end. The handle 4a is configured to have a diameter larger than the outer diameter of the first tubular container 2 so that the operation of rotating and pulling out the plug 4 by hand can be easily performed. It is configured. A large-diameter insertion portion 4b is connected to the lower end of the handle 4a.
Is connected to a small-diameter insertion portion 4c. The large-diameter insertion portion 4b is configured to have a diameter such that it can be pressed into the opening 2a of the first tubular container 2, and the small-diameter insertion portion 4b
c is configured to be able to be pressed into the opening 3a of the second tubular container 3. That is, the plug 4 is closed so that the openings 2a and 3a of the first tubular container 2 and the second tubular container 3 are closed.
Is attached.

As is apparent from FIG. 4A, a first groove 3b is formed on the inner surface of the second tubular container 3 so as to extend downward from the opening 3a. On the other hand, a second groove 4d extending in the vertical direction is formed in the small-diameter insertion portion 4c of the plug 4. The lower end of the second groove 4d is configured not to reach the lower end of the small-diameter insertion portion 4c. Also,
As shown in FIG. 4B, the second tubular container 3 is provided on the lower surface of the large-diameter insertion portion 4b so as to be continuous with the upper end of the second groove 4d.
A third groove 4e extending outward from the outer peripheral surface of is formed. In FIG. 1, the second groove 4d is indicated by a broken line. Regarding the second groove 4d, the small-diameter insertion portion 4c
Is deformed because it is pressed against the inner peripheral surface of the tubular container 3. In the state shown in FIG. 1, the first groove 3b is not shown because it is arranged at a position that does not overlap with the second groove 4d in the circumferential direction.

Therefore, in the initial state shown in FIG. 1, the inside of the second tubular container 3 and the inside of the first tubular container 2 are liquid-tightly shut off. The lower end of the first groove 3b is located below the lower end of the small-diameter insertion portion 4c in the state shown in FIG. 1, that is, in a state where the small-diameter insertion portion 4c is inserted into the second tubular container 3. ing.

In the present embodiment, the communication mechanism is composed of the first groove 3b and the plug 4, but in the present invention, the blood sample guided into the first tubular container is An appropriate communication mechanism that enables dispensing into the gap between the first and second tubular containers can be used, and is not particularly limited to the structure of the present embodiment.

For example, as shown in FIG. 6, the communicating mechanism comprises a notch 3c formed at the upper open end of the second tubular container 3 and an outer peripheral surface of the small-diameter insertion portion 4c of the plug 4 vertically. Groove 4 extending in the vertical direction and continuing downward
f.

In the cell function measuring container 1 of the present embodiment, as shown in FIG. 1, a first tubular container 2 and a second tubular container 3 are provided.
The stimulant 5 is accommodated in the gap between them, that is, in the first tubular container 2. As the stimulating substance 5, an appropriate stimulating substance that reacts with a blood sample to induce the production of the physiologically active substance is used depending on the type of the physiologically active substance to be measured. In the present embodiment, endotoxin is stored as the stimulant 5.

In the second tubular container 3, a blood anticoagulant 6 is stored. As the blood anticoagulant 6, heparin, a citrate compound, an oxalate compound, or the like can be used. Particularly, sodium heparin is preferable because it does not inhibit a biological reaction of cells. Further, a blood coagulant such as thrombin may be contained as the stimulating substance 5 in the first tubular container 2.

Next, a method for measuring cell functions using endotoxin as the stimulant 5 to induce cytokine production and measure cytokine will be described with reference to FIGS.

First, as shown in FIG.
Is pierced into the stopper 4, and the blood sample 8 is introduced into the second tubular container 3. In this case, when the pressure in the second tubular container 3 is reduced, the blood sample 8 can be introduced into the second tubular container 3 more quickly. The pressure inside the first tubular container 2 as well as inside the second tubular container 3 may be reduced.

The blood sample 8 is collected in the second tubular container 3 as described above. Since blood anticoagulant 6 is contained in second tubular container 3, blood sample 8 does not coagulate.

Next, the stopper 4 is rotated around the axial direction, and the first groove 3b provided in the second tubular container 3 and the second groove 4d provided in the stopper 4 are moved. Overlap.
Thereafter, the entire cell function measurement container 1 is turned upside down as shown in FIG. In this case, the first groove 3b and the second groove 4d face each other to form a flow path. Since this flow path is connected to the third groove 4e, the second tubular container 3
The inside and the inside of the first tubular container 2 communicate with each other. Therefore, the blood sample flows from the inside of the second tubular container 3 to the first tubular container 2 through the above-mentioned flow path. As a result, as shown in FIG. 2A, the liquid level of the blood sample 4 is the same in the first tubular container 2 and the second tubular container 3.

Next, the cell function measuring container 1 is turned upside down again (FIG. 2B). As a result, the blood sample 8 remains in the first tubular container 2, and the blood sample 8 also exists in the gap between the first tubular container 2 and the second tubular container 3. However, blood sample 8 stored between first tubular container 2 and second tubular container 3 is brought into contact with stimulating substance 5 described above. Therefore, in the blood sample, cytokine production is induced by stimulant 5, ie, endotoxin.

Therefore, a blood sample as a control can be collected in the second tubular container 3, and a sample to be measured is collected in the first tubular container 2. As described above,
By using the cell function measurement container 1 of the present embodiment, a single cell function measurement container can efficiently carry out the process from collection of a blood sample to preparation of cell functions.

In the above embodiment, a method of measuring cytokine as a physiologically active substance using endotoxin as the stimulant 5 has been described as an example. However, the cell function measuring container according to the present invention is applicable to various cell function measuring methods. Can be used.

That is, the physiologically active substance as the substance to be measured is not limited to cytokines.
Examples include arachidonic acid metabolites such as prostaglandins, reactive oxygen species, soluble adhesion factors, soluble receptors, and intragranular enzymes.

Further, the stimulating substance for inducing the production of the physiologically active substance can be appropriately selected according to the kind of the physiologically active substance. Examples of such stimulants include, besides endotoxin, various microorganisms such as BCG killed bacteria and Corynebacterium; synthetic lipid A; pyran copolymer;
Lectin (phytohemagglutinin, concanavalin A,
OK432 (picibanil); PSK (crestin); lentinan; zymosan; LPS (lipopolysaccharide); calcium ionophore; phorbol ester; immunoglobulin-immobilized carrier; formyl methionyl leucylphenylalanine (F)
Formyl peptides such as MLP); various cytokines and the like.

Also, asthma, hay fever, allergic rhinitis,
Specific antigens (eg, house dust; mite antigen; various pollen antigens such as ragweed pollen extract, cedar pollen extract, and rice extract) used for allergen tests such as atopic dermatitis, gastrointestinal allergy, and parasite allergy; Fungal antigens, parasite antigens such as Ascaris extract; food antigens such as ovalbumin, wheat, soybean, shrimp, crab, meat;
Bee venom) is also used.

In addition, the above-mentioned materials for inducing the production of a physiologically active substance are immobilized on various natural and synthetic polymer materials by a known immobilization method (covalent bonding method, physical adsorption method, etc.). No.

The shape of the material for inducing the production of the physiologically active substance is not particularly limited, and examples thereof include a liquid and a particle. Further, it may be immobilized on a carrier. In the case of the liquid, the material that induces the production of the physiologically active substance is usually used as a diluent, for example, a buffer such as a phosphate buffer or a Hanks buffer, MEM, or RPMI-
The medium is diluted with a normal medium such as 1640, physiological saline (for example, Otsuka Pharmaceutical Co., Ltd.), water for injection (for example, Otsuka Pharmaceutical Co., Ltd.) and used.

The state of the material for inducing the production of the physiologically active substance in the reaction vessel may be in a solid state or a liquid state. When the material that induces the production of a physiologically active substance is a water-soluble material, it may be made into a powder form after being applied or added to the inner wall surface of the container. For example, when a material that induces the production of a physiologically active substance is diluted with water for injection, it is preferable that the material that induces the production of a physiologically active substance is put into a container and then dried. When the material that induces the production of the physiologically active substance is a water-insoluble material, if air bubbles remain on the surface of the material that induces the production of the physiologically active substance, excessive hemolysis may occur when the material is brought into contact with blood by inversion mixing or the like. Therefore, the water-insoluble material is preferably immersed in, for example, the above-mentioned diluent.

When a material that induces the production of a physiologically active substance is immobilized on a carrier and used, and when the carrier to be immobilized is a water-soluble material, it is applied or added to the inner wall of the container and then powdered. Is also good. When the carrier is a water-insoluble material, if air bubbles remain on the surface of the material, excessive hemolysis may be caused and the measurement system may be affected. Therefore, it is preferable that the carrier is immersed in the above-described diluent, for example.

The stimulating substance is more preferably physically or chemically immobilized on the inner surface of the first tubular container or the outer surface of the second tubular container or on both surfaces thereof.
The method for physically immobilizing the stimulant on the surface of the container is not particularly limited. For example, a method in which endotoxin is dissolved in a carbonate buffer (pH 9.5) at a concentration of 10 mg / mL is immobilized. Add 1 mL to the surface of the container
After leaving still at 37 ° C. for about 18 hours, a method of washing with endotoxin-free water and the like can be mentioned.

Further, the stimulating substance is physically or chemically immobilized on a carrier having a size that cannot move between the first tubular container and the second tubular container by the communication mechanism. Is also good. As the carrier, a carrier larger than the size of the groove or the notch constituting the communication mechanism is used, and examples of the material include a synthetic resin. The method of physically immobilizing the stimulant on such a carrier is not particularly limited. For example, endotoxin is added to a carbonate buffer (pH 9.5) at a concentration of 10 mg / day.
A method in which 10 mL dissolved in a concentration of mL is mixed with 1 g of polystyrene beads having a diameter of about 2 mm, stirred at 37 ° C. for about 18 hours, and washed with endotoxin-free water, or the like can be mentioned.

The amount of the material for inducing the production of a physiologically active substance to be added to the reaction vessel is preferably set to an optimum concentration depending on the type of the material for inducing the production of the physiologically active substance. The first tubular container 2 may contain a measuring reagent for measuring the physiologically active substance. Examples of such a measuring reagent include a monoclonal antibody and a polyclonal antibody against the physiologically active substance. And enzyme immunoassay reagents utilizing enzymes such as peroxidase, alkaline phosphatase and the like, and chromogenic substrates of each enzyme.

In the above embodiment, the stimulus substance 5 is stored in the first tubular container 2, but in addition to the stimulus substance 5, a measurement reagent which reacts with a physiologically active substance produced by the stimulus substance is stored. Alternatively, the measurement reagent may be used to measure the concentration of a physiologically active substance.

[0050]

In the cell function measuring container according to the present invention, a second tubular container for collecting blood is inserted into the first tubular container, and a gap between the first and second tubular containers is provided. And a part of the blood sample guided into the second tubular container is moved to the gap between the first and second tubular containers by utilizing the communication mechanism. So
The blood sample can be separated into the second tubular container and a gap between the first and second tubular containers. In addition, the first and second
In the gap between the tubular containers, a physiologically active substance is produced by a reaction between the stimulating substance and the blood sample. Therefore, the blood sample in the second tubular container is used as a control,
It is possible to measure the physiologically active substance produced in the tubular container.

That is, in the conventional cell function measurement container or cell function measurement container, at least two containers were required for preparing the control sample and the measurement sample, whereas the cell function according to the present invention was used. In the measurement container, the use of only one cell function measurement container allows easy and efficient processing from collection of a blood sample to preparation of a control sample and a measurement target sample.

In a specific aspect of the cell function measurement container according to the present invention, the communication mechanism is provided on the first groove formed on the side surface of the second tubular container and on the outer peripheral surface of the small-diameter insertion portion of the plug. After the blood sample is collected in the second tubular container, the second groove is used for the second tubular container. The first groove and the second groove provided on the outer peripheral surface of the small-diameter insertion portion of the plug are overlapped to form a flow path, and the blood sample is transferred to the second tubular container only by turning upside down. And a gap between the first and second tubular containers. In a specific aspect of the cell function measurement container according to the present invention, the communication mechanism includes a groove provided on the outer peripheral surface of the small-diameter insertion portion of the plug,
Is formed by the cutout formed at the upper open end of the tubular container, so that after the blood sample is collected in the second tubular container, the cutout formed in the second tubular container, A flow path is formed so as to overlap the groove provided on the outer peripheral surface of the small-diameter insertion portion of the body, and the blood sample is placed in the second tubular container and in the first and second tubular containers simply by turning upside down. Can be separated into gaps between them. Therefore, the cell sample can be easily measured using the blood sample in the second tubular container as a control and the blood sample reacted with the stimulant in the first tubular container as the measurement target sample.

In the cell function measuring method according to the present invention, in the cell function measuring container according to the present invention, the blood guided to the second tubular container is communicated with the inside of the second tubular container by using the communication mechanism. The stimulant and the blood sample react in the gap between the first and second tubular containers to produce a physiologically active substance. Therefore, the cell function can be specified using one cell function measurement container.

In the cell function measuring method according to the present invention, the communication mechanism may include a first groove formed on the inner surface of the second tubular container, and second and third grooves formed on the stopper. In the case of comprising, a blood sample is collected in the second tubular container, the stopper is rotated, and the second groove of the stopper and the first groove of the tubular container overlap with each other. Thus, the blood sample is communicated with the first tubular container by merely communicating the first and second tubular containers with each other, then reversing the entire cell function measuring container upside down, and then reversing the cell function measuring container which has been inverted upside down again. Second
And the blood sample can react with the stimulating substance in the gap between the first and second tubular containers. In the case where the communication mechanism is constituted by a cutout formed at the upper open end of the second tubular container and a groove provided on the outer peripheral surface of the small-diameter insertion portion of the plug, Take a blood sample in the tubular container of
By rotating the stopper, the groove of the stopper and the notch of the tubular container overlap, thereby communicating between the first and second tubular containers, and thereafter, the entire cell function measurement container is turned upside down, The blood sample can be separated into the first tubular container and the second tubular container only by reversing the inverted cell function measuring container again, and the gap between the first and second tubular containers can be reduced. The blood sample can be reacted with a stimulant. Therefore, the cell function can be easily measured by measuring the control sample in the second tubular container and the biologically active substance-containing measurement target sample produced in the gap between the first and second tubular containers. Can be.

When endotoxin is used as the stimulating substance and an enzyme immunoassay reagent is used as a cell function measurement reagent, cytokine production can be induced by endotoxin, and various cell functions of a blood sample can be easily measured. Can be.

Further, the stimulating substance cannot move between the first tubular container and the second tubular container by the inner surface of the first tubular container and / or the outer surface of the second tubular container, or the communication mechanism. When physically or chemically immobilized on a carrier having a size, there is no possibility that the stimulating substance may be mixed into the second tubular container during the dispensing operation, and the cell function of the blood sample may be improved. Measurement can be performed with higher accuracy.

[Brief description of the drawings]

FIGS. 1A and 1B are longitudinal sectional views for explaining a cell function measuring method using a cell function measuring container according to an embodiment of the present invention.

FIGS. 2A and 2B are longitudinal sectional views for explaining a cell function measuring method using a cell function measuring container according to an embodiment of the present invention.

FIG. 3 is a perspective view showing the appearance of the cell function measurement container according to one embodiment of the present invention.

FIGS. 4A and 4B are an exploded perspective view for explaining a relationship between a second tubular container and a plug of the cell function measuring container according to one embodiment of the present invention, and a plug shown below. FIG.

FIG. 5 is a perspective view showing a first tubular container of the cell function measuring container used in one embodiment of the present invention.

FIGS. 6A and 6B are an exploded perspective view and a plug for explaining the relationship between a second tubular container and a plug of a cell function measurement container according to another embodiment of the present invention. The perspective view seen from the lower part.

[Explanation of symbols]

 DESCRIPTION OF SYMBOLS 1 ... Cell function measurement container 2 ... 1st tubular container 2a ... Opening 3 ... 2nd tubular container 3a ... Opening 3b ... 1st groove 3c ... Notch part 4 ... Plug 4a ... Handle 4b ... Large diameter insertion Portion 4c: small-diameter insertion portion 4d: second groove 4e: third groove 4f: groove 5: stimulating substance 6: anticoagulant 7: blood collection needle 8: blood sample

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) G01N 35/02 A61B 5/14 300C 300Z 300B 310 F Term (Reference) 2G045 AA01 BB34 BB60 CA25 DA36 HA06 HA09 HA13 HA14 2G058 AA09 CC17 CC18 CC19 EA08 FA03 4C038 KK00 KL01 KL07 KM00 KY01 KY11 TA10 UA03 UA06 UC02 UC04 UD04

Claims (9)

[Claims] 1. A cell function measurement container for measuring a cell function of a blood sample, comprising: a first tubular container having an opening at an upper end and having a bottom; and being inserted into the first tubular container. A second tubular container for collecting blood, having an opening at the upper end and having a bottom, a plug closing the first and second tubular containers, A stimulator that is accommodated in a gap between the two tubular containers and reacts with the blood sample to induce the production of a physiologically active substance. The blood sample guided into the second tubular container is provided in the first tubular container. ,
A cell function measurement container, further comprising a communication mechanism that enables dispensing into a gap between the second tubular containers. 2. A cell function measuring method using the cell function measuring container according to claim 1, wherein a step of collecting a blood sample in a second tubular container, and a second step of using the communication mechanism. At least a part of the blood sample collected in the tubular container is allowed to flow into the gap between the first and second tubular containers, and the blood sample is separated into the first tubular container and the second tubular container, Reacting a blood sample with a stimulating substance in a gap between the first tubular container and the second tubular container. 3. The plug body, wherein the communication mechanism includes the plug body and a first groove formed on the inner surface of the second tubular container so as to extend downward from an upper open end. Is connected to a large-diameter insertion portion that closes an upper end opening of the first tubular container in a liquid-tight manner, and is connected to a lower end of the large-diameter insertion portion, and closes an upper end opening of the second tubular container in a liquid-tight manner. A small-diameter insertion portion for closing, a second groove extending vertically in the outer peripheral surface of the small-diameter insertion portion, and a gap between the first and second tubular containers and the second groove; A third groove is formed on the lower surface of the large-diameter insertion portion so as to communicate
The cell function measurement container according to claim 1, wherein the lower end of the groove is positioned lower than the lower end of the small-diameter insertion portion when the small-diameter insertion portion is inserted into the second tubular container. 4. The communication mechanism comprises a plug and a first groove formed on the inner surface of the second tubular container so as to extend downward from an upper open end, wherein the plug is A large-diameter insertion portion for closing an upper end opening of the first tubular container in a liquid-tight manner, and a lower end of the large-diameter insertion portion being connected to the lower end of the second tubular container, and a liquid-tight closing of an upper end opening of the second tubular container. A second groove extending vertically in an outer peripheral surface of the small diameter insertion portion, and an upper end of the second groove has a first and second tubular container. Is connected to a third groove formed on the lower surface of the large-diameter insertion portion so as to communicate the gap with the second groove, and the lower end of the first groove is connected to the small-diameter insertion portion by the second tubular container. When the blood sample is inserted into the second tubular container, the blood sample is located below the lower end of the small-diameter insertion portion, and a part of the blood sample is Flowing from inside into the gap between the first and second tubular containers, rotating the plug, causing the second groove of the plug to overlap the first groove of the second tubular container, and 1, a step of communicating between the second tubular containers, and the first and second tubular containers closed by the stopper are turned upside down, and the blood sample is placed in the first tubular container from within the second tubular container. Flowing the blood sample into the container, and turning the cell function measuring container upside down upside down again to separate the blood sample into the first tubular container and the second tubular container, and the first tubular container and the second tubular container. Reacting a blood sample with a stimulant in a gap between the tubular container and the stimulating substance. 5. The communication mechanism comprises the plug and a notch formed in an upper opening end of a second tubular container, wherein the plug is an upper end opening of the first tubular container. A large-diameter insertion portion that is liquid-tightly closed, and a small-diameter insertion portion that is connected to a lower end of the large-diameter insertion portion and that closes an upper end opening of the second tubular container in a liquid-tight manner. The cell function measurement container according to claim 1, wherein the cell function measurement container has a groove extending vertically in the outer peripheral surface of the small-diameter insertion portion and continuing downward. 6. The communication mechanism comprises a plug and a cutout formed at an upper open end of a second tubular container, wherein the plug has an upper end opening of the first tubular container. It has a large-diameter insertion portion that is closed in a liquid-tight manner, and a small-diameter insertion portion that is connected to the lower end of the large-diameter insertion portion and that closes the upper end opening of the second tubular container in a liquid-tight manner. A groove extending vertically and continuing downward is formed on the outer peripheral surface of the small-diameter insertion portion, and a part of the blood sample is transferred from the second tubular container to the first and second portions using the communication mechanism. The step of flowing into the gap between the two tubular containers comprises rotating the plug and overlapping the groove of the plug with the notch of the second tubular container, thereby allowing communication between the first and second tubular containers. And turning the first and second tubular containers closed by the stopper upside down to transfer the blood sample to the second tubular container. Flowing the cell function measurement container upside down again from the inside of the container into the first tubular container, separating the blood sample into the first tubular container and the second tubular container, The step of reacting a blood sample with a stimulating substance in a gap between the first tubular container and the second tubular container. 7. The stimulant is endotoxin.
The cell function measurement container according to claim 1, 3 or 5. 8. The method according to claim 1, wherein the stimulating substance is physically or chemically immobilized on an inner surface of the first tubular container and / or an outer surface of the second tubular container.
8. The cell function measurement container according to 3, 5, or 7. 9. The stimulating substance is physically or chemically immobilized on a carrier having a size that cannot be moved between the first tubular container and the second tubular container by the communication mechanism. 9. The method of claim 1, 3, 5, or 7,
The cell function measurement container according to item 1.