JP2005333868A - Method and device for measuring malaria parasite - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a measuring method and a measuring device for simply detecting a malaria parasite. <P>SOLUTION: The method for measuring the malaria parasite comprises preparing a sample for the measurement by hemolyzing erythrocyte in the blood to separate the malaria parasite, irradiating the prepared sample for the measurement with light, detecting the first scattered light and the second scattered light having a scattering angle different from the scattering angle of the first scattered light, and detecting the malaria parasite in the sample for the measurement based on the detected first and second scattered lights. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention relates to a method and an apparatus for measuring malaria parasites, and more particularly, to a measurement method and an apparatus for easily detecting malaria parasites.

  Malaria is a parasitic infection widely distributed in the tropics and subtropics, and is transmitted by mosquitoes called anopheles. When blood is sucked into an anopheles with malaria parasite, the malaria parasite is injected into human blood together with the anemone saliva. The malaria parasite enters the hepatocytes where it grows and is released again into the blood. The form of the malaria parasite at this time is called merozoite (merozoite). As soon as merozoites are released into the blood, they invade the erythrocytes and grow while changing their morphology. This morphological change is called a life cycle, and each stage of the life cycle is called a ring form, a tropozoite, and a schizont. Malaria parasites that have developed to the schizont destroy red blood cells and become merozoites again, which are released into the blood. The released merozoite invades the red blood cells, and repeats the life cycle and repeats proliferation. The malaria parasite grows by repeating this cycle and continues to destroy the red blood cells in the blood.

  Conventionally, the most frequently used inspection method for malaria diagnosis is visual microscopic inspection. The blood smear is stained with Giemsa and visually observed with a microscope to detect and count red blood cells infected with malaria parasites. However, this method requires complicated steps such as smearing, fixing, and staining a blood sample in order to create a smear. In order to obtain an accurate test result, it is necessary to observe a large number of red blood cells over a long period of time, and it is difficult to process a large number of specimens quickly.

  On the other hand, as a method for rapidly examining the presence or absence of malaria infection, a method of detecting malaria parasites in blood by applying flow cytometry has been studied. For example, erythrocytes in a blood sample are hemolyzed with a reagent containing a hemolytic agent to release the malaria parasite in the erythrocyte, and the measurement sample is prepared by fluorescently staining the malaria parasite with a nucleic acid-staining dye. Introducing into a flow cytometer, irradiating cells flowing in the sheath flow with excitation light, measuring scattered light and fluorescence emitted by the cells, and discriminating malaria parasites from other cell components based on scattered light intensity and fluorescence intensity Patent Document 1 discloses this.

  In addition, using auramine O analog as the first fluorescent dye and a specific condensed benzene derivative as the second fluorescent dye, prepare a measurement sample by staining malaria-infected erythrocytes, and introduce the measurement sample into the flow cytometer Patent Document 2 describes a method of irradiating cells flowing in a sheath flow with excitation light and detecting malaria-infected erythrocytes using forward scattered light emitted from the cells and the intensity of fluorescence.

Japanese Patent Laid-Open No. 11-75892 Japanese Patent Laid-Open No. 6-300753

  An object of the present invention is to easily detect malaria parasites by a novel method using a combination of optical information different from the methods disclosed in Patent Document 1 and Patent Document 2.

  The present invention prepares a measurement sample in which red blood cells in blood are hemolyzed to liberate malaria parasites, irradiates the prepared measurement sample with light, and from the measurement sample irradiated with the light, the first And the second scattered light having a scattering angle different from that of the first scattered light, and based on the detected first scattered light and second scattered light, malaria parasite in the measurement sample is detected. The present invention provides a method for measuring a malaria parasite.

  The present invention also provides a measurement sample preparation unit for preparing a measurement sample in which red blood cells in blood are hemolyzed to liberate malaria parasites, a flow cell for flowing the prepared measurement sample, and a measurement flow flowing in the flow cell A light source for irradiating the sample with light, a first detector for detecting first scattered light from the measurement sample in the flow cell irradiated with the light, and the angle of scattering with the first scattered light Based on the second detector for detecting different second scattered light, the first scattered light detected by the first detector, and the second scattered light detected by the second detector. The present invention provides a malaria parasite measuring apparatus having a control unit for detecting a malaria parasite in a sample.

  According to the present invention, it is possible to easily detect malaria parasites by using a combination of optical information different from the prior art.

  Hereinafter, although the measurement of the blood sample by the malaria parasite measuring apparatus concerning one Embodiment of this invention is demonstrated, this invention is not necessarily limited to this.

Outline The apparatus for measuring malaria parasite of this example applies a flow cytometry method. Human blood is used as the specimen. In the apparatus for measuring malaria parasite, first, a sample for measurement is prepared by mixing a predetermined amount of specimen and a reagent (diluent and hemolytic agent). In the preparation of the measurement sample, the red blood cells in the specimen are hemolyzed by the action of the hemolytic agent, and the malaria parasite parasitized in the red blood cells is released into the liquid. Next, the measurement sample is caused to flow through the flow cell, and the measurement sample flowing through the flow cell is irradiated with laser light. The scattered light emitted from the measurement sample irradiated with laser light is received and photoelectrically converted by the light receiving element to detect it as an electrical signal, and the detected electrical signal is analyzed to detect and count malaria parasites in the specimen. To do.

Reagents The following reagents (hemolyzing agents and diluents) were used for preparing the measurement sample in the malaria parasite measuring apparatus of this example.

  As the hemolyzing agent, Stomatolyzer-FB (II) manufactured by Sysmex Corporation, which is commercially available as a hemolytic agent for an automatic blood cell analyzer, was used. This hemolytic agent contains a quaternary ammonium salt which is a kind of cationic surfactant. Red blood cells in the blood are hemolyzed by the action of the quaternary ammonium salt.

  As a diluent, a cell pack manufactured by Sysmex Corporation, which is commercially available as a diluent for an automatic blood cell counter, was used.

Malaria parasite measuring apparatus FIG. 1 shows an appearance of a malaria parasite measuring apparatus 100. The front surface of the apparatus is provided with a liquid crystal touch panel 101 for performing various setting inputs and displaying and outputting measurement results, a cover 102 for covering a sample preparation unit 200 described later, and a start switch 103. FIG. 2 shows the internal configuration of the malaria parasite measuring apparatus 100. In the space on the right side of the apparatus, a control unit 400 that controls the operation and analysis processing of the apparatus is provided. A detection unit 300 for detecting a signal from the sample solution is provided in the lower left space of the apparatus. In the remaining space, a sample preparation unit 200 for preparing a sample solution is provided.

  Hereafter, each part of the sample preparation part 200, the detection part 300, and the control part 400 is demonstrated.

Configuration of Sample Preparation Unit 200 FIG. 3 is an explanatory diagram showing the sample preparation unit 200. The sample preparation unit 200 includes a sample setting unit 221 on the front right side, a reagent setting unit 222 on the front left side, and an incubator 223 on the back side. A dispensing device 224 is also provided. When the operator opens the cover 102 of FIG. 1, the sample preparation unit 200 shown in FIG. 3 appears. In the sample preparation unit 200, the sample setting unit 221 is configured to set a sample container 225 containing a sample. A reaction vessel 226 is set in the incubator 223. The reagent setting unit 222 is configured to set a reagent container 227 containing a hemolytic agent and a reagent container 231 containing a diluent. The incubator 223 is configured to shake and agitate the liquid in the set reaction vessel 226 while maintaining a predetermined temperature. The dispensing device 224 sucks and discharges a predetermined amount of liquid from its tip, and is configured to be movable forward and backward, up and down, and left and right by a driving device (not shown). The sample container 233 is connected to a flow cell 301 of the detection unit 300 described later.

Configuration of Detection Unit 300 FIG. 4 is an explanatory diagram showing the configuration of the detection unit 300. The detection unit 300 includes a flow cell 301 for flowing a sample solution. The flow cell 301 is a portion irradiated with laser light and has an orifice portion 302 whose internal flow path is narrowed, a nozzle 303 that ejects sample liquid upward toward the orifice portion, a sheath liquid supply port 304, and a drain port. 305. The detection unit 300 includes a laser light source 306 for irradiating laser light. The laser light source 306 is a red semiconductor laser light source that emits laser light having a wavelength of 633 nm. The detection unit 300 further includes a condenser lens 307 that condenses the laser light emitted from the laser light source to the flow cell 301, and a photo that receives forward scattered light emitted from the sample liquid irradiated with the laser light and converts it into an electrical signal. A diode 308, a collector lens 309 for condensing forward scattered light on the photodiode 308, a pinhole 310, and a photo that receives side scattered light emitted from the sample liquid irradiated with the laser light and converts it into an electrical signal. The multiplier tube 311, the collector lens 312 for collecting the side scattered light to the photomultiplier tube 311, the filter 313, the pinhole 314, the photodiode 308 and the electric signal output from the photomultiplier tube 311 are amplified. As a forward scattered light signal and a side scattered light signal An amplifier 315 and 316 to be output to the control unit 400.

  In this example, the photodiode 308 is disposed at a position where it can detect scattered light scattered in the direction of the extension of the optical axis of the laser light incident on the flow cell 301 from the laser light source 306 in order to detect forward scattered light. On the other hand, since the photomultiplier tube 311 detects side scattered light, the photomultiplier tube 311 is disposed at a position where it can detect scattered light scattered in a direction substantially perpendicular to the optical axis of the laser light incident on the flow cell 301 from the laser light source 306. ing. The forward scattered light reflects the size of the test particles such as malaria parasites and blood cells contained in the sample liquid. The side scattered light reflects internal information such as the density inside the test particle and the presence or absence of a specific substance, or the surface state of the test particle.

Configuration of Control Unit 400 FIG. 5 is a block diagram illustrating the configuration of the control unit 400 and the relationship between the control unit 400 and each unit of the apparatus. The control unit 400 is a memory 401, a central processing unit (CPU) 402, a signal processing circuit 403 that processes a signal sent from the detection unit 300, and an operation control for controlling the operation of each unit of the malaria parasite measuring apparatus 100. A circuit 404 is included. The memory 401 stores an analysis program relating to analysis of signals obtained from particles such as malaria parasites and blood cells contained in the sample, and a control program for controlling the operation of each part of the apparatus. Further, the data processed by the signal processing circuit 403 and the processing result by the analysis program are stored. The CPU 402 executes an analysis program and a control program read from the memory 401, processes and analyzes signals detected from the sample solution in the detection unit 300, and controls signals for controlling operations of the respective units of the apparatus. Or sent to the circuit 404. An analysis result by the analysis program is output to the liquid crystal touch panel 101.

  Hereinafter, the details of the operation of the malaria parasite measuring apparatus 100 will be described. FIG. 6 is a flowchart showing overall control of the malaria parasite measuring apparatus 1 by the control program. When the operator presses the start switch 103, the control program is activated, and step S1 (preparation of the sample solution for measurement), step S2 (detection of the particle signal), and step S3 (analysis of the particle signal) are sequentially executed. Thereby, each part of the sample preparation part 200, the detection part 300, and the control part 400 is controlled, and a series of operation | movement of the malaria parasite measuring apparatus 100 is performed automatically. The operation of each part of the apparatus according to steps S1, S2, and S3 will be described below.

Step S1 (Preparation of sample solution for measurement)
In step S1, the sample preparation unit 200 is controlled to prepare a measurement sample. The operation of the sample preparation unit 200 in step S1 will be described with reference to FIG. First, the dispensing device 224 sucks 55 μL of hemolytic agent from the reagent container 227 of the reagent setting unit 222 and dispenses it into the reaction container 226. Next, the dispensing device 224 sucks 945 μL of the diluent from the reagent container 231 of the reagent setting unit 222 and dispenses it into the reaction container 226 containing the hemolytic agent. Next, 20 μL of the sample is sucked from the sample container 225 set in the sample setting unit 221 and dispensed into the reaction container 226 set in the incubator 223. Thereafter, the incubator 223 agitates the specimen for 14 seconds while keeping the liquid temperature in the reaction vessel 226 containing the specimen, the diluent, and the hemolytic agent at 40 ° C., thereby subjecting the specimen to hemolysis. The sample liquid for measurement thus prepared is sucked by the dispensing device 224 and supplied to the sample container 233. The measurement sample solution supplied to the sample container 233 is caused to flow into the flow cell 301 of the detection unit 300.

Step S2 (particle signal detection)
In step S2, the detection unit 300 is controlled to detect a signal reflecting the characteristics of each particle from the particles in the measurement sample liquid. The operation of the detection unit 300 in step S2 will be described with reference to FIG. As described above, when the sample solution is supplied to the sample container 233, the sample solution is guided to the nozzle 303 by the operation of a pump or a valve (not shown). Then, the sample liquid is discharged from the nozzle 303 to the flow cell 301. At the same time, the sheath liquid is supplied to the flow cell 301 through a sheath liquid supply port 304 from a sheath liquid container (not shown). As a result, the sample liquid is wrapped in the sheath liquid in the flow cell 301 and further squeezed by the orifice 302 and flows. By narrowing down the flow of the sample solution, particles such as malaria parasites and blood cells contained in the sample solution can be aligned and flowed to the orifice portion 302. Laser light emitted from the laser light source 306 is squeezed by the condenser lens 307 and irradiated to the sample liquid flowing through the orifice portion 302. The forward scattered light emitted from the particles in the sample liquid that has received the laser light is collected by the collector lens 309. The forward scattered light that has passed through the pinhole 310 is received and photoelectrically converted by the photodiode 308 to become a pulsed forward scattered light signal. Side scattered light emitted from the particles in the sample liquid that has received the laser light is collected by the collector lens 312. The side scattered light that has passed through the filter 313 and the pinhole 314 is received and photoelectrically converted by the photomultiplier tube 311 to become a pulsed side scattered light signal. The forward scattered light signal and the side scattered light signal are amplified by the amplifiers 315 and 316 as particle signals reflecting the characteristics of each particle, and sent to the control unit 400.

Step S3 (particle signal analysis)
The particle signals (forward scattered light signal and side scattered light signal) detected in step S <b> 2 and sent to the control unit 400 are input to the signal processing circuit 403 included in the control unit 400. For each particle signal, the signal processing circuit 403 regards a portion exceeding a predetermined signal intensity as a particle detection signal, and calculates a peak value of the signal intensity for each particle in the signal waveform. Thereby, the forward scattered light intensity and the side scattered light intensity are calculated for each particle in the sample. The forward scattered light intensity and the side scattered light intensity are stored in the memory 401 as data for each particle.

  Data of forward scattered light intensity and side scattered light intensity for each particle stored in the memory 401 is analyzed by an analysis program stored in the memory 401 in advance. Each step of the analysis program read and executed by the CPU 402 will be described with reference to the flowchart of FIG.

  Step S31: Data of forward scattered light intensity and side scattered light intensity corresponding to each particle in the measurement sample is acquired from the memory 401.

  Step S32: The data of the forward scattered light intensity and the side scattered light intensity are developed in a two-dimensional coordinate space, and a two-dimensional scattergram using the forward scattered light intensity and the side scattered light intensity as parameters is created.

  Step S33: A region M in which dots corresponding to the malaria parasite appear is set on the two-dimensional scattergram created in step S32. The coordinate data of the region M is stored in advance in the memory 401, and is read out by the analysis program in this step and applied on the two-dimensional scattergram. The position of the region M is determined based on the forward scattered light intensity and the side scattered light intensity obtained from the specimen that has been confirmed in advance by microscopic examination or the like to contain the malaria parasite.

  An example of the two-dimensional scattergram created in step S33 is shown in FIG. In the two-dimensional scattergram, the vertical axis represents the forward scattered light intensity, and the horizontal axis represents the side scattered light intensity. A region M where dots corresponding to the malaria parasite appear is set. The forward scattered light intensity reflects the size of the test particle. The side scattered light intensity reflects internal information such as the density inside the test particle and the presence or absence of a specific substance, or the surface state of the test particle. As a result of intensive studies, the inventors of the present invention have detected forward scattered light intensity and sides detected between a malaria parasite in a specimen and other particles (for example, non-hemolyzed leukocytes, debris of hemolyzed erythrocytes). It was found that malaria parasites can be detected without using fluorescence staining because of differences in the direction scattered light intensity. In FIG. 8, a group of dots appearing on the left side of the region M is a group of dots corresponding to the debris of hemolyzed red blood cells. A group of dots appearing above the region M is a group of dots corresponding to white blood cells.

  Step S34: For the area M, the number of dots appearing in the area is counted. The number of dots that appear in the region M reflects the number of malaria parasites in the sample solution.

  Step S35: Create data for outputting the created two-dimensional scattergram and the calculated number of malaria parasites to the liquid crystal touch panel 101 as an analysis result, and output the data to the liquid crystal touch panel 101. FIG. 9 shows a state in which the data is output to the liquid crystal touch panel 101. On the liquid crystal touch panel 101, a two-dimensional scattergram and the number of malaria parasites are displayed as analysis results.

Examples of Measurement Results FIGS. 8 and 10 are two-dimensional scattergrams obtained as a result of measuring specimen 1 and specimen 2 using the malaria parasite measuring apparatus 100 described above. Specimen 1 is blood collected from a malaria infected patient. Sample 2 is blood collected from a healthy person. In the two-dimensional scattergram of FIG. 8 obtained as a result of measuring the specimen 1, a group of dots corresponding to the malaria parasite appears in the region M. On the other hand, in the two-dimensional scattergram of FIG. 10 obtained as a result of measuring the specimen 2, almost no dots appear in the region M. This indicates that the malaria parasite is reliably detected by the malaria parasite measuring apparatus 100. In each two-dimensional scattergram, a group of dots appears on the left side of the region M. These dots correspond to remnants of red blood cells that have been hemolyzed by hemolysis. A group of dots appearing above the region M is a group of dots corresponding to white blood cells.

Identification of the growth stage of malaria parasite
As explained in the “Background” section, malaria parasites are the growth stages called ring form, tropozoite, and schizont in peripheral blood. Repeat while repeating. Hereinafter, the malaria parasite measuring apparatus 110 that can determine the growth stage of the malaria parasite included in the measurement sample will be described. The hardware configuration and operation of the malaria parasite measuring apparatus 110, the types and amounts of reagents used, and the control of the apparatus from the preparation of the measurement sample to the detection of the particle signal are the same as those of the malaria parasite measuring apparatus 100. Therefore, the flow of the analysis program from the analysis of the particle signal to the output of the analysis result for the malaria parasite measuring apparatus 110 will be described below. The flowchart is shown in FIG. In the malaria parasite measuring apparatus 110, portions common to the malaria parasite measuring apparatus 100 will be described using the same reference numerals.

  Step S41: Data of forward scattered light intensity and side scattered light intensity corresponding to each particle in the measurement sample is acquired from the memory 401.

  Step S42: The data of the forward scattered light intensity and the side scattered light intensity is developed in a two-dimensional coordinate space, and a two-dimensional scattergram is created using the forward scattered light intensity and the side scattered light intensity as parameters.

  Step S43: On the two-dimensional scattergram created in step S42, a region R in which dots corresponding to the ring form appear, a region T in which dots corresponding to the tropozoite appear, and a dot corresponding to schizont appear. Region S is set. The coordinate data of the regions R, T, and S are stored in advance in the memory 401, read by the analysis program in this step, and applied on the two-dimensional scattergram. The positions of the regions R, T, and S are respectively the forward scattered light intensity and the side scattered light intensity obtained from the specimen containing the ring foam, the specimen containing the tropozoite, and the specimen containing the tropozoite. It is determined based on this.

  An example of the two-dimensional scattergram created in step S43 is shown in FIG. In the two-dimensional scattergram, the vertical axis represents the forward scattered light intensity, and the horizontal axis represents the side scattered light intensity. Regions R, T, and S where dots corresponding to each growth stage of the malaria parasite appear are set. As described above, the forward scattered light intensity reflects the size of the test particle. The side scattered light intensity reflects internal information such as the density inside the test particle and the presence or absence of a specific substance, or the surface state of the test particle. As a result of intensive studies, the present inventors have made a difference in the detected forward scattered light intensity and side scattered light intensity for each growth stage in the malaria parasite, and determined the growth stage of the malaria parasite without using fluorescent staining. Found that is possible.

  Step S44: For the regions R, T, and S, the number of dots appearing in each region is counted, and malaria parasites at each growth stage are counted. Further, the total number of dots appearing in the regions R, T, and S is calculated, and the total number of malaria parasites is calculated.

  Step S45: Create data for outputting the created two-dimensional scattergram, the calculated number of malaria parasites at each growth stage, and the total number of malaria parasites to the liquid crystal touch panel 101 as analysis results, and store the data on the liquid crystal touch panel 101. Output. FIG. 13 shows how the data is output to the liquid crystal touch panel 101. On the liquid crystal touch panel 101, a two-dimensional scattergram, the number of malaria parasites at each growth stage (ring form, tropozoite, schizont), and the total number of malaria parasites are displayed as analysis results.

Examples of Measurement Results FIG. 12, FIG. 14 and FIG. 15 are two-dimensional scattergrams obtained as a result of measuring specimen 3, specimen 4 and specimen 5 using the malaria parasite measuring apparatus 110 described above. . Specimen 3 is obtained by adding cultured malaria parasite to blood collected from a healthy person. Specimen 4 is obtained by adding cultured malaria parasite (however, only ring foam) to blood collected from a healthy person. The sample 5 is blood collected from a healthy person. In the two-dimensional scattergram of FIG. 12 obtained as a result of measuring the specimen 3, dots appear in each of the regions R, T, and S. In the two-dimensional scattergram of FIG. 14 obtained as a result of measuring the specimen 4, a group of dots corresponding to the ring form appears in the region R. However, almost no dots appear in the regions T and S. In the two-dimensional scattergram of FIG. 15 obtained as a result of measuring the specimen 5, almost no dots appear in the regions R, T, and S. From this, the malaria parasite measuring apparatus 110 demonstrated above detects the malaria parasite in a sample reliably, and has also detected the malaria parasite of a specific growth stage (in the case of the said example, ring form) reliably. I understand that.

  In the malaria parasite measuring apparatus of each of the above examples, malaria parasites in the measurement sample are detected based on two types of scattered light, that is, forward scattered light and side scattered light, and fluorescence is not detected. Therefore, the measuring reagent used above contains a hemolytic agent for hemolyzing red blood cells, but does not contain a fluorescent dye for fluorescently staining a substance in a specimen. That is, in each of the above examples, since a staining solution containing a fluorescent dye is unnecessary, it is possible to reduce the types of reagents necessary for the measurement and to reduce the labor and time required for preparing the measurement sample. Conventionally, when detecting fluorescence, a highly sensitive detector for detecting faint light has been required, but such a detector is not necessary in the malaria parasite-measuring device of each of the above examples. Conventionally, in an automatic measurement apparatus, if the fluorescent dye used for measurement remains in the fluid system in the apparatus, it may affect the results of the next measurement, so the fluid system must be thoroughly cleaned. In the malaria parasite measuring apparatus in each of the above examples, since the use of a fluorescent dye is not essential, the cleaning operation in the apparatus can be simplified, the running cost can be reduced, and the apparatus configuration can be simplified. Can do. In addition, for the purpose of detecting another substance to be measured in combination with malaria parasite or increasing the accuracy of malaria parasite detection, information other than scattered light is also detected from the measurement sample and used for analysis. Also good. In this case, the specimen may be fluorescently stained with a staining liquid containing a fluorescent dye, and fluorescence may be detected from the measurement sample liquid together with two types of scattered light.

  In the malaria parasite measuring apparatus in each of the above examples, a hemolytic agent containing a quaternary ammonium salt that is a cationic surfactant is used. However, the hemolytic agent that can be used is not necessarily limited to this. As a substance that has an action of hemolyzing red blood cells, the hemolytic agent includes a cationic surfactant, a nonionic surfactant, or an anionic surfactant other than the quaternary ammonium salts used in the above examples. It can be used by selecting from various surfactants. These substances may be those that sufficiently lyse erythrocytes in a blood sample at the concentration at the time of hemolysis and do not cause excessive damage to the free malaria parasite. Moreover, in the malaria parasite measuring apparatus of each said example, a hemolyzing agent and a dilution liquid are prepared as a separate liquid as a reagent for malaria parasite measurement, and they are mixed in the step which prepares the sample for a measurement. Thus, you may comprise the reagent for a malaria parasite measurement as a reagent kit which consists of a some liquid. However, the configuration of the reagent used in the apparatus and method of the present invention is not necessarily limited to this, and a hemolyzing agent and a diluent may be mixed in advance to form a single malaria parasite-measuring reagent.

  In the malaria parasite measuring apparatus of each of the above examples, a semiconductor laser light source is used as a light source for irradiating the measurement sample solution. However, the present invention is not limited to this, and a gas laser light source represented by an argon ion laser light source is used. It is also possible to use.

  In the malaria parasite measuring apparatus of each example described above, scattered light scattered in the direction of the extension of the optical axis of the laser light incident on the flow cell 301 from the laser light source 306 is detected as forward scattered light. However, the forward scattered light is not necessarily limited to the scattered light scattered in the direction of the extension of the optical axis of the laser light incident on the flow cell 301, and is scattered at other angles as long as it reflects the size of the test particle. Scattered light may be used. In the malaria parasite measuring apparatus of each example described above, scattered light scattered in a direction substantially perpendicular to the optical axis of the laser light incident on the flow cell 301 from the laser light source 306 is detected as side scattered light. However, the side scattered light is not necessarily limited to the scattered light scattered in a direction substantially perpendicular to the optical axis of the laser light incident on the flow cell 301, but may reflect the internal information or surface state of the test particle. For example, scattered light scattered at other angles may be used. For example, it is possible to use scattered light that scatters at a larger angle than the forward scattered light, even if the direction does not intersect the optical axis at a substantially right angle.

It is a figure explaining the external appearance of the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure explaining the internal structure of the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure explaining the sample preparation part of the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure explaining the detection part of the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure explaining the control part of the malaria parasite measuring device which is one Embodiment of this invention. It is a flowchart explaining the whole control of the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a flowchart explaining each step of the analysis program performed by the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure which shows an example of the two-dimensional scattergram created by the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure which shows a mode that the result of the analysis was displayed on the liquid crystal touch panel of the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure which shows an example of the two-dimensional scattergram created by the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a flowchart explaining each step of the analysis program performed by the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure which shows an example of the two-dimensional scattergram created by the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure which shows a mode that the result of the analysis was displayed on the liquid crystal touch panel of the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure which shows an example of the two-dimensional scattergram created by the malaria parasite measuring apparatus which is one Embodiment of this invention. It is a figure which shows an example of the two-dimensional scattergram created by the malaria parasite measuring apparatus which is one Embodiment of this invention.

Explanation of symbols

100 Malaria parasite measuring apparatus 200 Sample preparation unit 300 Detection unit 400 Control unit

Claims (14)

Prepare a measurement sample that hemolyzes red blood cells in the blood to liberate malaria parasites,
Irradiating the prepared measurement sample with light,
From the measurement sample irradiated with the light, detect the first scattered light and the second scattered light having a scattering angle different from the first scattered light,
A malaria parasite-measuring method for detecting a malaria parasite in a measurement sample based on the detected first scattered light and second scattered light.   The malaria parasite measuring method according to claim 1, wherein the first scattered light is forward scattered light, and the second scattered light is side scattered light.   The method for measuring a malaria parasite according to claim 1, wherein, in preparing the measurement sample, a hemolytic agent is added to blood in order to hemolyze red blood cells.   The method for measuring a malaria parasite according to claim 1, wherein the measurement sample is irradiated with light using a semiconductor laser light source.   The malaria parasite measuring method according to claim 4, wherein the semiconductor laser light source is a red semiconductor laser light source.   The malaria parasite measuring method according to claim 1, wherein a growth stage of the malaria parasite is determined based on the detected first scattered light and second scattered light. A measurement sample preparation unit for preparing a measurement sample by hemolyzing red blood cells in blood to liberate malaria parasites;
A flow cell for flowing the prepared measurement sample;
A light source for irradiating the measurement sample flowing in the flow cell with light,
A first detector for detecting first scattered light from the measurement sample in the flow cell irradiated with the light;
A second detector for detecting second scattered light having a scattering angle different from that of the first scattered light;
Based on the first scattered light detected by the first detector and the second scattered light detected by the second detector, a control unit for detecting a malaria parasite in the measurement sample,
An apparatus for measuring malaria parasites.   The malaria parasite measuring apparatus according to claim 7, wherein the first scattered light is forward scattered light, and the second scattered light is side scattered light.   The malaria parasite measuring apparatus according to claim 7, wherein the first detector detects scattered light scattered in an extension line direction of an optical axis of light incident on the flow cell.   The malaria parasite measuring apparatus according to claim 7, wherein the second detector detects scattered light scattered in a direction substantially perpendicular to an optical axis of light incident on the flow cell.   The malaria parasite measuring apparatus according to claim 7, wherein the measurement sample preparation unit includes a hemolytic agent adding means for adding a hemolytic agent to blood.   The malaria parasite measuring apparatus according to claim 7, wherein the light source is a semiconductor laser light source.   The malaria parasite measuring apparatus according to claim 7, wherein the semiconductor laser light source is a red semiconductor laser light source. The malaria parasite measuring apparatus according to claim 7, wherein the control unit determines a growth stage of the malaria parasite based on the first scattered light and the second scattered light.

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