REACTION VESSEL AND FLUORESCENCE MEASURING APPARATUS USING THE REACTION VESSEL

Abstract

A reaction vessel has plural wells each configured to accommodate a sample on the same plane or on the same straight line. Among the wells of a well plate, a specific well other than a well at a center of point symmetry is an orientation recognizing well in which a fluorescent dye is placed. The reaction vessel can identify a position of the orientation recognizing well by using fluorescence emitted from the fluorescent dye, and can thereby recognize an orientation of the reaction vessel.

Claims

1. A reaction vessel in which a plurality of wells for accommodating a sample are provided on a same plane or on a same straight line, wherein, the reaction vessel comprises an orientation recognizing well which is a well other than a well provided at a center of point symmetry among the plurality of wells, a fluorescent dye is accommodated in the orientation recognizing well.

2. The reaction vessel according to claim 1, wherein the plurality of wells are arranged in matrix in the plane, and wherein the orientation recognizing well is a well provided at a corner or near the corner of the plane.

3. A fluorescence measuring apparatus, comprising: a reaction vessel setting part on which the reaction vessel according to claim 1 is set; and a measurement part configured to measure an intensity of light of a measurement target wavelength from inside of each of the wells of the reaction vessel set on the reaction vessel setting part, and to detect fluorescence emitted from the fluorescent dye in the orientation recognizing well of the reaction vessel.

4. The fluorescence measuring apparatus according to claim 3, comprising a measurement value correction part configured to correct a measurement value for the orientation recognizing well based on a difference between a baseline of the measurement value for the orientation recognizing well of the reaction vessel measured by the measurement part and baselines of measurement values for the other wells measured by the measurement part when a wavelength of the fluorescence emitted from the fluorescent dye of the orientation recognizing well of the reaction vessel is the same as the measurement target wavelength.

5. The fluorescence measuring apparatus according to claim 3, wherein the wavelength of the fluorescence emitted from the fluorescent dye in the orientation recognizing well of the reaction vessel is different from the measurement target wavelength.

6. The fluorescence measuring apparatus according to claim 3, further comprising a well position identification part configured to identify a position of the orientation recognizing well based on measurement values of each of the wells obtained by the measurement part.

7. The fluorescence measuring apparatus according to claim 6, further comprising a well position determination part configured to determine whether or not an actual position of the orientation recognizing well identified by the well position identification part matches with an original position set in advance for the orientation recognizing well.

8. The fluorescence measuring apparatus according to claim 7, wherein the well position determination part is configured to issue a warning to a user when the actual position identified by the well position identification part does not match with the original position.

9. The fluorescence measuring apparatus according to claim 7, further comprising: a sample information storage part configured to store sample information, which is information related to the sample accommodated in each of the wells of the reaction vessel set in the reaction vessel setting part, in association with the position of the well in which each sample is accommodated; and a correspondence relation correction part configured to, when the well position determination part determines that the actual position identified by the well position identification part does not match with the original position, correct a correspondence relation of the position of the well to the sample information of the sample information storage part based on the actual position of the orientation recognizing well identified by the well position identification part, so that the sample information stored in the sample information storage part is associated with the position of the well in which each sample is actually accommodated.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0028] FIG. 1 is a perspective view showing one embodiment of a well plate.

[0029] FIG. 2 is a plan view of the same embodiment.

[0030] FIG. 3 is a schematic cross-sectional configuration view showing one embodiment of a fluorescence measuring apparatus.

[0031] FIG. 4 is a diagram showing an example of detection signal waveforms of a normal well and a orientation recognizing well according to the same embodiment.

[0032] FIG. 5 is a flowchart showing an example of operation of the same embodiment.

[0033] FIG. 6 is a plan view of a well plate showing an arrangement of each well when the well plate is placed in a wrong orientation.

[0034] FIG. 7 is a flowchart showing another example of the operation of the same embodiment.

EMBODIMENT OF THE INVENTION

[0035] Hereinafter, one embodiment of a reaction vessel and a fluorescence measuring apparatus is described using drawings.

[0036] First, one embodiment of the reaction vessel is described with reference to FIG. 1.

[0037] A reaction vessel 2 of this embodiment formed by plural vessels 4 which are arranged in the same plane and are integrated with each other. Each vessel 4 includes a well 6 that is open at a top. The well 6 is for accommodating a sample and a reaction reagent therein. In this embodiment, the wells 6 are arranged in a matrix of 8 rows in the vertical direction and 12 columns in the horizontal direction. In order to identify a position of each well 6, a left end and a top end of an upper surface of a well plate 2 are respectively labeled with identification codes A to H indicating rows in the vertical direction and identification codes 1 to 12 indicating columns in the horizontal direction.

[0038] Among the wells 6 provided in the reaction vessel 2, a fluorescent dye 8 is accommodated in the well 6 provided at a specific position other than a center of point symmetry. In this embodiment, the fluorescent dye 8 is accommodated in the well 6 at H12 (12th from the left in the row H) located at a lower right end of the reaction vessel 2. Hereinafter, the well 6 in which the fluorescent dye 8 is accommodated is referred to as a orientation recognizing well 6s so as to be distinguished from the other wells 6. Note that the position of the orientation recognizing well 6s where the fluorescent dye 8 is accommodated may be the position of the well other than the center of point symmetry in all the wells 6.

[0039] The fluorescent dye 8 placed in the orientation recognizing well 6s is solidified by drying, for example. The fluorescent dye 8 preferably has a property of being dissolved and colored when the reaction reagent is added to the orientation recognizing well 6s. However, the fluorescent dye is not necessarily required to be colored by the addition of the reaction reagent, and one that emits fluorescence when irradiated with excitation light may be used. The fluorescent dye 8 is not necessarily be solid-phased, and may be sealed in the orientation recognizing well 6s by any means. As the fluorescent dye, SYBR (registered trademark) Green I, SYBR (registered trademark) Gold, EvaGreen (registered trademark), and the like can be used.

[0040] Because the fluorescent dye having the property of being colored when the reaction reagent is added, is fixed in the orientation recognizing well 6s, when the reaction reagent is added to the orientation recognizing well 6s, the orientation recognizing well 6s provided at the specific position is colored as shown in FIG. 2. Accordingly, an orientation of the reaction vessel 2 can be visually confirmed. When the fluorescent dye that emits fluorescence without adding the reaction reagent but by irradiation with the excitation light is used as the fluorescent dye 8, the reaction reagent is not necessarily added to the orientation recognizing well 6s, but instead, the position of the orientation recognizing well 6s can be distinguished by measuring a fluorescence intensity obtained by irradiating each well 6 with the excitation light.

[0041] Next, an embodiment of a fluorescence measuring apparatus using the reaction vessel 2 is described with reference to FIG. 3.

[0042] The fluorescence measuring apparatus includes a reaction vessel setting part 9 for setting the reaction vessel 2. The reaction vessel setting part 9 is made of heat conductive metal material, and has temperature thereof adjusted by, for example, a Peltier element or a heater. The reaction vessel setting part 9 is provided with recesses 10 each of which accommodates each vessel 4 of the reaction vessel 2.

[0043] A measurement part 12 is provided above the reaction vessel setting part 9. The measurement part 12 includes an optical sensor 14. The optical sensor 14 includes a light emitting element that emits excitation light vertically downward and a light receiving element for detecting fluorescence from the sample excited by the excitation light.

[0044] The measurement part 12 is configured to bring the optical sensor 14 at a position immediately above each well 6 so that each well 6 in the reaction vessel 2 is scanned in order during the measurement, and to measure an intensity of light of a measurement target wavelength from the sample accommodated in each well 6. In addition, the measurement part 12 has a function of detecting by the optical sensor 14, fluorescence from the fluorescent dye placed in the orientation recognizing well 6s of the reaction vessel 2. A detection signal obtained by the optical sensor 14 is taken into an arithmetic processing unit 18. The arithmetic processing unit 18 is realized by a dedicated computer or a general-purpose personal computer. A display unit 30 is connected to the arithmetic processing unit 18. The display unit 30 can be realized by a liquid crystal display, for example.

[0045] The arithmetic processing unit 18 has a function of performing various analysis processes based on the detection signal from the optical sensor 14. The analysis process performed by the arithmetic processing unit 18 includes, for example, absolute quantitative analysis for obtaining a concentration of a measurement sample, relative quantitative analysis for obtaining a relative expression level with respect to a gene, and SNP analysis for determining a genotype. A user can set a desired analysis process to be performed on the sample to be measured as a measurement condition.

[0046] Alternatively, the measurement part 12 may be provided so as to be moved in a horizontal plane below the reaction vessel setting part 9. In that case, in order to enable the measurement of the light of the measurement target wavelength from the sample accommodated in each well 6 by the optical sensor 14 of the measurement part 12, an opening for measurement is provided at a bottom part of the recess of the reaction vessel setting part 9.

[0047] The arithmetic processing unit 18 includes a measurement value correction part 20, a well position identification part 22, a well position determination part 24, a correspondence relation correction part 26, and a sample information storage part 28. The measurement value correction part 20, the well position identification part 22, the well position determination part 24, and the correspondence relation correction part 26 are functions obtained by an arithmetic element such as a CPU provided in the arithmetic processing unit 18 executing a predetermined program. The sample information storage part 28 is a function realized by a partial area of a storage unit provided in the arithmetic processing unit 18.

[0048] The measurement value correction part 20 is configured to correct a measurement value of the intensity of the light of the measurement target wavelength of the sample accommodated in the orientation recognizing well 6s of the well plate 2, when the wavelength of the fluorescence emitted from the fluorescent dye placed in the orientation recognizing well 6s of the reaction vessel 2 is the same as the measurement target wavelength. When the wavelength of the fluorescence emitted from the fluorescent dye accommodated in the orientation recognizing well 6s of the reaction vessel 2 is the same as the measurement target wavelength, and when the fluorescence intensity from each well 6 of the reaction vessel 2 is measured by the optical sensor 14, a fluorescence intensity baseline of the orientation recognizing well 6s in which the fluorescent dye is accommodated in advance becomes higher than that of the normal well 6 as shown in FIG. 4. The measurement value correction part 20 obtains a difference S between the fluorescence intensity baseline of the normal well 6 and the fluorescence intensity baseline of the orientation recognizing well 6s, and subtracts the difference S from the measurement value of the sample in the orientation recognizing well 6s, thereby correcting the measurement value of the sample in the orientation recognizing well 6s. Accordingly, the fluorescence intensity of the sample can be measured using the orientation recognizing well 6s in the same manner as the normal well 6. As the baseline of the normal well 6, for example, an average value of the baselines of all the wells 6 other than the orientation recognizing well 6s among the wells 6 provided in the reaction vessel 2 can be used.

[0049] The well position identification part 22 is configured to identify an actual position of the orientation recognizing well 6s on the reaction vessel setting part 9 by comparing the measurement values of the fluorescence intensity for respective wells 6 of the reaction vessel 2. When the wavelength of the fluorescence emitted from the fluorescent dye accommodated in the orientation recognizing well 6s of the reaction vessel 2 is the same as the measurement target wavelength, for the orientation recognizing well 6s in which the fluorescent dye is accommodated in advance, the measurement value of the intensity of the light having the measurement target wavelength is larger than the measurement value of the intensity of the light having the measurement target wavelength for the other wells 6. For this reason, by comparing the fluorescence intensities measured by the optical sensor 14 for respective wells 6, it is possible to specify which position the orientation recognizing well 6s is arranged on the reaction vessel setting part 9. In addition, when the wavelength of the fluorescence emitted from the fluorescent dye placed in the orientation recognizing well 6s of the reaction vessel 2 is different from the measurement target wavelength, the position of the orientation recognizing well 6s can be identified by searching the well 6 in which the light having the wavelength emitted from the fluorescent dye is detected.

[0050] The well position determination part 24 is configured to determine whether or not the position of the orientation recognizing well 6s identified by the well position identification part 22 is correctly arranged at a specified position. In the arithmetic processing unit 18, a specific position where the orientation recognizing well 6s is provided in the reaction vessel 2, that is, for example, the position H12 is registered in advance as the specified position. This specified position is a position where the orientation recognizing well 6s should be arranged when the reaction vessel 2 is set in the reaction vessel setting part 9 in the correct orientation. The well position determination part 24 is configured to determine whether or not the actual position of the orientation recognizing well 6s identified by the well position identification part 22 matches with the specified position.

[0051] When the reaction vessel 2 is set in the correct orientation in the reaction vessel setting part 9, the actual position of the orientation recognizing well 6s identified by the well position identification part 22 matches with the specified position. However, as shown in FIG. 6, when the orientation of the reaction vessel 2 is wrong, the orientation recognizing well 6s is arranged in a position (position A1 in FIG. 6) different from the specified position (position H12). Therefore, the actual position of the orientation recognizing well 6s identified by the well position identification part 22 does not match with the specified position. With this configuration, it is possible to automatically determine whether or not a setting orientation of the reaction vessel 2 is correct on an apparatus side.

[0052] The correspondence relation correction part 26 is configured to correct a correspondence relation between the sample information regarding each sample accommodated in each well 6 of the reaction vessel 2 and the position of the well 6 in which each sample is accommodated.

[0053] Before the measurement is started, the user registers, for example, a sample ID of the sample stored in each well 6 and a measurement condition for each sample as sample information. The sample information of each sample is registered in association with the position of the well 6 used for measuring each sample. The sample information registered in the apparatus by the user is stored in the sample information storage part 28.

[0054] However, as shown in FIG. 6, when the user sets the reaction vessel 2 in the wrong orientation in the reaction vessel setting part 9, the actual position on the reaction vessel setting part 9 for the well 6 in which each sample is accommodated becomes different from the information stored in the sample information storage part 28. By referring to FIG. 6, the well 6 provided at the position A1 of the reaction vessel 2 is arranged at the position H12 on the reaction vessel setting part 9, and the well 6 provided at the position A2 of the reaction vessel 2 is arranged at the position H11 on the reaction vessel setting part 9.

[0055] In response to this, the correspondence relation correction part 26 determines in which orientation the reaction vessel 2 is set on the reaction vessel setting part 9 based on the position of the orientation recognizing well 6s, identified by the well position identification part 22, on the reaction vessel setting part 9, and corrects the sample information of each sample stored in the sample information storage part 28 so as to associate the sample information with the actual position on the reaction vessel setting part 9 for the well 6 in which each sample is accommodated.

[0056] An example of operation of this embodiment is described with reference to FIG. 5.

[0057] First, the user inputs the sample information, and the sample information of each sample is stored in the sample information storage part 28 in association with the position of the well 6 in which each sample is accommodated (step S1). Thereafter, the user sets the reaction vessel 2 on the reaction vessel setting part 9 (step S2). This completes preparation of measurement. When the user adds the reagent to each well 6 of the well plate 2 before setting the reaction vessel 2 in the reaction vessel setting part 9, only the orientation recognizing well 6 is colored, accordingly, the user can easily recognize the orientation of the well plate 2.

[0058] When the preparation of measurement is completed, the user performs some operation to indicate the completion. Examples of the operation indicating the completion of the preparation of measurement include operation to input the completion to the arithmetic processing unit 18, and operation to close a cover (not shown) covering an upper part of the reaction vessel setting part 9, and the like. When the user performs the above operation, the well position identification part 20 recognizes that the preparation of measurement is completed, and scans all the wells 6 in the reaction vessel 2 using the optical sensor 14 of the measurement part 12, thereby identifying the position of the orientation recognizing well 6s (step S3).

[0059] After the position of the orientation recognizing well 6s is identified, the well position determination part 24 determines whether or not the position of the orientation recognizing well 6s is in the specified position (step S4), and if the position is in the specified position, the measurement is started (step S7). In the measurement, the measurement value of the sample in the orientation recognizing well 6s is corrected by the measurement value correction part 20.

[0060] If the position of the orientation recognizing well 6s is not at the specified position, the well position determination part 24 issues a warning to the user (step S5). Examples of the warning include displaying the indication on the display unit 30 connected to the arithmetic processing unit 18, generating a warning sound, and the like. It is not always necessary to give the warning to the user.

[0061] When the position of the orientation recognizing well 6s is not at the specified position, the correspondence relation correction part 26 corrects the sample information of each sample stored in the sample information storage part 28 so that the sample information is associated with the actual position on the reaction vessel setting part 9 for the well 6 in which each sample is accommodated (step S6). Thereafter, the measurement is started (step S7).

[0062] Not limited to the above, the correspondence relation correction part 26 may correct the correspondence relation between measurement data and the sample information so that the measurement data obtained by the measurement is correctly associated with the sample after the measurement is completed.

[0063] The correspondence relation correction part 26 is not an essential component. Therefore, even when the setting orientation of the reaction vessel 2 is wrong, the correspondence relationship between the sample information and the well position information may not be automatically corrected.

[0064] An example of operation of a fluorescence detection apparatus that does not include the correspondence relation correction part 26 is described with reference to FIG. 7.

[0065] Further, in the example of FIG. 7, after the user inputs sample information (step S11) and sets the reaction vessel 2 in the reaction vessel setting part 9 (step S12), the well position identification part 20 identifies the position of the orientation recognizing well 6s (step S13). After the position of the orientation recognizing well 6s is identified, the well position determination part 24 determines whether or not the position of the orientation recognizing well 6s is in a specified position (step S14), and if the orientation recognizing well 6s is in the specified position, the measurement is started (step S15).

[0066] On the other hand, if the position of the orientation recognizing well 6s is not at the specified position, the well position determination part 24 issues a warning to the user and waits without starting the measurement (step S16). The user recognizes that the setting orientation of the reaction vessel 2 is wrong by the warning, and sets the well plate 2 again (step S12). Thereafter, the well position identification part 20 identifies the position of the orientation recognizing well 6s (step S13), and the well position determination part 24 determines whether or not the position of the orientation recognizing well 6s is at the specified position (step S14), and further, if the orientation recognizing well 6s is at the specified position, the measurement is started (step S15).

DESCRIPTION OF REFERENCE SIGNS

[0067] 2: Reaction vessel [0068] 4: Vessel [0069] 6: Well [0070] 6s: Orientation recognizing well [0071] 8: Fluorescent dye [0072] 9: Reaction vessel setting part [0073] 10: Recess [0074] 12: Measurement part [0075] 14: Optical sensor [0076] 16: Guide rail [0077] 18: Arithmetic processing unit [0078] 20: Measurement value correction part [0079] 22: Well position identification part [0080] 24: Well position determination part [0081] 26: Correspondence relation correction part [0082] 28: Sample information storage part [0083] 30: Display unit