JPS628038A - Apparatus for analyzing particle - Google Patents

Abstract

PURPOSE:To make it possible to easily perform the axial adjustment of the center axis of a flow cell and the optical axis of irradiation light, by detecting and displaying the intensity distribution of irradiation light. CONSTITUTION:Specimen particles pass through the flow part 2 of a flow cell 1 and a laser beam source 3 is arranged in the direction crossing said flow at right angles. The laser beam L allowed to irradiate the flow part 2 through an image forming lens 4 is scattered by specimen particles to reach a photoelectric detector 7 through a beam splitter 5 and a condensing lens 6 and the information relation to the size of the specimen particles is mainly obtained. The luminous fluxes divided and reflected by the beam splitter 5 are formed on an array type photoelectric detector 9 as an image by a lens 8 and, when the output signal I thereof is displayed on a monitor 10, a left-and-right symmetric smooth Gauss intensity distribution wave form P can be observed. Next, when the center axis of the flow part 2 does not coincide with the optical axis 01 of emitted beam, the output signal I of the detector comes to asymmetric Gauss distribution having disturbance generated in the right side and, therefore, by observing this distribution wave form P, the adjustment of the center axis of the flow cell and the optical axis of irradiation beam becomes easy.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Application Field] The present invention relates to a particle analysis device that makes it possible to easily determine the alignment state in a flow cytometer or the like.

[Prior Art] In a conventional particle analysis device used in a flow cytometer or the like, a particle size of, for example, 200 pm
Irradiation light is irradiated onto a specimen such as blood cells wrapped in sheath fluid and passing through a flow section with a minute cross section of 0.00 am, and the resulting forward and side scattered light is used to determine the shape, number, size, etc. of the specimen. It is possible to obtain particle-like properties such as refractive index.

Further, for a specimen that can be stained with a fluorescent agent, important information for analyzing the specimen can be obtained by detecting the fluorescence of the specimen from side scattered light in a direction substantially perpendicular to the irradiation light.

Laser light is generally used as the irradiation light in such devices, and the intensity distribution of the laser light from a normal laser light source exhibits a Gaussian distribution. On the other hand, since the sample particles are designed to pass through the center of the flow section while being wrapped in the sheath liquid, if the optical axis of the laser beam and the center of the flow section match, the sample particles will receive a strong The most intense laser light is irradiated, allowing for efficient measurements. However, if the optical axis of the laser beam and the center of the flow section are relatively misaligned, or the flow of sample particles deviates from the center of the flow section, the intensity of the laser light irradiated to the sample particles will follow a Gaussian distribution. The scattered light obtained also becomes weak, and even if the same particle is measured, the same light detection signal may not be obtained.

Therefore, in order to perform accurate measurements using a flow cytometer or the like, it is necessary to precisely align the flow axis of sample particles with the optical axis.

For this reason, in conventional devices, the operator manually adjusts the focus and optical axis by visual inspection while flowing a standard sample before measurement, which makes it difficult to perform sufficiently accurate focus and optical axis adjustments. Since it is impossible to confirm changes in the shape of the irradiation beam during measurement and relative deviation between the irradiation beam and the flow section, it is not possible to determine whether or not spurious signals have entered the measurement. , there are concerns about the reliability of the data.

[Object of the Invention] An object of the present invention is to provide a particle analysis device that can easily adjust the central axis of a flow cell and the optical axis of the irradiated light by detecting and displaying the intensity distribution of the irradiated light. There is a particular thing.

[Summary of the Invention] The gist of the present invention for achieving the above-mentioned object is to provide an irradiation optical system that irradiates a light beam to sample particles flowing through a flow section in a flow cell, and a light beam scattered by the sample particles. A photometric optical system that measures light, a detection means that uses a one-dimensional photoelectric conversion element to detect the intensity distribution of the irradiated light that has passed through the flow section, and a display means that displays the output signal waveform of the detection means. This is a particle analysis device featuring:

[Embodiments of the invention] The present invention will be explained in detail based on illustrated embodiments.

Figure 1 shows the overall configuration of the sample particles, which are surrounded by a sheath liquid in a high-speed laminar flow and hydrodynamically focused inside the flow section 2 perpendicular to the plane of the paper in the center of the flow cell I. passes through, and a laser light source 3 is arranged in a direction perpendicular to this flow. An imaging lens 4 is arranged on the optical axis 01 in order to guide the laser beam emitted from the laser beam s3 to the circulation section 2. In addition, a beam splitter 5 is provided on the forward scattered light side of the laser light scattered by the sample particles. A condensing lens 6 and a photoelectric detector 7 are arranged in sequence, and the laser light irradiated at the flow section 2 through the imaging lens 4 is scattered by the sample particles and sent to the beam splitter 5.
The light passes through a condensing lens 6 and reaches a photoelectric detector 7, where information mainly on the size of the sample particles can be obtained. Note that a light-shielding plate (not shown) is usually placed near the condenser lens 6 to remove the straight component of the laser beam that is not scattered by the sample particles, and to convert the information obtained by the photoelectric detector 7 into an S/
It is designed to improve the N ratio.

Furthermore, the optical axis 02 of the luminous flux split by the beam splitter 5
A condensing lens 8 and an array photoelectric detector 9 are arranged above, and the array photoelectric detector 9 has a CR on which a reference index M is displayed.
A monitor 10 such as T is connected. The light beam split and reflected by the beam splitter 5 is then split into a condensing lens 8.
The arrayed photoelectric detector 9, which is a one-dimensional photoelectric conversion element, is
imaged on top.

In addition, the central axis of the flow of sample particles and the optical axis O1 of the laser beam
On the optical axis o3, which is substantially orthogonal to each other, an optical system such as an objective condenser lens 11 for side scattered light, an aperture (not shown), a wavelength selection means, a photoelectric detector, etc. is arranged, and 90% of the sample particles are The scattered light and fluorescence in the direction of 90 ° are measured.
°The granularity of the sample particles can be observed using scattered light, and the biochemical properties of the sample particles can be observed using fluorescence.

By the way, laser light normally has a Gaussian intensity distribution, and the imaging beam in the flow section 2 also has a Gaussian intensity distribution. #Shoot.

If the central axis of the flow section 2 and the optical axis 01 of the irradiated light almost coincide, the intensity distribution of the imaging beam on the arrayed photoelectric detector 9 also exhibits a symmetrical Gaussian intensity distribution, and the arrayed photoelectric detection When the output signal I of the device 9 is displayed on the monitor 10, a symmetrical and smooth Gaussian intensity distribution waveform P can be observed.

FIG. 2 is an explanatory diagram of the output distribution waveform P of the arrayed light tm output device 9 when the central axis of the flow section 2 and the optical axis 01 of the irradiated light do not coincide, for example, the optical axis O1 of the laser beam. is shifted to the right side of the flow section 2 when viewed from the laser light source 3 side, the laser beam hits the inner wall 2R of the flow cell 1, the intensity distribution of the laser beam is disturbed, and the output signal of the arrayed photoelectric detector 9 is The result is an asymmetric Gaussian distribution with disturbances on the right side. Therefore, by observing this distribution waveform P using the monitor lO, it is possible to confirm whether or not the laser beam is hitting the inner wall of the flow cell l.

FIG. 3 is an explanatory diagram of the procedure for actually aligning the central axis of the circulation section 2 and the optical axis O1 of the irradiated light. First of all,
The flow cell l is moved so that a smooth Gaussian distribution waveform P can be obtained on the monitor lO, that is, so that the laser beam does not hit the inner wall of the flow cell l.Normally, the imaging beam diameter in the laser beam distribution section 2 is: In order to prevent disturbances during measurement, it is set shorter than the horizontal length of the flow section 2, so if a smooth Gaussian waveform is obtained on the monitor lO, the center of the flow section 2 and the laser It is not possible to determine whether the optical axis 01 of the light is exactly aligned with the optical axis 01 of the light.

Therefore, first, the flow cell l is moved in the direction A on the left side as seen from the laser light source 3, and when the left inner wall of the circulation section 2 reaches the position a indicated by the broken line, it is moved to the right side of the circulation section 2. Assume that the inner wall 2R is within the beam diameter of the laser beam. Then, the laser beam hits 12R of the flow cell 1, causing disturbance in the Gaussian distribution, and the displayed waveform on the monitor 10 becomes an asymmetric Gaussian distribution waveform Pa with disturbance on the right side.

With such a position a marked on the scale, the flow cell 1 is moved in the opposite direction to the right side B, so that the left inner wall 2L of the flow cell 1 enters the beam diameter of the laser beam, and the monitor 10
The position of the left inner wall 2L of the flow cell I when the displayed waveform above becomes an asymmetric Gaussian distribution waveform pb with disturbance on the left side is calibrated. Then, when the flow cell 1 is moved in the direction A so that the center of the distribution waveform P is located at a position C between this position a and the position C, the distribution waveform obtained on the monitor 10 will be It exhibits a symmetric and smooth Gaussian intensity distribution. This Gaussian distribution waveform Pc indicates that the optical axis O1 of the laser beam and the center of the flow section 2 are completely aligned.

In this way, the laser beam irradiated onto the flow section 2 is imaged on the arrayed photoelectric detector 9, and the output signal is sent to the monitor 10.
By observing this, it is possible to easily adjust the optical axis between the flow cell 1 and the irradiation optical system. After completing the optical axis adjustment, the sample particles are actually passed through the flow section 2, irradiated with laser light, and the forward scattered light transmitted through the beam splitter 5 is detected by the photoelectric detector 7 via the condenser lens 6. You can get accurate measurements by doing this.
□Ru.

In this way, measurement is started with the shafts aligned and the flow section 2 in a clean state, and good measured values can be obtained for a while, but as the measurement continues, the sample inside the flow section 2 Dirt from the liquid may adhere and the wall surface may no longer be in a clean state.

In this situation, the Gaussian intensity of the laser light
; Disturbances occur in the distribution, and even though the same specimen particles are being measured, there will be differences in the measured scattered light intensity, and there is a risk that the measured values will lack reliability.

However, according to the present invention, this can be prevented by monitoring the distribution waveform P on the monitor 10 during measurement. FIG. 4 shows an example of the distribution waveform P on the monitor 10 when dirt adheres to the inner surface of the flow cell 1. When dirt adheres to the inner surface of the flow cell I, the distribution waveform P on the monitor 10 is disturbed. , it is no longer a smooth Gaussian distribution.

Therefore, by monitoring the displayed waveform on the monitor 10 during measurement, it is possible to know the state inside the flow cell 1 during measurement, and measurement is performed only when the distribution waveform P on the monitor 10 is in a normal state. By doing so, highly reliable measurement values can be obtained.

The above has described the optical axis adjustment between the laser irradiation optical system and the flow section 2 of the 70-cell 1. However, by providing the index M on the monitor 10 as shown in FIG. It is also possible to detect the alignment state with the detection optical system. That is, an index M indicating the center position is written on the monitor 10, and by moving the distribution waveform P with respect to the index M, it is possible to determine the 1-axis state and easily adjust the optical axis.

It is also possible to manually move the flow cell 1 to align the axis while monitoring the distribution waveform on the monitor 10, or to detect the inner wall of the flow cell 1 by signal processing the output signal of the arrayed photoelectric detector 9. Therefore, it is also possible to move the flow cell l using a motor drive mechanism or the like and automatically adjust the optical axis.

[Effects of the Invention] As explained above, the particle analysis device according to the present invention has a CO
By installing a detection means such as a D-senna and a monitor that displays the output signal of the detection means, and monitoring the distribution waveform on the monitor, the optical axis between the laser irradiation optical system and the flow cell flow section can be easily adjusted. Furthermore, by monitoring disturbances such as dirt on the inner surface of the flow cell flow section during measurement, highly accurate analysis is possible. Moreover, if a reference index is provided on the monitor, the optical axes of the laser irradiation optical system and the COD detection optical system can be easily adjusted, and measurement accuracy can be further improved. Further, if desired, by providing a mechanism driven by the output signal of the detection means, it becomes possible to perform the optical axis adjustment fully automatically, and furthermore, the measurement operation can be made easier.

[Brief explanation of drawings]

The drawings show an embodiment of the particle analysis device according to the present invention, and FIG. 1 shows the overall configuration, and FIG. 2 shows the relative positional relationship between the irradiated light and the inner wall of the blow cell when the axes are not aligned. Figure 3 is an explanatory diagram of the distribution waveform at that time. Figure 3 is an explanatory diagram of the relative positional relationship between the irradiated light and the inner wall of the flow cell during alignment operation, and the distribution waveform. Figure 4 is an illustration of the distribution waveform when the inner surface of the flow cell is dirty. It is a distribution waveform diagram. 1 is a flow cell, 2 is a flow section, 3 is a laser light source, 5
is a beam splitter, 7 is a photoelectric detector, 9 is an arrayed photodetector, and 10 is a monitor. Figure 2

Claims (1)

[Scope of Claims] 1. An irradiation optical system that irradiates a light beam onto sample particles flowing through a flow section in a flow cell, a photometric optical system that measures scattered light from which the light beam is scattered by the sample particles, and a photometric optical system that measures the scattered light that is scattered by the sample particles; What is claimed is: 1. A particle analysis device comprising: a detection means for detecting the intensity distribution of irradiation light that has passed through the part using a one-dimensional photoelectric conversion element; and a display means for displaying an output signal waveform of the detection means. 2. The light beam is a laser beam, and the detection means detects a disturbance in the Gaussian intensity distribution of the laser beam due to scattering at the edge portion of the flow section, and displays it on the display means, so that the irradiation optical system The particle analysis device according to claim 1, wherein optical axis adjustment is performed with respect to the flow section. 3. The particle analysis apparatus according to claim 1, wherein an index indicating a reference position is provided on the display means, and optical axis adjustment between the irradiation optical system and the detection means is performed. 4. The particle analysis device according to claim 1, wherein the state inside the flow section can be detected by monitoring the output signal waveform of the display means during measurement.