Patent Application
20220178832
The present invention relates to a biological sample analysis device and a biological sample analysis method of analyzing light generated from a biological substance contained in a sample.
Conventionally, microbial monitoring has been performed for environmental management of pharmaceutical production plants, food plants, and the like. As an example of this microbial monitoring, a luminescent reagent is added to adenosine triphosphate (ATP) contained in a microorganism, bioluminescence of the luminescent reagent is measured, an obtained luminescence intensity is converted into an ATP amount, and thus correlation with bacterium can be taken.
As a device that analyzes light generated by a biological substance such as ATP, a device disclosed in Patent Literature 1 is considered. This biological sample analysis device adds a luminescent reagent to a sample containing a biological substance, and detects a luminescence intensity at a luminescence peak and a luminescence intensity in a state where luminescence is reduced after a predetermined time (for example, about 10 minutes) from the luminescence peak. In this device, a luminescence intensity obtained by subtracting the “luminescence intensity in a state where the luminescence is reduced” from the “luminescence intensity at the luminescence peak” is used for conversion into the number of bacteria.
However, in the above method, it is necessary to wait for a predetermined time (for example, about 10 minutes) to elapse from the luminescence peak in order to detect the “luminescence intensity in a state where the luminescence is reduced”, and measurement of one sample takes long time. This problem is particularly noticeable when a plurality of samples are measured.
In a case where the container storing the sample includes resin, the container stores light such as ultraviolet rays or fluorescent lamps outside or stores bioluminescence, and the “luminescence intensity in a state where the luminescence is reduced” includes the light stored in the container, and this deteriorates measurement accuracy.
Patent Literature 1: JP 2008-268019 A
The present invention has been made to solve the above problems, and a main object of the present invention is to provide a biological sample analysis device that stores a sample containing a biological substance and a luminescent reagent in a container, detects luminescence generated by reacting the sample and the luminescent reagent, and analyzes the biological substance, the biological sample analysis device shortening measurement time of the sample and improving measurement accuracy.
Abiological sample analysis device of the present invention stores a sample containing a biological substance and a luminescent reagent in a container, detects luminescence generated by reacting the sample and the luminescent reagent, and analyzes the biological substance, the biological sample analysis device including a photodetector that detects the luminescence and outputs a light intensity signal, and a calculator that subtracts the light intensity signal obtained before the sample and the luminescent reagent react from the light intensity signal obtained after the sample and the luminescent reagent react to remove light stored in the container, and calculates a value related to an amount of the biological substance.
In the biological sample analysis device configured as described above, the light intensity signal obtained before the sample and the luminescent reagent react is subtracted from the light intensity signal obtained after the sample and the luminescent reagent react to calculate the value related to the amount of the biological substance, and thus, there is no need to detect the “luminescence intensity in a state where the luminescence is reduced” as in the conventional art. As a result, measurement time of the sample can be shortened. Further, since the light stored in the container is removed by subtracting the light intensity signal obtained before the sample and the luminescent reagent react from the light intensity signal obtained after the sample and the luminescent reagent react, there is no need to consider the light stored in the container, and measurement accuracy can be improved. Here, the light stored in the container is light other than bioluminescence generated by reaction of the sample and the luminescent reagent, and includes phosphorescence and fluorescence emitted from the container.
Here, in order to improve analysis efficiency of the biological sample, it is conceivable that the biological sample analysis device sequentially measures the luminescence of the sample stored in a plurality of the containers.
In this case, the stored light amounts in the plurality of containers are different from each other. Thus, the stored light amount to be subtracted differs for each container. Therefore, the calculator desirably subtracts the light intensity signal obtained before the sample and the luminescent reagent react from the light intensity signal obtained after the sample and the luminescent reagent react in each of the plurality of containers. This configuration can improve the measurement accuracy in each of the plurality of containers.
In the biological sample analysis device of the present invention, it is conceivable that the biological substance includes adenosine triphosphate (ATP), and ATP-derived luminescence generated by a reaction between the sample and an ATP luminescent reagent is detected.
In order to improve the analysis efficiency by automatically introducing the reagent into the sample, the biological sample analysis device of the present invention desirably further includes a dispensing mechanism that dispenses a reagent into the sample, a standard solution having a known ATP amount, and a zero solution having a zero ATP amount. Specifically, the dispensing mechanism dispenses an ATP scavenging solution, a spore reaction solution, an ATP extract, a luminescent reagent, and the like into the sample, the standard solution, and the zero solution.
Conventionally, solution amounts of each reagent are different among the standard solution, the zero solution, and the sample. As described above, since the solution amounts of each reagent are different among the standard solution, the zero solution, and the sample, pH in the solution is different, a luminescence intensity is different, and accurate light intensity cannot be detected.
In order to suitably solve this problem and detect an accurate light intensity, the dispensing mechanism is desirably controlled to equalize a mixing ratio of the ATP scavenging solution, the spore reaction solution, and the ATP extract to be added to the sample, a mixing ratio of the ATP scavenging solution, the spore reaction solution, and the ATP extract in the standard solution, and a mixing ratio of the ATP scavenging solution, the spore reaction solution, and the ATP extract in the zero solution.
The ATP extract has an effect of inactivating the ATP scavenging solution.
Therefore, the dispensing mechanism is desirably controlled to dispense the ATP extract after adding the ATP scavenging solution and the spore reaction solution to the standard solution.
By dispensing the reagents in that order, the standard solution containing the ATP scavenging solution can be created.
Furthermore, a biological sample analysis device of the present invention desirably further includes a housing body that accommodates a measurement system instrument for biological sample analysis inside and includes an opening, a door that opens and closes the opening of the housing body, and uneven structures respectively provided on contact portions of the opening of the housing body and the door, the uneven structures being fitted to each other in a state where the door closes the opening.
In this configuration, light entering inside of the device from between the housing body and the door can be blocked by the uneven structures, and measurement accuracy can be improved.
In addition, it is conceivable that the biological sample analysis device of the present invention further includes a disposal box in which pipette tips that inject the reagent into the sample are discarded.
In this configuration, in order to reliably discard the pipette tips in the disposal box, the disposal box desirably includes disposal spaces respectively partitioned for the pipette tips to be discarded, the disposal box including a slope that inclines the pipette tips discarded in the disposal spaces in a predetermined direction. Note that, in a configuration in which the slope is not provided, directions of the pipette tips discarded in the disposal spaces vary, and this variation may interfere with the pipette tips to be discarded.
A biological sample analysis method of the present invention stores a sample containing a biological substance and a luminescent reagent in a container, detects luminescence generated by reacting the sample and the luminescent reagent, and analyzes the biological substance, the method including subtracting a light intensity signal obtained before the sample and the luminescent reagent react from the light intensity signal obtained after the sample and the luminescent reagent react to remove light stored in the container, and calculating a value related to an amount of the biological substance.
In the present invention configured as described above, the biological sample analysis device stores a sample containing a biological substance and a luminescent reagent in a container, detects luminescence generated by reacting the sample and the luminescent reagent, and analyzes the biological substance, the biological sample analysis device being capable of shortening the measurement time of the sample and improving the measurement accuracy.
Hereinafter, an embodiment of a biological sample analysis device of the present invention will be described with reference to the drawings.
<Device Configuration>
A biological sample analysis device 100 according to the embodiment analyzes light generated by a biological substance contained in a sample to measure an amount of the biological substance. Hereinafter, an ATP amount measurement device that measures an amount (amol (=10−18 mol)) of adenosine triphosphate (ATP) as a biological substance will be described.
Specifically, as illustrated in
As illustrated in
As illustrated in
The uneven structures 15 and 16 are provided so as to surround substantially an entire periphery of the opening C1h. In the embodiment, the uneven structure 15 provided at the contact portion of the housing body C1 includes a body outer protrusion 151 provided so as to surround the opening C1h, and a body inner protrusion 152 provided so as to surround the opening C1h at an inner side of the body outer protrusion 151. Further, the uneven structure 16 provided at the contact portion of the door C2 includes a door outer protrusion 161 provided so as to surround the opening C1h at an outer side of the body outer protrusion 151 of the housing body C1, and a door inner protrusion 162 inserted between the body outer protrusion 151 and the body inner protrusion 152 of the housing body C1. The uneven structures 15 and 16 allow a path of light from outside to meander, and to be blocked before reaching inside of the device, and the inside of the device becomes a darkroom. This can reduce stray light and improve measurement accuracy. In addition, a space between the door C2 and the opening C1h may be sealed with a seal member (not illustrated) to make the inside of the housing C a darkroom.
Further, the housing body C1 is provided with a temperature control mechanism 7 that holds a plurality of specimen tubes FC storing specimens and controls a temperature of these plurality of specimen tubes FC, a reagent setting part 8 in which reagent containers RC1 and RC2 storing respective reagents are set, and a pipette tip setting part 9 provided with a pipette tip PT used for the dispensing mechanism 6.
The temperature control mechanism 7 accommodates and holds the plurality of specimen tubes FC, for example, in a matrix. The temperature control mechanism 7 includes a metallic (for example, aluminum) holder block 71 that holds the specimen tubes FC, a heat source 72 such as a heater provided in the holder block 71, and a temperature sensor 73 such as a thermocouple that detects a temperature of the holder block 71. On the basis of the temperature detected by the temperature sensor 73, the heater 72 as the heat source is controlled by a controller COM for the temperature of the holder block 71 to be a predetermined temperature.
In the reagent setting part 8, the reagent container RC1 storing a pretreatment reagent for subjecting a specimen to pretreatment and the reagent container RC2 storing a luminescent reagent are set. Examples of the pretreatment reagent include an ATP scavenging solution for scavenging ATP (free ATP) other than living cells (viable bacteria) contained in the specimen, a spore reaction solution for germinating bacteria in a spore state, and an ATP extract for extracting ATP from living cells.
As illustrated in
As illustrated in
The holder drive mechanism 5 moves the holder 3 to sequentially position the containers 2 held by the holder 3 at detection positions Xdet detected by the photodetector 4. Specifically, the holder drive mechanism 5 rotates the holder 3 around the predetermined rotation center. As illustrated in
As illustrated in
The nozzle 61 includes a tip holder 611 that detachably holds the pipette tip PT that contacts and holds the sample and each reagent. The tip holder 611 includes an internal flow path, a base end to which a flow path is connected, and a distal end opening to which a pipette tip PT is connected.
The nozzle moving mechanism 63 linearly moves the nozzle 61 in a horizontal direction (an X-axis direction and a Y-axis direction) and linearly moves the nozzle 61 in a vertical direction (a Z-axis direction). Specifically, the nozzle moving mechanism 63 includes a movable member 631 that holds the nozzle 61, a slide mechanism 632 provided in the X-axis direction, the Y-axis direction, and the Z-axis direction, and an actuator 633 that moves the movable member 631 in each of the directions along the slide mechanism 632. Each operation in the ATP measurement is executed by controlling the actuator 633 and the pump mechanism 62 by the controller COM. Each operation in the ATP measurement naturally includes attachment and detachment of the pipette tip PT to and from the tip holder 611.
Furthermore, as illustrated in
The light shielding mechanism 13 includes a container-side light shield 131 provided in each container 2, and a movable-side light shield 132 that moves forward and backward with respect to the container 2 located at the detection position Xdet.
The container-side light shield 131 is constituted by a member having no light permeability, and covers an entire periphery of an upper part of each container 2. In the embodiment, the container-side light shield 131 having a cylindrical shape is provided on a container holding part of the holder 3, and the container 2 is accommodated in the container-side light shield 131, and thus the container-side light shield 131 covers the entire periphery of the upper part of the container 2 held by the holder 3.
The movable-side light shield 132 is constituted by a member having no light permeability, and covers the entire periphery of a lower part of the container 2 located at the detection position Xdet, except for the upper part covered by the container-side light shield 131. The movable-side light shield 132 ascends and descends between a light shielding position at which the movable-side light shield covers the lower part of the container 2 located at the detection position Xdet and a retraction position at which the movable-side light shield is separated downward from the lower part of the container 2 and does not interfere with the movement of the holder 3 when the holder 3 moves. Note that the ascend and descend of the movable-side light shield 132 is performed by, for example, a lifting device 14 using an actuator. The lifting device 14 is controlled by the controller COM in conjunction with operations of the holder drive mechanism 5 and the dispensing mechanism 6.
In the biological sample analysis device 100 according to the embodiment, a disposal box 10 as a disposal tip storage for disposing the pipette tip PT of the dispensing mechanism 6 is integrally provided with the holder 3. Specifically, the disposal box 10 is provided at an inner side of the plurality of containers 2, the inner side serving as a dead space in the holder 3. The disposal box 10 has an arc-shaped opening 10x along an arrangement direction of the plurality of containers 2 in plan view. The disposal box 10 in plan view has a substantially octagonal ring shape illustrated in
In the holder 3, the holding holes 3h for inserting and holding fingers are formed at an inner side of the arc-shaped opening 10x. In this configuration, in a state where the holder 3 is held by the holding holes 3h, the disposal box 10 and the container 2 are positioned at an outer side of the holding hand, and inadvertent contact with the pipette tip PT having been discarded and the container 2 having been measured can be easily prevented.
Then, the pipette tip PT used for dispensing is detached above the disposal box 10 of the holder 3. Specifically, the detachment may be performed by moving the nozzle 61 to a chip detachment member (not illustrated) arranged above the disposal box 10. Alternatively, the chip detachment member is provided in the movable member 631, the movable member 631 is moved to above the disposal box 10, and then the detachment may be performed using the chip detachment member.
When each pipette tip PT is detached, the controller COM controls the holder drive mechanism 5 and the dispensing mechanism 6 such that the pipette tip PT is not unevenly distributed at one spot in the disposal box 10. As this control mode, it is conceivable that (1) the holder 3 is rotated by a predetermined angle every time each pipette tip PT is detached to change a disposal position with respect to the disposal box 10, (2) the holder 3 is rotated by a predetermined angle every time a predetermined number of pipette tips PT are detached while the disposal position of the predetermined number of pipette tips PT keeps the same to change the disposal position with respect to the disposal box 10, and the like. This control can scatter, as a whole, the pipette tips PT discarded in the disposal box 10, and prevent the pipette tips PT from being unevenly distributed at one spot and protruding from the disposal box 10.
<Analysis Method>
Next, an analysis method will be described together with operation of the biological sample analysis device 100 configured as described above.
For example, a large volume (for example, from 50 ml to 200 ml) of a specimen is concentrated to a predetermined amount (for example, from 1 μl to 1,000 μl) to generate a sample.
In this concentration step, as illustrated in (A) of
As described above, the specimen tube FC storing the concentrated sample is set in the temperature control mechanism 7. The door C2 is closed in a state where a predetermined number of specimen tubes FC are set, and the measurement is started. Although each container 2 held by the holder 3 in this state is empty, the container 2 for standard solution measurement stores a standard solution having a known ATP amount.
When the measurement is started, the controller COM causes the dispensing mechanism 6 to dispense each pretreatment reagent into each of the specimen tubes FC held by the temperature control mechanism 7 in accordance with a predetermined sequence. As a result, the sample in the specimen tubes FC is subjected to predetermined pretreatment (ATP extraction). Note that the pipette tips PT are replaced for each pretreatment reagent, and the used pipette tips PT are discarded in the disposal box 10.
Specifically, a mixed solution of the ATP scavenging solution and the spore reaction solution is dispensed into the sample in the specimen tube FC, and the sample is kept at a predetermined temperature and is on standby until a reaction of each reagent is completed. Thereafter, the ATP extract is dispensed into the sample in the specimen tube FC, and the sample is kept at a predetermined temperature and is on standby until the extraction of ATP is completed. Instead of the mixed solution of the ATP scavenging solution and the spore reaction solution, the ATP scavenging solution and the spore reaction solution may be separately dispensed.
As for a calibration solution, during the standby after the reagent is dispensed into the sample, each pretreatment reagent is dispensed into a standard solution having a known ATP amount and a zero solution having a zero ATP amount in accordance with a predetermined sequence. Specifically, after dispensing the ATP scavenging solution and the spore reaction solution into the standard solution and the zero solution, the ATP extract is dispensed. Since the standard solution is stored in the container 2s for standard solution measurement and the zero solution is stored in the container 2b for blank measurement, the dispensing mechanism 6 dispenses each pretreatment reagent into the container 2s and the container 2b. An order of dispensing each pretreatment reagent into the zero solution is not limited to the above order.
At this time, as shown in
Thereafter, the dispensing mechanism 6 dispenses the sample in each of the specimen tubes FC after the pretreatment into each of the containers 2 held by the holder 3.
Then, the controller COM causes the holder drive mechanism 5 to move the container 2 to be measured to the detection position Xdet. After moving the container 2 to be measured to the detection position Xdet, the controller COM causes the lifting device 14 to move the movable-side light shield 132 of the light shielding mechanism 13 to a light shielding position. After this state, the controller COM causes the dispensing mechanism 6 to introduce the luminescent reagent into the container 2 located at the detection position Xdet. Thus, light emitted from the sample in the container 2 located at the detection position Xdet is detected by the photodetector 4. Before measurement of luminescence of each container 2, blank measurement and standard solution measurement are performed, and zero point calibration and span calibration are performed.
A light intensity signal obtained by the photodetector 4 is subjected to arithmetic processing by a calculator of the controller COM to calculate an ATP amount (amol).
Specifically, a calculator COM1 of the controller COM subtracts a “light intensity signal obtained before the luminescent reagent is added” from a “light intensity signal obtained after the luminescent reagent is added to the sample” to remove light stored in the container 2, and calculates a value related to the amount of the biological substance.
The “light intensity signal obtained after the luminescent reagent is added to the sample” is an integrated average signal as an average value of integrated signals for a predetermined time (for example, several seconds to several tens of seconds) from a time at which the luminescent reagent is introduced. The “light intensity signal obtained before the luminescent reagent is added” is an integrated average signal as an average value of integrated signals for a predetermined time (for example, several seconds to several tens of seconds) before the luminescent reagent is introduced. Here, the “light intensity signal obtained before the luminescent reagent is added” is based on the light stored in the container 2. For example, when the container 2 is placed outside the biological sample analysis device 100 before the measurement is started, light of ultraviolet rays, a fluorescent lamp, or the like is stored in the container 2 in some cases. Therefore, the calculator COM1 subtracts a “second light intensity signal that is a light intensity signal only from the container 2” from a “first light intensity signal including the light intensity signal derived from the container 2 and a light intensity signal of biological origin”. As a result, the biological sample analysis device 100 can accurately calculate only the light intensity signal of biological origin. The same applies to signal processing in the blank measurement and the standard solution measurement. The light intensity signal is not limited to the integrated average signal, and may be a simple integrated signal for a predetermined time (for example, several seconds to several tens of seconds) from the time at which the luminescent reagent is introduced, or may be a signal subjected to other arithmetic processing.
In this manner, the calculator COM1 of the controller COM calculates ATP [amol] for each of the containers 2 by the following equation (see
Samplesignal is a signal obtained from the sample measurement, and is a signal obtained by subtracting the “light intensity signal obtained before the luminescent reagent is added to the sample” from the “light intensity signal obtained after the luminescent reagent is added to the sample”.
STDsignal is a signal obtained from the standard solution measurement, and is a signal obtained by subtracting the “light intensity signal obtained before the luminescent reagent is added to the standard solution” from the “light intensity signal obtained after the luminescent reagent is added to the standard solution”.
Zerosingle is a signal obtained from the blank measurement, and is a signal obtained by subtracting the “light intensity signal obtained before the luminescent reagent is added to the zero solution” from the “light intensity signal obtained after the luminescent reagent is added to the zero solution”. In
By the above calculations, variations in stored light amounts of the container 2s for standard solution measurement, the container 2b for blank measurement, and the container 2 for sample measurement can be removed, and the ATP amount can be accurately calculated.
After measurement of luminescence of one container 2 is completed, the controller COM causes the lifting device 14 to move the movable-side light shield 132 of the light shielding mechanism 13 to the retraction position, and then causes the holder drive mechanism 5 to move the container 2 to be measured next to the detection position Xdet. In this way, the luminescence of the sample in each container 2 is sequentially measured. Here, the pipette tip PT is replaced each time the luminescence of the sample in each container 2 is measured, and the used pipette tip PT is discarded in the disposal box 10.
After the measurement is completed for all the samples in this manner, the door C2 is opened to replace the specimen tubes FC held by the temperature control mechanism 7, and the containers 2 held by the holder 3 are replaced. Here, when the containers 2 held by the holder 3 are replaced, the holder 3 is removed from the device body by holding the holding holes 3h of the holder 3. Since the holder 3 includes the used and discarded pipette tips PT in the disposal box 10 of the holder 3, the discarded pipette tips PT can also be taken out from the device body at the same time by detaching the holder 3 from the device body.
Next, using a container irradiated with ultraviolet rays and a container not irradiated with ultraviolet rays, ATP solutions adjusted to 0, 1, 2, 4, 10, and 20 amol/μL are measured using the biological sample analysis device 100.
It can be seen that the luminescence amount in dark measurement under influence of stored light (light intensity signal obtained before the luminescent reagent is added) is larger in the container irradiated with ultraviolet rays. Further, it can be seen that the luminescence amount in peak measurement under influence of stored light (light intensity signal obtained after the luminescent reagent is added) is also larger in the container irradiated with ultraviolet rays.
Meanwhile, by subtracting the luminescence amount of the dark measurement from the luminescence amount of the peak measurement, the luminescence amount of the container irradiated with ultraviolet rays and the luminescence amount of the container not irradiated with ultraviolet rays are substantially matched. Therefore, it can be seen that the ATP amount can be calculated by removing the stored light amount of the container by the above calculation method.
<Effects of Embodiment>
In the biological sample analysis device 100 according to the embodiment configured as described above, the light intensity signal obtained before the luminescent reagent is added is subtracted from the light intensity signal obtained after the luminescent reagent is added to the sample to calculate the value related to the amount of the biological substance, and thus, there is no need to detect the “luminescence intensity in a state where the luminescence is reduced” as in the conventional art. As a result, measurement time of the sample can be shortened. Further, since the light stored in the container 2 is removed by subtracting the light intensity signal obtained before the luminescent reagent is added from the light intensity signal obtained after the luminescent reagent is added to the sample, there is no need to consider the light stored in the container 2 storing the sample, and the measurement accuracy can be improved.
Note that the present invention is not limited to the embodiment.
For example, the holder 3 holds the plurality of containers 2 in a circular shape, but may hold the plurality of containers 2 in an annular shape such that the plurality of containers 2 are arranged, for example, in a rectangular shape, a polygonal shape, or an elliptical shape.
In the embodiment, the disposal box 10 has two openings 10x, but may have one or three or more openings 10x.
As illustrated in
Further, the disposal box 10 may have a slope 10y that inclines each pipette tip PT discarded in each disposal space 10s in a predetermined direction. The slope 10y is inclined such that upper ends of the pipette tips PT discarded in the disposal spaces 10s do not interfere with each other. In the example in
In the embodiment, the holder 3 and the disposal box are integrally formed. However, the holder 3 and the disposal box 10 may be separately provided.
In the embodiment, the luminescent reagent is added to the container storing the biological sample, but the biological sample may be added to the container storing the luminescent reagent.
In addition, bacterial species contained in a sample whose ATP is measured by the biological sample analysis device according to the embodiment may be identified.
Specifically, as shown in
As pretreatment for analysis with a DNA sequencer, amplification by a DNA amplification method (PCR) is considered. Here, the residual liquid is collected using, for example, DNA collecting beads, and the collected DNA is amplified by PCR. The residual liquid contains an ATP extract. As the ATP extract, for example, a surfactant, a mixed solution of ethanol and ammonia, methanol, ethanol, trichloroacetic acid, perchloric acid, a Tris buffer solution, or the like can be suitably used. Examples of the surfactant include sodium dodecyl sulfate, potassium lauryl sulfate, sodium monolauroyl phosphate, sodium alkylbenzene sulfonate, benzalkonium chloride, benzethonium chloride, cetylpyridinium chloride, cetyltrimethylammonium bromide, and myristyl dimethylbenzylammonium chloride. Some ATP extracts inhibit action of enzymes that break down DNA. As for a sample after ATP measurement, there is a possibility that the ATP extract inactivates the enzyme of PCR, and thus pretreatment of removing the ATP extract may be performed.
Furthermore, it is also possible to measure an ATP amount of bacteria in a spore state (spore bacteria) by the biological sample analysis device according to the embodiment. That is, the ATP amount of the spore bacteria can be measured by subtracting the ATP amount before the spore bacteria germinate from the ATP amount after the spore bacteria germinate.
Specifically, an ATP amount of only bacteria in a normal state before the spore bacteria germinate (viable bacteria) (Y [amol]) can be measured by performing ATP measurement without putting the spore reaction solution into the sample or before the spore bacteria germinate in a case where the spore reaction solution has been put into the sample. An ATP amount of both the spore bacteria and the viable bacteria (X [amol]) can be measured by performing ATP measurement after the spore reaction solution of the sample is added to germinate the spore bacteria. Then, the ATP amount of the spore bacteria can be calculated by X-Y [amol]. A large value of X-Y indicates generation of spore bacteria, and a user can take measures such as cleaning with a sporocide. Further, viable bacteria in the sample are killed using a certain method (for example, a heat shock method). Then, the ATP amount of the spore bacteria can also be measured by germinating the spore bacteria by heating or germinating the spore bacteria by adding nutrients.
The present invention is not limited to the embodiment, and it goes without saying that various modifications can be made without departing from the gist of the present invention.
The present invention can shorten the measurement time of the sample and improve the measurement accuracy.
| Number | Date | Country | Kind |
|---|---|---|---|
| 2019-123882 | Jul 2019 | JP | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/JP2020/019535 | 5/15/2020 | WO | 00 |
