JPS6138447A - Particle analyser - Google Patents

Abstract

PURPOSE:To contrive to enhance accuracy by removing the variation in the output of a beam source, by correcting the detection output signal, which was obtained by measuring the beam scattered by a specimen irradiated with beam from the beam source, by the output signal from the photoelectric detector for detecting said irradiated beam. CONSTITUTION:The parallel beam from a laser beam source 10 is allowed to irradiate the specimen S in a flowing part 2 and forward scattered beam is received by a lens 4 and a photoelectric detector 5 while lens 13a, 13b and photoelectric detectors 14a, 14b are respectively provided in two directions crossing incident beam at right angles to measure laterally scattered beam. A beam splitter 11 is obliquely provided between the beam source 10 and a lens 3 so as to guide a part of laser irradiated beam to a photoelectric detector 12. The outputs of the detectors 14a, 14b and the output of the detector 12 are respectively connected to a differential amplifier and the difference between the signals A, B by scattered beam and the signal C by irradiated beam is amplified to remove the effect by the variation of the beam source 10.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a particle analysis device used in a flow cytometer or the like, which irradiates a specimen with light and corrects the measured value of the generated scattered light based on the intensity of the irradiated light.

A flow cytometer is a device that elucidates the properties and structure of cells by irradiating, for example, laser light onto a cell suspension solution flowing at high speed, and detecting and analyzing photoelectric signals from the scattered light. It is used in fields such as medicine, oncology, and genetics.

In the conventional optical system for particle analysis, a sample S such as a blood cell, for example, is wrapped in a sheath liquid and passes through a flow section 2 having a minute rectangular cross section of, for example, 70 pm x 20 gm in the center of the flow cell l shown in Fig. fj51. 2, collimated light from a laser light source (not shown) is focused through a lens 3, and forward scattered light in the straight direction of the irradiated light is received by a photodetector 5 through a lens 4. Information on the size of the specimen S is obtained. Further, side scattered light in a direction perpendicular to the straight direction of the irradiated light is detected by the lens 6 and the photoelectric detector 7, and information on the internal state of the specimen S can be obtained. Furthermore, when performing cytochemical analysis by fluorescently labeling cells with a flow cytometer, measurement can be performed using a combination of a barrier filter, a lens 6, and a photodetector 7 (not shown).

The photoelectric detector 7 on the side of the forward NK scattered light can sufficiently detect it with a semiconductor detector having normal detection ability, but since the side scattered light is extremely weak compared to the forward scattered light,
For example, a photomultiplier tube such as a photomultiplier is used as the photoelectric detector 7 and the like. Further, in order to obtain a large amount of received side scattered light, it is conceivable to increase the output of the light source, but for example, in the case of a laser light source, the output is required to be several watts. On the other hand, in order to reduce the light source output, it is desired to increase the ability to collect side scattered light, but this can usually be achieved by increasing the numerical aperture of the focusing optical system. However, when the numerical aperture of the condensing optical system is increased in this way, the sample S is
When moving in the optical axis direction, variations in the amount of measured light tend to occur.

FIG. 3 shows how an image is formed when sample particles move along the measurement optical axis. In Fig. 3, when the light emitted by the particle at position 0 is guided to the photoelectric detector 7 by the condenser lens 6, the intensity of the optical signal is equal to the solid angle 2π (1-cosφl).
is proportional to. Similarly, the light emitted by a particle at a position of 0° is proportional to the solid angle 2π (1−cosφ2). Therefore, if a particle moves from O to Oo on the fl+1 constant optical axis, the output signal of the photoelectric detector 7 will fluctuate, making the measurement inaccurate, even though the same particle is being fixed at X1ll. Become. This is especially true when the solid angle φ is large, that is, when the condenser lens 6
The effect increases as the numerical aperture increases.

Furthermore, in addition to the variations in measured values due to the movement of particles as described above, the detection signal of the scattered light obtained varies considerably due to the influence of output variations of laser photoelectric and the like. That is, the conventional apparatus has the disadvantage that it is susceptible to errors due to the influence of the light source, etc., as well as errors due to variations in the specimen.

In particular, an object of the present invention is to correct the intensity of the obtained scattered light by the intensity of the irradiated light from the light source, in order to eliminate the above-mentioned fluctuations in the light source output, thereby achieving stable M1 with high accuracy.
The purpose is to provide a particle analysis device that can perform particle analysis, and its gist is to provide a flow cell having a flow section through which a sample passes;
A photoelectric detector for measuring the light scattered by the specimen by the irradiated light from the light source, a photoelectric detector for detecting the light intensity of the irradiated light, and an output signal from the photoelectric detector for detecting the scattered light. The present invention is characterized in that it includes an arithmetic processing circuit for performing correction using an output signal from a photoelectric detector that detects the irradiated light.

The present invention will be explained in detail based on the embodiments shown in FIGS. 4 and 5.

Here, FIG. 4 is a block diagram of the optical system, and FIG. 5 is a block circuit diagram of the signal processing system. Note that the same reference numerals as in FIG. 2 represent the same members.

In FIG. 4, reference numeral 10 denotes a laser light source, and parallel light from this light source 10 is transmitted through a lens 3 to the sample S in the flow section 2.
The forward scattered light is received by the lens 4 and the photoelectric detector 5 in the same manner as in the case of Fig. g:32. A beam splitter 11 is installed obliquely between the light source 1O and the lens 3, and guides a portion of the laser irradiation light to the photoelectric detector 12. In order to photometer the side scattered light, a lens 13a and a photoelectric detector 14a, and a lens 13b and a photoelectric detector 14b are installed in two directions perpendicular to the amount of light incident on the flow section 2, respectively.

Then, as shown in FIG. 5, photoelectric detectors 14a and 14b
The outputs of
It is connected to the. On the other hand, the output of the photoelectric detector 12 passes through a variable resistor 18 and a capacitor 19 to a differential amplifier 17a.
17b in parallel. differential amplifier 17a,
Analog-to-digital converters 20a and 20b are connected to 17b, respectively, and the outputs of these analog-to-digital converters 20a and 20b are connected to a series of serially configured arithmetic section 21, storage section 22, and display section 23. Connected to circuit equipment.

Since the embodiment of the present invention has the above-described configuration, when the sample S passes through the flow section 2 of the flow cell 1, the forward scattered light by the sample S is determined by the lens 4 and the photoelectric detector 5, and is processed by a processing circuit (not shown). Data is processed. In addition, side scattered light is detected by photoelectric detectors 1 arranged in opposite directions.
3a and 13b into optical intensity signals A and B, which are input to the differential amplifiers 17a and 17b, and then sent to the photoelectric converter 12.
The irradiated light intensity signal C is also input to the differential amplifiers 17a and 17b. The resistance values of the variable resistors 15a, 15b, 18 are set in advance so that the outputs of the differential amplifiers 17a, 17b are minimized, and the optical signals A, B, and
The optical signal C caused by the irradiation light is transmitted to a capacitor 16a,
16b and 19 remove unnecessary low frequency components. The differential amplifier 17a amplifies the difference between the scattered light signal A and the irradiated light signal C, and the differential amplifier 17b amplifies the difference between the scattered light signals B and C. influence can be removed.

Therefore, the output signals E obtained by the differential amplifiers 17a and 17b have low noise and high precision, and are converted into digital signals F and G by the analog-to-digital converters 20a and 20b, respectively. It is calculated in section 21. Then, for example, the average value of the sum of the digital signals F and G is stored moment by moment in the storage unit 22 as side scattered light data of the specimen S, and is finally displayed on the display unit 23 such as a CRT as histogram data. It is now possible to do so.

In the above embodiment, side scattered light signals in multiple directions are corrected by the irradiation light signal, but the scattered light signals in multiple directions do not necessarily have to be output signals, and forward scattered light may also be corrected. It is possible.

In this way, the particle analysis device according to the present invention extracts a part of the irradiated light and corrects the output of the scattered light based on its intensity, thereby eliminating the influence of the instability of the light source.
It is possible to perform highly accurate and stable sample analysis. Furthermore, if side scattered light is detected from multiple directions and the outputs are averaged, more accurate measurement results can be obtained.

[Brief explanation of the drawing]

Fig. 1 is a perspective view of a flow cell, Fig. 2 is a sectional view of a conventional device, Fig. 3 is an explanatory diagram of the light collection efficiency of a lens, and Fig. 4 and the following show an embodiment of a particle analysis device according to the present invention. , FIG. 4 is a sectional view of the optical system, and FIG. 5 is a block circuit diagram of the signal processing system. Code 1 is the flow cell, 2 is the distribution section, 3.4.13a, 1
3b is a lens, 5, 12.14a, 14b is a photoelectric detector, 10 is a laser light source, 11 is a beam splitter, 17
a and 17b are amplifiers, 20a and 20b are analog-to-digital converters, 21 is an arithmetic unit, 22 is a storage unit, 23 is a display unit, and S is a sample. Patent applicant Canon Co., Ltd. Representative Patent attorney Yukihiko Hibiya', Ll'H
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Claims (1)

[Claims] 1. A flow cell having a flow section through which a specimen passes, a photoelectric detector for measuring light scattered by the specimen by irradiation light from a light source, and a photoelectric detector for detecting the light intensity of the irradiation light. A particle comprising a photoelectric detector and an arithmetic processing circuit for correcting an output signal from the photoelectric detector that detects the scattered light with an output signal from the photoelectric detector that detects the irradiated light. Analysis device. 2. The particle analysis device according to claim 1, wherein the detection of the light intensity of the irradiation light is performed by interposing a beam splitter in the irradiation optical path and extracting a part of the irradiation light. 3. The particle analysis device according to claim 1, wherein the scattered light to be corrected is side scattered light. 4. The particle analysis device according to claim 3, wherein the side scattered light is detected on both sides of the specimen, and these outputs are averaged.