JPH04204254A - Cell sorter - Google Patents

Abstract

PURPOSE:To separate and collect many kinds of cells accurately at the same time by making the tip of the nozzle of a flow cell long in a needle shape, attaching a magnetic body to the tip, arranging electromagnets so as to face the magnetic body, and controlling the magnetic fields of the electromagnets so that the tip of the nozzle is moved when the magnetic forces are imparted to the electromagnets. CONSTITUTION:A slender, needle-shaped nozzle 3a of a flow cell 3 has flexibility in certain degree. The angle of the tip can be freely changed. A permanent magnet 26 is attached to the vicinity of the tip part of the nozzle 3a. Both poles of the permanent magnet 26 and the poles of the same polarity of the electromagnets are arranged so as to face each other. Therefore, one pole of the permanent magnet 26 is attracted to one of the two poles having the same polarity in the electromagnets 27a and 27b. The other pole of the permanent magnet 26 is repelled from the other one of the poles of the electromagnets 27a and 27b. Then, the tip of the nozzle 3a is moved in the lateral direction. The flying track of liquid droplets 29 is changed by the movement of the nozzle 3a. In this way, the many kinds of cells can be accurately separated and collected.

Description

[Detailed Description of the Invention] [Industrial Application Field] The present invention is a cell sorting bag! (cell sorter) sorting mechanism.

(Prior art) A cell sorter is a device that can sort and collect specific types of cells in a living state, and is used for cell analysis, DNA amount measurement, monoclonal antibody production, research such as 5-cell fusion, blood cell analysis, It is used in clinical tests such as cancer screening, and in a wide range of other fields of medicine and biology.

A schematic diagram of a typical device configuration is shown in FIG. 4, and its operation will be explained with reference to FIG. A sample liquid 1 containing cells and a sheath liquid 2 are guided from their respective containers to a flow cell 3 through piping by pressurized air. In the flow cell 3, the sheath liquid 2 surrounds the sample liquid 1 in a cylindrical shape to form a sheath flow, and is ejected from a nozzle at the lower end of the flow cell 3 as a jet stream 4 while remaining in a sheath shape.

The sheath flow is formed so that the cells flow precisely one by one along the central axis of the jet stream 4. The lower end of the flow cell 3 or the jet stream 4 is irradiated with laser light 7 emitted from a laser light source 5 and narrowed down by a lens system 6 . The cells contained in the sample liquid 1 are coated with various types of fluorescent substances (fluorescent probes), and when the cells pass through the laser beam 7, scattered light and fluorescence are generated. The scattered light passes through a condensing optical system consisting of a condensing lens 8 and a beam block 9, and is detected by a photodetector 10. On the other hand, regarding fluorescence,
Red fluorescence is detected by condensing lens 11. Half mirror 12° condensing lens 13. The green fluorescence is collected by a condensing optical system consisting of a filter 14 and detected by a photodetector 15, and the green fluorescence is sent from a half mirror 12 through a condensing lens 16. It is collected by a filter 17 and detected by a photodetector 18. Signals from the photodetectors 10, 15, 18 are sent to a signal processing circuit 19, where the intensity of scattered light and fluorescence is analyzed and cells can be identified.

At this time, minute vibrations are applied to the flow cell 3 by the piezoelectric vibrator 20, and the jet stream 4 exits the nozzle of the flow cell 3 and travels a certain distance, and then the liquid 2
It becomes 1. The number of droplets 21 formed per unit time matches the frequency of vibration applied to the flow cell 3. Immediately before forming the droplet 21 containing the cells to be sorted, a voltage is applied by a controller (not shown) that feeds back the previous analysis results to give it a positive or negative charge, and the charged liquid 111i21 is a pair of opposing deflectors tfi to which a high voltage is applied by the controller 22;
23a.

23b, and its trajectory changes as it receives electrostatic force. The droplets 24 whose trajectories have changed are dropped into different sorting containers 25, and desired cells can be sorted into each sorting container 25.

However, the above device has the following problems.

[The invention tries to solve the problem 1! ! I) In the above cell sorter, there are only two types of cells that can be sorted at -degrees by changing the polarity (±) of the voltage applied to the droplet 21 or the deflection plates 23a and 23b. - It is desirable to be able to sort out many types of cells at once.

On the other hand, in order to sort out many types of cells, it is necessary to adjust the deflection angle by appropriately adjusting the magnitude of the voltage applied to the deflection plate, as described in JP-A-62-255868, for example. There is a method in which the droplets are changed into multiple stages and the finely divided receiver is dropped onto a plate, but in this case, when one droplet is charged, subsequent droplets are also affected by the charge due to electrostatic induction. Therefore, there is a problem in that the voltage waveform applied to the deflection plate becomes quite complex and difficult to control. Furthermore, even if the electric field size is set correctly, there are variations in the size or mass of droplets that fall one after another, and this causes subtle changes in the trajectory of the droplets. It was not possible to perform very fine fractionations.

The present invention has been made in view of the above-mentioned points, and its object is to provide a cell sorter that can perform highly accurate fractionation with a simple device configuration and control mechanism.

[Means for Solving the Problems] In order to solve the above problems, the cell sorter of the present invention attaches a magnetic material near the tip of a flow cell nozzle processed into an elongated needle shape, and installs an electromagnet to face the magnetic material. The flow system and separation mechanism are configured such that at least one electromagnet is disposed and the droplets falling from the nozzle tip are deflected when the electromagnet attracts the magnetic material.

(Function) In the cell sorter of the present invention configured as described above, since the nozzle of the flow cell has a certain degree of flexibility, the angle of the nozzle tip can be changed, thereby temporarily changing the falling position of droplets. This makes it possible to accurately guide droplets to any desired location.

In other words, in a cell sorter, the pressure applied to the liquid in the flow cell is sufficiently large, so the trajectory of the droplet leaving the flow cell is determined by the angle at which the droplet exits without being affected by gravity, and therefore the droplet size and The cell sorter of the present invention is hardly affected by changes in trajectory due to variations in mass, etc. Therefore, by selecting the angle of the flow cell nozzle, it is possible to easily sort out many types of cells at once.

(Example) The present invention will be explained below based on Examples.

The cell sorter of the present invention differs from the conventional one in the shape of the flow cell nozzle and the mechanism for moving the tip of the nozzle using magnetic force; the other configurations are almost the same as that shown in FIG. Here, only the parts related to the present invention are schematically shown in FIG. 1, and parts common to those in FIG. 4 are denoted by the same reference numerals. In FIG. 1, the nozzle 3a of the glass flow cell 3 has an elongated needle shape, and has an outer diameter of several tens of digits and a length of several tens of digits.
Process it into Therefore, this nozzle 3a has a certain degree of flexibility, and the angle of the tip can be changed freely. For example, the nozzle 3 is located near the tip of the nozzle 3a.
The permanent magnet 26 is attached so that a passes through the permanent magnet 26, and both magnetic poles (N, S) of the permanent magnet 26 are connected to two electromagnets 27a, 27.
Magnetic poles of the same polarity (both N or S) are arranged at a predetermined interval between opposing gears. In principle, 26 does not need to be a permanent magnet, but may be any magnetic material, and at least one electromagnet is sufficient, such as electromagnet 27a.
, 27b are controlled by controlling the magnitude and direction of the current of the controller 28 which operates based on the signal from the signal processing circuit 19 shown in FIG. S pole), the electromagnet 27
a.

It is attracted to one of the two magnetic poles of the same polarity (for example, N pole) of electromagnet 27b, and is repelled by the other positive magnetic pole (for example, N pole) of electromagnets 27a and 27b (for example, N pole) of the other permanent magnet 26. As a result, the tip of the nozzle 3a moves laterally. FIG. 2 is a schematic diagram showing this situation, and the trajectory of the droplet changes as shown in 29 as the nozzle 3a moves. The tip of the nozzle 3a is approximately 30° from the vertical direction.
It can be oriented at any angle. The receiver is plate 30
(FIG. 1) is placed at a distance of 70 to 80 degrees below the tip of the nozzle 3a, and is divided finely according to the falling angle of the droplet 29 as shown in FIG. When the tray 30 is divided into four sections, for example, about 10 types of cells can be separated and collected by calculating backward from the movable angle of the tip of the nozzle 3a.

Note that when the nozzle 3a at the tip of the flow cell 3 moves, the nozzle 3a
This effect can be eliminated by making the diameter of the nozzle 3a sufficiently small to improve separation of the droplets, and by synchronizing and controlling the period of ultrasonic vibration applied to the flow cell 3 and the movement of the tip of the nozzle 3a.

The above example assumes that the droplet 21 is deflected in one dimension (straight line), but if the number of permanent magnets and electromagnets is increased to deflect the droplet into a two-dimensional droplet, even more types of cells can be separated and collected. It is possible.

FIG. 3 is a diagram showing an embodiment of nozzle movement different from that in FIG. 2. As shown in FIG. 3, the same effect can be obtained by a method in which the nozzle is twisted by magnetic force.

As described above, the cell sorter of the present invention comprises a flow system and a cell sorting mechanism using a simple device, and is capable of sorting many types of cells.

(Effects of the Invention) Conventional cell sorters that charge droplets ejected from a flow cell, change their trajectory through a pair of deflection plates to which a high voltage is applied, and sort out droplets containing cells. In contrast, in the cell sorter according to the present invention, as described in the embodiment, the nozzle tip of the flow cell is connected to a needle. Attach the magnetic material to a long shape,
This is arranged facing at least one ′@electromagnet magnetic body,
Based on the analysis results of the measurement optical system, the nozzle tip moves when a magnetic force is applied to the electromagnet, so by controlling the magnetic field generated by the 1M1 stone, droplets can be dropped to any desired location. As a result, it has become possible to accurately separate and collect many types of cells at once.

[Brief explanation of the drawing]

FIG. 1 is a partial schematic diagram showing the flow system and separation mechanism in the cell sorter of the present invention, and FIG. 2 is a schematic diagram showing changes in droplet tracks as the nozzle moves in the cell sorter of the present invention. FIG. 3 is a schematic diagram showing different embodiments of nozzle movement of the present invention, and FIG. 4 is a schematic diagram for explaining the configuration and operation of a conventional cell sorter. 1: Sample liquid, 2: Sheath liquid, 3: Flow cell, 3a : Nozzle, 4-jet flow, 5: Laser light source, 6: Lens system, 7: Laser light, 8, 11, 13,
16: Condensing lens, 9: Beam block, 10.15
．． 18: Photodetector, 12: Half mirror, 14, 1'
7: Filter, 19: Signal processing circuit, 20: Piezoelectric vibrator, 21: Droplet, 22.28: Controller, 23a, 2
3b: deflection plate, 25: separation container, 26: permanent magnet,
27a, 27b: Electromagnet, 24.29: Droplet with changed trajectory, 30: Receiver is a plate. Proxy Bentonshi Yamaguchi Otori "--1, Nosen 1 Zuman 2 Sendai 3rd Entq Ibakusamemon 1gki Itlikuchi Tomb (1st 4th concave)

Claims (1)

[Claims] 1) The sample liquid containing cell particles and the sheath liquid are introduced from the nozzle of the flow cell, and the sample liquid is ejected as a sheath-shaped jet flow surrounded by the sheath liquid, and the flow cell is vibrated to transform the jet flow into a liquid. a flow system that ejects particles in a line in the form of droplets; a projection optical system that irradiates the jet flow with laser light; and a measurement optical system that detects and analyzes scattered light and fluorescence emitted from the fine particles by irradiation with the laser light; A cell sorter comprising a sorting mechanism that separates and collects the droplets, and sorts the droplets according to the type of fine particles contained in the droplets based on a signal from the measurement optical system. , the flow system is characterized in that the needle-like nozzle of the flow cell has at least one magnetic body attached near its tip, and the sorting mechanism has at least one electromagnet disposed opposite to the magnetic body. cell sorter.