JPS6260071B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION Field of Industrial Application The present invention relates to a secretion production detection device used to select cells with strong proliferative ability and strong antibody-producing ability from among antibody-producing cells that produce secretions such as antibodies. It is.

BACKGROUND OF THE INVENTION Secretory products, such as monoclonal antibodies, are used in studies in antibody chemistry, antigen chemistry, in studies of receptors for hormones and neurotransmitters in pharmacology, and in virology, parasitology, and bacteriology.
In diagnosis, it can be used for molecular biochemical analysis of immunodeficiency diseases or detection of antigens (malignant tumors, etc.), and in treatment, it can be used for organ transplantation (histocompatibility) and passive immunization (injection of antibodies). It is widely used in the treatment of malignant tumors.

Secretions such as monoclonal antibodies are obtained from cells that produce them, and conventional methods for selecting monoclonal antibody-producing cells include, for example, the following method.

Splenocytes (2.5×10 ⁸ cells), tumor cells (2.5×
Cells such as 10 ⁷ cells were fused using polyethylene glycol as a fusion promoter, and after centrifugation, the fusion solution was discarded.
HAT culture solution (Hypoxanthine Aminopferin and
Add culture medium (containing Thymidine) to disperse the cells. After that, the cell suspension obtained by dispersion is dispensed, for example, 0.2 ml into each well provided in a tray, and then placed in a CO ₂ incubator (cultivator).
Culture for 2 weeks. This culture kills unfused cells and proliferates fused cells.
Whether the fused cells are proliferating is determined by placing each well of the tray under a microscope and observing the presence or absence of colonies. For wells in which fused cells were found to be proliferating, the supernatant of the culture was collected using a micropipette.
Dispense into each well of the tray for antibody detection. A tray for antibody detection in which each well is filled with culture supernatant is attached to the antibody detection section, and a plurality of amounts contained in the supernatant are measured. This measurement is usually performed using ELISA (Enzyne linked immuno sorbent).
assay), and the amount of antibody produced by the cells contained in each well can be determined. At this time, the well into which the cell solution was dispensed is, for example, 480
Normally, only about 5 wells produce the desired antibody.

Problems to be Solved by the Invention After culturing, each well of the tray is placed under a microscope to observe the cell colony, and the supernatant of the culture solution is also taken out with a micropipette to detect antibodies. The conventional method for detecting antibody production involves dispensing the antibody into each well of a tray, attaching the tray to the antibody detection unit, and measuring the amount of antibody contained in the supernatant. It is time-consuming, there is a high risk of contamination with bacteria due to exposure to outside air outside of a sterile environment, and it is complicated to match each well of the tray used for culture with each well of the tray for antibody detection.

Means for Solving the Problems The present invention was made to solve the above-mentioned problems, and consists of a tray transport section, a preparative dispensing section, a colony observation section, an assay section, and a controller. The tray transport section includes a tray stand on which a tray with a plurality of wells is placed, and a tray stand for transporting the tray stand to predetermined positions in the preparative dispensing section, colony observation section, and assay section. The preparative dispensing unit includes a manipulator that grips a pipette and positions the pipette above each of a plurality of wells provided in a tray on the tray stand, and a manipulator that aspirates and discharges a fixed amount of liquid from the pipette. The colony observation section consists of an imaging section with a microscope and a processing device that calculates the degree of proliferation from the captured image data, and the controller moves the tray stand on which the culture tray is placed to the imaging section with the microscope. a means for transmitting a signal to the tray conveying section to move the pipette to a position below the section, and a pipette tip corresponding to the culture supernatant of each well of the culture tray whose proliferation level has been determined by the processing device and the detection tray. a means for transmitting a signal to the manipulator to move the detection tray above each well; a means for transmitting a signal to the pump for injecting and discharging the solution into the pipette; and an assay section driving means for transmitting a signal for moving the detection tray into the assay section. The device consists of a means for transmitting a signal to the cell line, and a means for calculating the antibody production ability from the amount of antibody measured in the assay section and the degree of proliferation measured in the colony observation section. The present invention relates to a secretion production ability detection device characterized in that it is a control device.

Effects of the Invention The effects of the present invention will be explained with reference to the drawings. 1st
The figure is a perspective view showing an embodiment of the present invention. The cell solution is placed in each well 1 and the tray 2 after being cultured in a CO ₂ incubator is placed on the tray stand 4 of the tray transport section 3 . A colony observation section 6 is provided to observe the number and size of colonies, and the tray stand 4 is moved to the tray 2 by the drive means 5 of the tray conveyance section 3.
The wells 1 provided in the colony observation section 6 are moved so that they are located below the microscope-equipped image capturing section. As shown in FIG. 1, the driving means 5 of the tray conveying section 3 is composed of stepping motors 7, 7', a feed screw 8, and a base member 9, and the tray used for culture is controlled by the number of input pulses to the stepping motor 7. (Culture tray) 2 position is determined. Air drive or wire drive can also be used as the drive means 5. A tray chuck 10 for holding the tray 2 may be provided on the tray stand 4, and the tray chuck 10 has the following features:
As shown in FIG. 2 (A is a plan view of the tray stand, B is a side view of the tray stand), opening/closing operations in the θ direction and movement in the x and z directions are possible by motor drive. Air drive can also be used instead of motor drive.
In addition, in Fig. 2, 11 and 11' are motors, and 12
1 is a ball screw, and 13 is a peon rack.

The number and size of colonies in each well 1 are observed by the processing device in the colony observation section 6, and the degree of proliferation of each is determined. The colony observation section 6 measures the intensity of the solution-transmitted light, and can determine the degree of proliferation. The processing device in the colony observation section 6 usually uses means within a controller.

A fixed amount of the culture supernatant in each well on the culture tray 2 for which the degree of cell proliferation has been determined is dispensed by a pipette 25 moved by a pipette manipulator 15 in the aliquot/dispensing section 14. and are dispensed into corresponding wells of the antibody detection tray. As the pipette manipulator 15, a horizontal multi-joint type arm, an orthogonal type arm, etc. can be used. The pump 16 controls the suction amount and discharge amount based on the pressure and pressurization time. When collecting the culture supernatant, use pipette 1.
To avoid contamination, the tip of pipette 6 should be used once and has come into contact with the liquid.
The pipette is moved to position 7 using the pipette manipulator 15 and removed, thereby replacing it with a new pipette.

The antibody detection tray collected from each well of culture tray 2 and dispensed into each corresponding well is
Placed on the tray stand 19 in the assay section 18,
The antibody is transported to the antibody detection position inside the assay section 18 by a drive means consisting of a feed screw 21 operated by a drive motor or a drive means consisting of an air drive, and is contained in the supernatant liquid of each well. The amount of antibody is measured. As a detection device in the assay section 18, an ELISA measurement device, a liquid chromatograph, or the like can be used.

The controller 22 is a device including, for example, an input device 23 and a control device 24, and controls each section according to the above-mentioned sections and operating conditions between the sections inputted by the input device 23.

A block diagram and an operational flow diagram showing the signal system in the present invention are shown in FIGS. 3 and 4, respectively.

In FIGS. 3 and 4, the control signal S ₁ is a signal for operating the stepping motor 9 in the tray transport section 5, and the control signal S ₂ transmitted by the controller is a signal for operating the tray chuck 16 in the tray transport section. Motor 11,1 in Figure 2
This is a signal that controls the opening/closing in the θ direction and the movement in the x and y directions by driving 1'.

In accordance with the control signal _S2 , the culture tray 2 is placed on a tray stand by a tray chuck, and in response to the control signal _S1 , the culture tray 2 is moved to a position below the TV camera with a microscope in the colony observation section.

In the colony observation section, an image of the colony observed by a TV camera with a microscope (contains values obtained by image processing of the number, size, and brightness of cell colonies in the well) is sent to the controller ₂₂ as a measurement signal T1. , the controller 22 performs image processing on the measurement signal T ₁ to calculate the proportion of living cells occupying the entire colony screen.

Next, the manipulator 15 is actuated by the signal _S3 , and the pipette 4 is moved to a position above one of the plurality of wells provided on the tray 2 in which the degree of cell proliferation has been determined.

The pump 16 is activated by the control signal _S4 , and a certain amount of the culture supernatant from each well is taken out by the pipette 25, and then transferred to each corresponding well in the antibody detection tray 20 again by the control signal _S3 . , the culture supernatant in the pipette 25 is dispensed into each well of the antibody detection tray 20 in accordance with the control signal _S4 .

According to the control signals _S3 and _S4 , the pipette is changed every time the cell culture supernatant is dispensed into each well of the antibody detection tray.

When the cell culture supernatant has been dispensed into all wells on the antibody detection tray, the antibody detection tray is moved to the tray stand of the assay section by control signals _S1 and _S2 .

Next, in accordance with the control signal _S5 , the antibody detection tray placed on the upper part of the tray stand 19 is moved into the assay section by actuation of the driving means. The measurement signal _T2 contains the amount of antibody contained in the cell solution supernatant measured in the assay section, and the value of the amount of antibody is stored in the controller 22, and from this value, the amount of antibody contained in the cells contained in each well is determined. Calculate the production capacity (=antibody amount measurement value/proliferation degree measurement value).

Through the above process, the degree of cell proliferation and the amount of antibody in each well are measured, and the antibody production ability of the cells contained in each well is automatically detected from these values.

Effects of the Invention In selecting monoclonal antibody-producing cells, it is necessary to select cells with high antibody-producing ability. Using the device of the present invention, wells containing cells with a high proliferation rate and strong antibody-producing ability can be extracted in a short time from a huge amount of cell culture fluid that cannot be processed manually. It is possible to detect it reliably. Furthermore, since the cells are handled in the device in a sterile environment, there is little risk of contamination with bacteria, and the target cells can be stably detected and selected accordingly.

From the above, it is clear that monoclonal antibodies can now be obtained efficiently and will greatly contribute to the application of monoclonal antibodies.

This device can of course be used to detect antibody production ability, but by replacing the antibody detection section with another detection device, it can also be used to detect the target substance production ability of microorganisms that secrete target substances. is possible.

[Brief explanation of the drawing]

FIG. 1 is a perspective view of the antibody production ability detection apparatus of the present invention, FIG. 2A is a plan view of a tray stand in a tray transport section, and FIG. 2B is a side view thereof. FIG. 3 is a block diagram showing the signal system in the present invention, and FIG. 4 is an operational flow diagram. DESCRIPTION OF SYMBOLS 1... Well, 2... Tray, 3... Tray conveyance part, 4... Tray stand, 5... Drive means, 6...
Colony observation section, 14... Preparation/dispensing section, 15...
Manipulator, 16...pump, 18...assay section, 19...tray stand, 20...antibody detection tray, 21...driving means, 22...controller.

Claims (1)

[Claims] 1. A secretion production ability detection device consisting of a tray transport section, a preparative dispensing section, a colony observation section, an assay section, and a controller, wherein the tray transport section includes a tray stand on which a tray with a plurality of wells is mounted, and a tray stand on which a tray with a plurality of wells is placed. It consists of a drive means for transporting the tray stand to each predetermined position in the preparative dispensing section, the colony observation section, and the assay section, and the preparative dispensing section grips a pipette and moves the pipette to the tray on the tray stand. It consists of a manipulator positioned above each of the plurality of wells provided in the well, and a pump for sucking and discharging a certain amount of liquid from the pipette. The controller includes a processing device within the controller that calculates Means for transmitting a signal to a manipulator to move the culture supernatant in each well of the culture tray whose proliferation rate has been determined by the processing device and above each corresponding well of the detection tray; A means for transmitting a signal to the pump to discharge a signal, a means for transmitting a signal to the assay part driving means to move the detection tray into the assay part, and a means for transmitting a signal to the assay part driving means to send a signal to the pump to move the detection tray into the assay part, and a means to send a signal to the assay part driving means to send a signal to the pump, and a means to send a signal to the assay part driving means to send a signal to the assay part to move the detection tray into the assay part. What is claimed is: 1. A secretion production ability detecting device, comprising means for calculating antibody production ability based on blood pressure, and controlling each of the parts according to input operating conditions of each part.