JPH0320642A - Apparatus and method for inspecting specimen - Google Patents

Abstract

PURPOSE:To reduce the number of optical sensors by receiving different optical data from respective specimens by a combination optical sensor. CONSTITUTION:An optical polarizer 2 is controlled by a drive circuit 3 to collectively irradiate the A-point in a flow cell 7 with the laser beam from a laser beam source 1. The forward scattering beam condensed by a lens 9 in the scattering beam generated from the specimen at the A-point is detected in its intensity by a photodetector 6 through an optical fiber 10, a lens 20, a half mirror 21 and a lens 22. Next, when the intensity of the output pulse of the photodetector 6 becomes a predetermined threshold value or less, a signal processing circuit 5 judges that the passage of a particle through the A-point is completed to send a signal to a control circuit 4 and the polarizer 2 is driven by the circuit 3 so that the irradiation position with laser beam becomes a B-point. Subsequently, at the B-point, the intensity of lateral scattering beam is detected by the photodetector 6 through a lens 11, a filter 12, a mirror 13 and the lens 22. In the same way, red and green fluorescences at points C, D are measured and the obtained measured data is analyzed by a memory and operation circuit 23.

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to a sample testing device and method, such as a flow cytometer.

[Prior Art] Conventional sample testing devices, such as flow cytometers, include, for example, those that remove red blood cells from collected whole blood, and those that add serum to a latex reagent and agglutinate the latex through an antigen-antibody reaction. A sample of blood cells, latex particles, and other particles in this sample are separated one by one using a sheath flow system and flowed at high speed, and each flowing particle is irradiated with light to detect the scattering that occurs. Measures optical information such as light and fluorescence.

A sample can be tested by statistically processing the measurements obtained for a large number of particles.

[Problems to be Solved by the Invention] However, conventional flow cytometers use dedicated photodetectors for each item to detect scattered light and fluorescence. A photoelectron image multiplier is generally used as a photodetector, but it is expensive and also has variations in sensitivity of each C, so it is necessary to adjust the A3 level between them to compensate for variations in sensitivity. Ta.

[Means and effects for solving the problems] The present invention to solve the above problems is an apparatus for separating individual specimens in a sample, irradiating each specimen with light one by one, and inspecting the specimens from the optical information obtained thereby. In this case, the number of optical sensors can be reduced by receiving different optical information from each sample using a dual-purpose optical sensor.

[Example] Embodiments of the present invention will be described in detail below with reference to the drawings.

FIG. 1 is a block diagram of an embodiment of the present invention. Laser light emitted from a laser light source 1 serving as a light irradiation means for irradiating a specimen with light is incident on a light deflector 2 provided in an optical path.

As the optical deflector 2, an acousto-optic deflector (AOD) is used. The acousto-optic deflection element can change the degree of polarization of incident light according to the control frequency. The laser beam deflected by the optical deflector 2 is irradiated onto the flow section 8 in the flow cell 7 . Note that the O-order light emitted from the optical deflector 2 is
It is cut at 5. The drive circuit 3 generates a control frequency that controls the degree of polarization of the optical deflector 2, thereby changing the degree of polarization of the laser beam. In this embodiment, by changing the frequency to four predetermined values, it is possible to irradiate laser light to four different positions along the flow direction of the flow section.

Microparticles in the sample, such as blood cells and latex particles, are passed through the flow section 8 in the flow cell 7 using a sheath flow method well known in this field. The particles are separated one by one or one by one, and are sequentially passed through the flow section at a fixed passing speed. The flow direction of the specimen is from the top to the bottom in the figure.

Individual specimens flowing sequentially through points A, B, C, and D in the figure are irradiated with light at each position, and forward scattered light, side scattered light, red fluorescence, and green fluorescence are measured, respectively. On the optical axis, A
Behind the point, a stopper 24 and a lens 9 are arranged,
The end of the optical fiber 10 is placed at a position conjugate to point A with the lens 9 in between. Further, in the lateral direction of point B, a lens 11, a filter 12 that selectively transmits the wavelength of the irradiated laser beam, and a mirror 13 are arranged in this order.

A lens 14, a filter 15 that selectively transmits red fluorescent wavelengths, and a half mirror 16 are disposed on the side of point C, and a lens 17, that transmits green fluorescent wavelengths, is similarly placed on the side of point D. A filter 18 and a half mirror 19 are arranged.

Next, a control method for measuring by deflecting the laser beam and sequentially switching the irradiation position in synchronization with the passage of the specimen will be described.

First, the optical deflector 2 is controlled to irradiate a fixed laser beam onto the location A in the figure. When the specimen approaches point A, scattered light and fluorescence are generated, and at point A, forward scattered light is measured. Among the scattering destinations generated from the point A, forward scattered light collected by the lens 9 enters one end of the optical fiber 10.

Note that a stopper 24 is provided on the optical axis to directly cut the laser beam, thereby preventing the powerful laser beam from entering the optical fiber. The light that entered the optical fiber 10 and exited from the other end passes through a lens 20, a half mirror 21, and a lens 22.
After that, the intensity is detected by a photodetector 6.

In the signal processing circuit 5, the output pulse intensity of the photodetector 6 is 0.
, or below a predetermined threshold, A. When it is determined that the particles have finished passing through the point, a signal is sent to the control circuit 4, and the optical deflector 2 is driven by the drive circuit 3 so that the laser beam irradiation position becomes point B. By instantaneously switching the control frequency of the optical deflector 2, the irradiation position can also be instantaneously switched, and the laser beam can be irradiated to point B in advance of the passage of the specimen.

At point B, side scattered light is measured. Of the scattered light diverging from point B, the side scattered light scattered in the lateral direction with respect to the laser optical axis is incident on the lens 11, and only the wavelength of the laser light is selected by the filter 12, and the side scattered light is transmitted to the mirror l3, lens 2
2, the intensity is detected by a photodetector 6.

As before, when the pulse output of the photodetector 6 becomes O or below a predetermined threshold in the signal processing circuit 5, the laser irradiation point is switched from point 8 to point C.

Thereafter, measurements at point C and point D are performed in the same manner.

Red fluorescence is measured at point C, and green fluorescence is measured at point D. After the measurement at point D is completed, the laser irradiation position is returned to point A and fixed, waiting for the next particle to pass.

Measurements are repeated in the same manner for a large number of samples passing through the circulation section one after another, and the obtained measurement data is analyzed by performing statistical processing etc. in the storage/arithmetic circuit 23. A detailed analysis method is omitted here because various methods have been developed and are known in this field.

Note that the present invention is not limited to the above-mentioned embodiments, and it is not necessary to use a single photodetector to detect all the light, but to detect multiple types of light. You should do it. Generally, the intensity of forward scattered light is high, and the intensity of other light, especially fluorescence, is weak. Therefore, in order to measure both forward scattered light and fluorescence with a single photodetector, a photodetector with a wide dynamic range is required. Therefore, for example, a separate photodetector is used to detect forward scattered light and side scattered light, and a single highly sensitive photodetector is used to detect red and green fluorescence, and measurements are performed sequentially at different times. Then, a photodetector such as a general photodetector can be used. in this case,
Since forward scattered light and fluorescence can be detected simultaneously at the same point, the number of measurements per particle is reduced, which also has the effect of shortening the total measurement time.

[Effects of the Invention] According to the present invention, a dedicated photodetector was conventionally required for each type of light to be detected, but multiple types of light can be detected with a dual-purpose photodetector.

As a result, the number of photodetectors can be reduced, and the device can be made more compact and lower in cost. Furthermore, errors due to variations in the sensitivity of individual photodetectors do not occur.

[Brief explanation of the drawing]

FIG. 1 is a block diagram of an embodiment of the present invention, and the main symbols in the figure are: 1...laser light source, 2...optical deflector, 3...
・Drive circuit, 4...Control circuit, 5...Signal processing circuit, 6...Photodetector, 7...Flow cell, 8
... Distribution section, 10 ... Optical fiber, 12, 15, 15 ... Optical wavelength selection filter, 24,
25...Stotsuba,

Claims (3)

[Claims] (1) A means for separating individual analytes in a sample, a means for irradiating each separated analyte with light one by one, and a means for sequentially measuring different optical information of each analyte using a dual-purpose optical sensor. A sample testing method characterized by having: (2) A light deflection means for deflecting the irradiated light along the passing direction of the specimen in the test portion through which each specimen passes; a control means for controlling the deflection angle of the light deflection means; and each of the test portions. A specimen testing device comprising: a dual-purpose detection means for detecting different optical information from the specimen. (3) It is characterized by having the following steps: separating the individual specimens in the sample and irradiating them with light one by one; and measuring different optical information of each specimen non-simultaneously using a dual-purpose optical sensor. Specimen testing method.