JP2018134035A - Measuring apparatus, measuring method and computer program - Google Patents

Abstract

An object of the present invention is to improve the measurement accuracy of a measuring device for measuring the amount of microorganisms. [Solution] A measuring device for measuring the amount of microorganisms in a sample liquid includes a light source for irradiating the sample liquid with excitation light, and receives light emitted by the excitation light to generate an electrical signal. It includes a light receiving section that performs the conversion, and a calculation section that calculates the amount of microorganisms based on the number of pulses that are included in the electrical signal and have an amplitude equal to or greater than a first predetermined value. The calculation unit is configured to perform calculation based on the number of pulses having an amplitude of at least a first predetermined value when the electrical signal includes a pulse having an amplitude of at least a second predetermined value that is larger than the first predetermined value. The system is configured to calculate the amount of microorganisms that is smaller than the amount of microorganisms. [Selection diagram] Figure 1

Description

  The present invention relates to a measurement technique for measuring the amount of microorganisms in a sample solution.

  In order to stabilize the ship, the ship that is not loaded with cargo travels with ballast water and discharges the ballast water in the sea area where the luggage is loaded. Ballast water is usually discharged in a different area from the area where the ballast water is mounted, so if the ballast water contains microorganisms such as plankton and bacteria, the microorganisms are discharged into areas other than the original habitat. Will be. This may cause problems such as ecosystem destruction.

  In order to deal with such problems, international rules regarding the regulation of ballast water were formulated, and the “International Convention for the Control and Management of Ship Ballast Water and Sediment (Ballast Water Management Convention)” was adopted. ing.

According to the “Guidelines for Ballast Water Sampling (G2)” related to the Ballast Water Management Treaty, the “Ballast Water Emission Standard (D-2)” describes the aquatic organisms living in the ballast water discharged from ships. An allowable number of individuals is defined for each minimum size of aquatic organisms. Specifically, for example, the discharge standard for aquatic organisms with a minimum size of 50 μm or more (hereinafter referred to as “L size organisms”) is 10 pieces / m ³ or less, and aquatic organisms with a minimum size of 10 μm or more and less than 50 μm. The emission standard for organisms (hereinafter referred to as “S size organisms”) is 10 / mL or less.

  As inspection methods for confirming whether or not these emission standards are satisfied, for example, bulk FDA (fluorescein diacetate), chlorophyll fluorescence, PAM, ATP, and pulse counting FDA are known. In the pulse counting FDA, microorganisms in a sample solution are stained and fluorescence of the microorganisms obtained by irradiating the sample solution with excitation light is detected by a detector. Then, the number of living organisms is measured by counting pulses having a certain amplitude or more (for example, Patent Documents 1 to 3 below).

JP 2014-42463 A JP 2014-55796 A JP, 2014-230514, A

  In various inspection methods, the measurement of the S size biomass is generally performed on a sample solution from which L size organisms have been separated by passing through a mesh filter (for example, having a diagonal distance of 50 μm). Since the opening of the mesh filter is smaller than the L-size organism, theoretically, the L-size organism is not included in the sample water after separation. However, in reality, flexible microorganisms are included in L-size organisms. Such microorganisms may pass through the mesh filter by being deformed. When such an event occurs, L-size organisms are mixed into the sample liquid for measuring the S-size biomass, resulting in a deterioration in measurement accuracy.

  Such a problem is not limited to the inspection of ballast water, but is common when measuring the amount of microorganisms of a specific size separated in advance from microorganisms of other sizes.

  SUMMARY An advantage of some aspects of the invention is to solve at least a part of the problems described above, and the invention can be implemented as, for example, the following forms.

  According to the 1st form of this invention, the measuring apparatus for measuring the amount of microorganisms in a sample solution is provided. The measuring apparatus includes a light source for irradiating a sample liquid with excitation light, a light receiving unit that receives light fluorescently emitted by the excitation light and converts the light into an electric signal, and a first light included in the electric signal. A calculation unit that calculates the amount of microorganisms based on the number of pulses having an amplitude greater than or equal to a predetermined value. The calculation unit is calculated based on the number of pulses having an amplitude greater than or equal to the first predetermined value when the electrical signal includes a pulse having an amplitude greater than or equal to the second predetermined value that is greater than the first predetermined value. The amount of microorganisms smaller than the amount of microorganisms to be calculated is calculated as the amount of microorganisms.

  According to such a measuring apparatus, when a large-sized microorganism that is not the target of measurement is mixed in the sample liquid, the measurement accuracy of the amount of microorganisms is improved by calculating the amount of microorganisms excluding the mixed amount of the microorganisms. it can. For example, in the inspection of ballast water, when L-size organisms are mixed in a sample liquid for measuring S-size organisms, the influence of L-size organisms on the amount of microorganisms to be measured can be reduced.

  According to the second aspect of the present invention, in the first aspect, when the electric signal includes a pulse having an amplitude greater than or equal to the second predetermined value, the calculation unit is greater than or equal to the first predetermined value and The amount of microorganisms is calculated based on the number of pulses having an amplitude less than the second predetermined value and the number of pulses having an amplitude greater than or equal to the second predetermined value. According to such a form, the amount of microorganisms obtained by subtracting the amount of contamination of large-sized microorganisms that are not measured can be calculated using the detected amount actually measured in the sample liquid to be measured. Therefore, the measurement accuracy can be further improved.

  According to the third aspect of the present invention, in the first aspect, the calculation unit is predetermined for each condition relating to the sample solution when a pulse having an amplitude equal to or larger than the second predetermined value is included in the electrical signal. The amount of microorganisms is calculated using the calculated coefficient. According to such a form, the amount of microorganisms obtained by subtracting the amount of large-sized microorganisms that are not to be measured can be simply calculated. Therefore, the calculation load of the measuring device is reduced.

  According to a fourth aspect of the present invention, in any one of the first to third aspects, the measurement device may include a case where a predetermined number or more of pulses having an amplitude greater than or equal to the second predetermined value are included in the electrical signal. An informing unit for informing is provided. According to this embodiment, when a large amount of microorganisms that are not to be measured are accidentally mixed in the sample solution, this can be notified to the user. If the user who receives this notification prepares the sample solution again and measures it again, the measurement accuracy can be improved.

  According to the 5th form of this invention, the measuring method for measuring the amount of microorganisms in a sample solution is provided. This measuring method has a step of irradiating the sample liquid with excitation light, a step of converting light emitted by the excitation light into an electrical signal, and an amplitude greater than or equal to a first predetermined value included in the electrical signal. Calculating the amount of microorganisms based on the number of pulses. The step of calculating the amount of microorganisms includes the step of calculating the number of pulses having an amplitude greater than or equal to a first predetermined value when a pulse having an amplitude greater than or equal to a second predetermined value greater than the first predetermined value is included in the electrical signal. A step of calculating a microbial amount having a value smaller than the microbial amount calculated based on the microbial amount as a microbial amount. According to the 5th form, there exists an effect similar to a 1st form.

  According to the sixth aspect of the present invention, a computer program for measuring the amount of microorganisms is provided. This computer program has a function of calculating the amount of microorganisms based on the number of pulses having an amplitude greater than or equal to a first predetermined value of an input electrical signal, and an amplitude greater than or equal to a second predetermined value greater than the first predetermined value. A function of calculating a microbial amount having a value smaller than the microbial amount calculated based on the number of pulses having an amplitude greater than or equal to a first predetermined value as a microbial amount, Make it a computer. According to the 6th form, there exists an effect similar to a 1st form.

  Any of the second to fourth embodiments can be applied to the fifth or sixth embodiment. In addition to the above-described embodiments, the present invention can be realized in various forms such as a control device for controlling the measurement device and a storage medium in which the computer program is recorded.

It is a functional block diagram which shows schematic structure of the measuring apparatus as one Embodiment of this invention. It is a schematic diagram which shows the principle of a measuring apparatus. It is a histogram about the sample liquid containing only L size organisms. It is a histogram about the sample liquid after L size organisms are removed by the mesh filter. FIG. 5 is a diagram in which the histogram of FIG. 4 is separated into a histogram derived from an L size organism and a histogram derived from an S size organism.

  FIG. 1 is a functional block diagram showing a schematic configuration of a measuring apparatus 10 as an embodiment of the present invention. The measuring apparatus 10 is an apparatus for measuring the amount of microorganisms in a sample solution using a pulse counting FDA technique. In the present embodiment, ballast water is used as the sample solution, and the measuring device 10 is used for measuring the S size biomass.

  The measurement apparatus 10 includes a light source unit 20, a light receiving unit 30, a CPU 40, and a sample container housing unit 50. The light source unit 20 irradiates excitation light toward the sample container 51 housed in the sample container housing part 50. As will be described in detail below, the light receiving unit 30 receives light fluorescently emitted by the excitation light and converts it into an electrical signal.

  The CPU 40 controls the overall operation of the measurement apparatus 10 by executing a program stored in the ROM 41 or the RAM 42. In particular, the CPU 40 also functions as a calculation unit that detects the number of light emission based on the number of pulses of the electrical signal obtained by the light receiving unit 30, and calculates the amount of microorganisms based on the number of light emission. However, at least a part of the functions of the CPU 40 may be realized by a hardware circuit specialized for a specific function.

  The sample container storage unit 50 stores the sample container 51 therein. The sample container 51 is formed of a transparent material that transmits light (for example, glass, quartz, acrylic resin, or the like). In the sample container 51, a sample solution to be measured for the amount of microorganisms and a luminescent reagent are accommodated. In this embodiment, ballast water from which L-size organisms have been removed by a mesh filter is used as the sample solution. In the sample container 51, a rotor 52 for agitating the sample solution is inserted. At the time of measurement, the rotor 52 is rotationally driven by the magnetic stirrer 43 in the sample container 51.

  The sample container storage unit 50 includes holding plates 54 and 55 (see FIG. 2). The holding plates 54 and 55 support the sample container 51 on at least two surfaces when the sample container 51 is accommodated in the sample container accommodating part 50. The holding plates 54 and 55 are provided at positions where light from the light source unit 20 is not blocked.

  The measuring apparatus 10 further includes an operation unit 61 and a display unit 62. The operation unit 61 is a user interface for receiving instructions from the user. The display unit 62 is a display for displaying measurement results and the like.

  FIG. 2 is a schematic diagram showing the principle of the measuring apparatus 10. The light source unit 20 includes a light source 21, a collimator 22, and a band pass filter 23. The light source unit 20 is arranged so that excitation light is incident in a direction orthogonal to the irradiated surface 51 a of the sample container 51. Arbitrary light-emitting means can be used for the light source 21, and the light source 21 may be LED, for example. The light from the light source 21 enters the collimator 22 and is collimated. The collimator 22 is not necessary when a light source capable of irradiating parallel light (for example, a parallel light LED, a laser light source, etc.) is used as the light source 21. The collimated light is incident on the band-pass filter 23, and then the slit-like parallel light is applied to the sample liquid in the sample container 51 as excitation light.

  The light receiving unit 30 includes a photomultiplier tube 31, a band pass filter 32, a condensing lens 33, a slit 34, and a relay lens 35. When the sample liquid is irradiated with excitation light, the fluorescent substance contained in the microorganisms floating in the sample liquid is excited and emits fluorescence. The fluorescently emitted light is collected by the relay lens 35 and imaged. The light imaged by the relay lens 35 passes through the slit 34 and is collected by the condensing lens 33. The slit 34 is installed in order to narrow the observation surface of the fluorescence emitted light in a slit shape. Thereby, since the light receiving area of the light receiving surface 31a is narrowed, the area of background fluorescent light emission that becomes noise also narrows. Therefore, the ratio (SN ratio) of the fluorescence emission signal of the microorganism to the background is improved, and the detection accuracy of the fluorescence emission is improved.

  The light that has passed through the slit 34 enters the photomultiplier tube 31 through the bandpass filter 32. The light receiving surface 31 a of the photomultiplier tube 31 is provided perpendicular to the incident direction of incident light from the light source unit 20. Instead of the photomultiplier tube 31, an arbitrary light receiving sensor (for example, a silicon photodiode (SiPD), an avalanche photodiode (APD), etc.) may be used.

  The measurement apparatus 10 described above is used as follows, for example. First, the user uses a pipette or the like to collect 100 ml as a sample from ballast water having a temperature of about 20 ° C. (L size organisms have been removed by the mesh filter) and put it into the sample container 51. Next, a fluorescent staining reagent is added into the sample container 51. As the fluorescent staining reagent, generally known calcein AM (Calcein-AM, manufactured by Promocell GMBH, Germany), FDA, or the like can be used. Next, the user puts the rotor 52 into the sample container 51. Next, the user accommodates the sample container 51 in the sample container accommodating portion 50 and attaches the lid 53 (see FIG. 1). Thus, the measurement preparation is completed.

  Next, the user operates the operation unit 61 to activate the measurement apparatus 10. As a result, the magnetic stirrer 43 is driven, the rotor 52 starts to rotate, and the sample liquid is stirred. Next, the light source 21 is turned on, and the sample liquid in the sample container 51 is irradiated with excitation light. The excitation light has a wavelength of 450 nm to 490 nm, for example. The specimen (microorganism) in the sample container 51 emits fluorescence by irradiation with excitation light. Then, as described above, this fluorescence is detected by the photomultiplier tube 31 via the relay lens 35, the slit 34, the condensing lens 33, and the bandpass filter 32.

  The photomultiplier tube 31 converts received light energy into electrical energy (electrical signal) by using the photoelectric effect. This electrical signal is amplified by an amplifier circuit (not shown) and input to the CPU 40. The amplified electric signal may be input to the CPU 40 after noise is removed by a band-pass filter or the like.

CPU40 detects the light emission number based on the pulse number of an electrical signal as a process of the calculation part mentioned above, Based on the said light emission number, the amount of microorganisms (here, the number of individuals of S size living organisms) in a sample solution is calculated. calculate. Specifically, the CPU 40 counts the number of pulses included in the input electrical signal and having an amplitude (pulse height) greater than or equal to the first predetermined value Th _S and detects the number of light emissions (light reception count value). To do. The first predetermined value Th _S is a threshold value for determining whether or not the pulse corresponds to the S size organism, and is set to a value that can distinguish the pulse of the S size organism from the background. In the pulse counting FDA method, since the fluorescence intensity (that is, the amplitude of the pulse) depends on the size of the microorganism, the size of the microorganism can be determined based on the size of the amplitude. In the present embodiment, since the L-size organism is removed in advance by the mesh filter, the sample solution should not originally contain the L-size organism. For this reason, it can be determined that a pulse having an amplitude equal to or greater than the first predetermined value Th _S corresponds to an S-size organism. Further, the number of luminescence and the amount of microorganisms present in the sample solution have a correlation. Therefore, the number of S-size organisms in the sample liquid can be estimated by grasping the correlation in advance through experiments or the like. That is, the CPU 40 converts the received light count value based on the correlation, and calculates the amount of microorganisms (here, the number of individuals of S-size organisms). The correlation may be stored in the ROM 41 or the RAM 42 in the form of a function, a map, or the like. The amount of microorganisms thus calculated is displayed on the display unit 62. The display unit 62 displays information on whether or not the amount of microorganisms satisfies a reference value (for example, ballast water discharge standard) stored in advance in the ROM 41 or RAM 42 instead of or in addition to the amount of microorganisms. Also good.

  The measurement apparatus 10 described above can improve measurement accuracy by correcting the amount of microorganisms when a large-sized microorganism that is not a measurement target is mixed in the sample solution. In the present application, “correction” is not limited to correcting the value once calculated, but includes calculating the value by a method different from the normal time. In the following, correction in order to reduce the influence of L-size organisms on the measurement value when L-size organisms are mixed in the sample liquid for measuring S-size organism mass in the ballast water test will be described. .

The CPU 40 calculates a microorganism based on the number of pulses having an amplitude equal to or greater than the first predetermined value Th _S when the input electric signal includes a pulse having an amplitude equal to or greater than the second predetermined value Th _L. A microbial amount having a value smaller than the amount is calculated as the microbial amount. In other words, CPU 40, when included a pulse having a second amplitude greater than the predetermined value Th _L to input electrical signals, and is configured to perform correction to reduce the microbial load. The second predetermined value Th _L is a threshold value for determining whether or not the pulse corresponds to the L size organism, and is set to correspond to the minimum size of the L size organism. That is, the CPU 40 corrects the amount of microorganisms when the L-size organism is included in the sample liquid that should contain only the S-size organism.

FIG. 3 is a histogram for a sample solution containing only L-size organisms (that is, not containing S-size organisms). The horizontal axis is the pulse height, and the vertical axis is the appearance frequency of the pulses. The number NL ₁ of L-size organisms can be expressed by the following equation (1).
NK ₁ = K _L × CL _1a (1)
Here, K _L is a coefficient is determined based on the correlation described above. CL _1a is the number of pulses having an amplitude greater than or equal to a second predetermined value Th _L as shown in FIG.

Theoretically, the pulse number CL _1a corresponds to the individual size of the L-size organism. In practice, however, a pulse having an amplitude less than the second predetermined value Th _L may be generated for an L-size organism. For example, such an event may occur when an L-size organism passes through a position shifted from the focal point of the optical system of the light receiving unit 30. For this reason, the pulse generated from the L-size organism includes a pulse having an amplitude equal to or larger than the first predetermined value Th _S and smaller than the second predetermined value Th _L. The number of pulses having an amplitude greater than or equal to the first predetermined value Th _S and less than the second predetermined value Th _L is represented as CL _{1b in} FIG.

  FIG. 4 is a histogram of the sample liquid after the L-size organism is removed by the mesh filter. The vertical and horizontal axes are the same as those in FIG. As described above, this histogram is created based on the number of pulses counted by the CPU 40 when the sample container 51 is attached to the measurement apparatus 10 and the sample liquid in the sample container 51 is measured. However, in this histogram, a pulse derived from an S-size organism and a pulse derived from an L-size organism are mixed by mixing an L-size organism in the sample solution.

FIG. 5 is a diagram in which the histogram of FIG. 4 is separated into a histogram H _L derived from an L size organism and a histogram H _S derived from an S size organism. The number CA ₂ of pulses (including pulses derived from L-size organisms and pulses derived from S-size organisms) having an amplitude greater than or equal to the first predetermined value Th _S and less than the second predetermined value Th _L is given by the following equation: It can be represented by (2).
CA ₂ = CL _2b + CS ₂ (2)

Here, CL _2b is the number of pulses derived from an L size organism having an amplitude equal to or greater than the first predetermined value Th _S and less than the second predetermined value Th _L , and CS ₂ is the first predetermined value The number of pulses derived from an S-size organism having an amplitude equal to or greater than Th _S and less than the second predetermined value Th _L. In FIG. 5, CL _2a is the number of pulses derived from an L-size organism having an amplitude equal to or greater than a second predetermined value Th _L. The pulse number CA ₂ and the pulse number CL _2a can be actually counted based on the pulse height of the pulse included in the electric signal input to the CPU 40. On the other hand, the pulse number CL _2b and the pulse number CS ₂ cannot be actually counted because it is impossible to determine whether each pulse is derived from an L size organism or an S size organism.

When Expression (2) is changed, the following Expression (3) is obtained.
CS ₂ = CA ₂ -CL _2b (3)

Here, the height of the pulse derived from the L-size organism in the sample liquid is the number of pulses CL _1b (greater than or equal to the first predetermined value Th _S and the second predetermined value) in the histogram for the L-size organism shown in FIG. Assuming that the pulses occur at the same ratio as the ratio between the number of pulses having an amplitude less than the value Th _L and the number of pulses CL _1a (the number of pulses having an amplitude greater than or equal to the second predetermined value Th _L ), the first The number of pulses CL _2b derived from an L-size organism having an amplitude equal to or greater than the predetermined value Th _S and less than the second predetermined value Th _L can be estimated by Expression (4).
CL _2b = CL _2a × CL _1b / CL _1a (4)

Therefore, the pulse number CS ₂ is expressed by the following equation (5) from the equations (3) and (4).
_{CS 2 = CA 2 -CL 2a ×} CL 1b / CL 1a ··· (5)

Further, the number NS ₂ of S-size organisms in the sample liquid mixed with L-size organisms can be expressed by the following equation (6).
NS ₂ = K _S × CS ₂ (6)
Here, K _S is a coefficient and is determined based on the above-described correlation.

Therefore, from the equations (5) and (6), the number NS ₂ of S-size organisms in the sample liquid mixed with L-size organisms can be expressed by the following equation (7).
_{NS 2 = K S × (CA} 2 -CL 2a × CL 1b / CL 1a) ··· (7)

In the equation (7), the pulse number CA ₂ and the pulse number CL _2a can be actually counted based on the height of the pulse included in the electric signal input to the CPU 40. Therefore, the coefficient K _S , the pulse number If the CL _1b and the pulse number CL _1a are experimentally obtained in advance, the CPU 40 can calculate the individual number NS ₂ of S-size organisms contained in the sample solution using the equation (7). Expression (7) may be stored in the ROM 41 or the RAM 42.

According to the measuring apparatus 10 described above, the CPU 40, as can be best understood from the equation (3), when the L-size organism is mixed in the sample liquid, by subtracting the mixed portion, S40 The individual amount of size organisms can be corrected. As a result, measurement accuracy is improved. Moreover, the measuring apparatus 10 has the number of pulses having the amplitude less than or equal to the first predetermined value Th _S and less than the second predetermined value Th _L (the number of pulses CA ₂ ) and the second predetermined value Th _L or more. Correction is performed based on the number of pulses having amplitude (number of pulses CL _2a ). That is, correction is performed using the detected amount actually measured in the sample liquid to be measured. Therefore, correction according to the characteristics of the sample liquid can be performed, and the measurement accuracy can be further improved.

In the measurement apparatus 10 described above, the correction of the amount of microorganisms may be simply performed using a coefficient predetermined for each condition relating to the sample solution. For example, the individual amount of the S-size organism may be calculated by multiplying the coefficient set for each condition related to the sample solution by the number of pulses having an amplitude equal to or greater than the first predetermined value Th _S. Alternatively, the S size is obtained by multiplying the coefficient set for each condition related to the sample solution by the number of pulses having an amplitude equal to or larger than the first predetermined value Th _S and smaller than the second predetermined value Th _L. The individual amount of the organism may be calculated. These coefficients may be stored in the ROM 41 or the RAM 42. The conditions related to the sample solution can be various factors that affect the characteristics of the L-size organism (for example, the ease of slipping through the mesh filter), such as the sample solution collection area and the collection season. May be. In the case of such a configuration, the measurement apparatus 10 may receive a condition related to the sample solution input by the user by the operation unit 61, select a coefficient according to the input condition, and use it for calculating the amount of microorganisms. According to such a configuration, correction can be performed simply and the calculation load on the CPU 40 is reduced.

Further, the measuring device 10 described above, CPU 40, when a pulse having a second amplitude greater than the predetermined value Th _L that contain more than a predetermined number of electrical signals, may function as a notifying unit for performing notification. The notification can be in any form. For example, a message may be displayed on the display unit 62, or notification by sound may be performed. According to such a configuration, when a large amount of L-size organisms are accidentally mixed in the sample solution, this can be notified to the user. If the user who receives this notification prepares the sample solution again and measures it again, the measurement accuracy can be improved. Such a notification function may be employed in a measuring apparatus that does not include the correction function described above.

  The specific configuration of the measurement apparatus 10 described above is merely an example, and the correction function described above can be applied to any microorganism amount measurement apparatus that uses the pulse counting FDA method. In addition, the measurement target of the measuring apparatus 10 is not limited to ballast water, but may be an arbitrary sample solution for measuring the amount of microorganisms of a specific size separated in advance from microorganisms of other sizes.

  Although some embodiments of the present invention have been described above, the above-described embodiments of the present invention are intended to facilitate understanding of the present invention and are not intended to limit the present invention. The present invention can be changed and improved without departing from the spirit thereof, and the present invention includes equivalents thereof. Moreover, in the range which can solve at least one part of the subject mentioned above, or the range which exhibits at least one part of an effect, the combination of each component described in the claim and the specification, or omission is possible. .

DESCRIPTION OF SYMBOLS 10 ... Measuring apparatus 20 ... Light source part 21 ... Light source 22 ... Collimator 23 ... Band pass filter 30 ... Light receiving part 31 ... Photomultiplier tube 31a ... Light receiving surface 32 ... Band pass filter 33 ... Condensing lens 34 ... Slit 35 ... Relay Lens 40 ... CPU
41 ... ROM
42 ... RAM
DESCRIPTION OF SYMBOLS 43 ... Magnetic stirrer 50 ... Sample container accommodating part 51 ... Sample container 51a ... Irradiation surface 52 ... Rotor 53 ... Cover 54,55 ... Holding plate 61 ... Operation part 62 ... Display part H _L ... H histogram derived from L size organism _S ... Histogram derived from S size organism Th _S ... First predetermined value Th _L ... Second predetermined value CL _1a ... Number of pulses having an amplitude equal to or greater than the second predetermined value Th _L CL _1b ... First predetermined value Number of pulses having an amplitude equal to or greater than Th _S and less than the second predetermined value Th _L CS ₂ ... S-size organism having an amplitude equal to or greater than the first predetermined value Th _S and less than the second predetermined value Th _L Number of pulses derived from CL _2a ... Number of pulses derived from an L-size organism having an amplitude greater than or equal to a second predetermined value Th _L CL _2b ... More than or equal to a first predetermined value Th _S and less than a second predetermined value Th _L Of pulses derived from L-size organisms with an amplitude of

Claims (6)

A measuring device for measuring the amount of microorganisms in a sample solution,
A light source for irradiating the sample liquid with excitation light;
A light receiving unit that receives the fluorescence emitted by the excitation light and converts the light into an electrical signal;
A calculation unit that calculates the amount of microorganisms based on the number of pulses having an amplitude greater than or equal to a first predetermined value included in the electrical signal,
The calculation unit calculates the number of pulses having an amplitude greater than or equal to the first predetermined value when a pulse having an amplitude greater than or equal to a second predetermined value greater than the first predetermined value is included in the electrical signal. A measuring device configured to calculate a microorganism amount having a value smaller than a microorganism amount calculated based on the microorganism amount as the microorganism amount. The measuring device according to claim 1,
When the electric signal includes a pulse having an amplitude greater than or equal to the second predetermined value, the calculation unit may calculate a pulse having an amplitude greater than or equal to the first predetermined value and less than the second predetermined value. A measuring device that calculates the amount of microorganisms based on the number and the number of pulses having an amplitude greater than or equal to the second predetermined value. The measuring device according to claim 1,
The calculation unit calculates the amount of the microorganism using a coefficient determined in advance for each condition relating to the sample solution when a pulse having an amplitude equal to or greater than the second predetermined value is included in the electrical signal. apparatus. A measuring device according to any one of claims 1 to 3,
A measurement apparatus comprising a notification unit that performs notification when a predetermined number or more of pulses having an amplitude greater than or equal to the second predetermined value are included in the electrical signal. A measurement method for measuring the amount of microorganisms in a sample solution,
Irradiating the sample liquid with excitation light;
Converting light emitted by the excitation light into an electrical signal;
Calculating the amount of microorganisms based on the number of pulses having an amplitude greater than or equal to a first predetermined value included in the electrical signal,
The step of calculating the amount of microorganisms has an amplitude greater than or equal to the first predetermined value when a pulse having an amplitude greater than or equal to a second predetermined value greater than the first predetermined value is included in the electrical signal. A measurement method comprising a step of calculating, as the amount of microorganisms, the amount of microorganisms smaller than the amount of microorganisms calculated based on the number of pulses. A computer program for measuring the amount of microorganisms,
A function of calculating the amount of microorganisms based on the number of pulses having an amplitude equal to or greater than a first predetermined value of an input electrical signal;
When the electric signal includes a pulse having an amplitude equal to or larger than a second predetermined value larger than the first predetermined value, the electric signal is calculated based on the number of pulses having an amplitude equal to or larger than the first predetermined value. A computer program for causing a computer to realize a function of calculating the amount of microorganisms smaller than the amount of microorganisms as the amount of microorganisms.

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