JP2010190796A - Background light reducing member and light measuring method - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a background light reducing member and a light measuring method for reducing effectively background light generated in liquid such as a buffer existing around a measuring object without exerting an influence on the measuring object, even when a container has a small size, and dispensing with replacement of the buffer. <P>SOLUTION: This background light reducing member 1 is a member used for reducing background light generated in the buffer B around the measuring object S, when measuring through a bottom part of the container 40, light emitted from the measuring object S arranged in the buffer B in the bottomed container 40 by taking a fluorescent or light-emitting material. The member includes a hydrophobic contact part 10 inserted into the container 40 to be in contact with the buffer B and arranged so as to enclose a prescribed axis C along an insertion direction, and reduces background light owing to contact between the buffer B and the contact part 10 in the container 40. <P>COPYRIGHT: (C)2010,JPO&amp;INPIT

Description

  The present invention relates to a background light reducing member and a light measurement method.

  Conventionally, fluorescence measurements are used for screening compound libraries in drug discovery. In such fluorescence measurement, first, a measurement object such as a cell is cultured and placed on the bottom of a transparent bottomed container such as a petri dish or a Bayer bottle. Then, a buffer (buffer solution, external solution) containing a fluorescent indicator is additionally injected. If this is the case, the fluorescence of the fluorescent indicator that has not been absorbed by the cells or the like becomes background light, making it difficult to identify and measure the fluorescence from the measurement object at the bottom of the container. Washing (buffer replacement, washout) is repeated by removing the buffer by suction and adding a buffer containing no fluorescent indicator. Thereafter, screening can be performed by introducing a compound (reagent) or the like to be screened and measuring the fluorescence state of the measurement object through the bottom of the container.

  In the fluorescence measurement as described above, many compounds to be screened have autofluorescence. When a compound having autofluorescence is dispensed and tested, the measured fluorescence value is derived from a measurement object such as a cell or from the autofluorescence of the compound in the buffer (ie, background light). It is difficult to judge. As a result, there arises a problem that false positives increase. In particular, in screening of high-throughput compounds, it causes a decrease in detection accuracy and further a decrease in throughput.

  In order to solve the problem of autofluorescence of such a compound, for example, a method of reducing the influence of autofluorescence of a compound by mixing a black dye or various kinds of dyes in a buffer has been put into practical use (for example, a patent (See FIG. 2 of Document 1). Alternatively, polymer latex beads, inorganic particles, etc. are added to the buffer, and are applied to the object to be measured and laminated to form an optical separation layer, thereby covering the object to be measured from above and thereby from the separation layer. A method for shielding the background fluorescence derived from the upper buffer has also been proposed (see, for example, FIG. 6 of Patent Document 1).

  Further, in Patent Document 2, after adding a buffer containing a fluorescent indicator to a transparent container in which a measurement object such as a cell is accommodated, a masking member having light-shielding properties and liquid permeability is added to the measurement object in the transparent container. A method is disclosed in which the background light that is going to travel from the buffer above the measurement object to the bottom of the transparent container is shielded by being arranged at the top.

Japanese Patent No. 345068 International Publication No. 2005/095929 Pamphlet

  However, among the methods described in Patent Document 1, in the method of mixing a black pigment or the like in a buffer, the black pigment or the like itself affects the function of the cell that is the measurement target and causes inhibition of various reactions. In some cases, it is difficult to support all measurement systems. In addition, in the case of quenching with a dye, the absorption band varies depending on the wavelength. Therefore, it is essential to select an optimum dye depending on the type of fluorescent indicator and the measurement system.

  On the other hand, in the method of adding polymer latex beads or the like in a buffer and laminating them on the object to be measured to form an optical separation layer, particles such as polymer latex beads directly contact the cells that are the object to be measured. Doing so may cause adverse effects on the cells. Further, when the reagent is further dispensed after the separation layer is formed with polymer latex beads or the like, craters are generated in the separation layer due to the discharge pressure of the pipette, etc., and the separation layer is disturbed, resulting in variations in the thickness of the separation layer. As a result, there is a possibility that high-resolution detection of fluorescence derived from cells or the like cannot be performed.

  Further, according to the inventor's research on the method described in the cited document 2, a remarkable effect can be obtained when the inner diameter of the transparent container is relatively large, but the effect gradually decreases as the inner diameter of the transparent container decreases. It has been found. Possible causes are as follows. Usually, since the location where the measurement object is arranged in the transparent container needs to be hydrophilic, the inner surface of the transparent container is subjected to a hydrophilic treatment. For this reason, a concave distortion (meniscus) due to surface tension is formed on the liquid surface in the transparent container. The fluorescence or the like generated in the liquid such as the buffer is reflected / scattered by the concave distortion, and part of the fluorescence proceeds to the bottom of the transparent container. And the smaller the inner diameter of the transparent container, the more noticeable the distortion of the liquid level in the transparent container, and the amount of background light at the bottom of the container increases.

  In addition, the conventional method for performing buffer replacement is time-consuming and causes a reduction in throughput. Moreover, there is also a problem that a measurement object such as a cell is detached from the bottom of the container by repeatedly replacing the buffer in the container.

  The present invention has been made in view of the above-described problems, and background light generated in a liquid such as a buffer around the measurement object does not affect the measurement object, and the container has a small diameter. Even so, an object of the present invention is to provide a background light reducing member and a light measurement method that can be effectively reduced and further eliminate the need for buffer replacement.

  In order to solve the above-described problems, the background light reducing member according to the present invention emits light emitted when a measurement object arranged in a liquid in a bottomed container takes in a fluorescent or luminescent substance. This is a member used to reduce the background light generated in the liquid around the measurement object when measuring through the bottom of the object, and is inserted into the bottomed container to come into contact with the liquid in the bottomed container And having a hydrophobic contact portion arranged so as to surround a predetermined axis along the insertion direction, and reducing background light due to the contact between the liquid in the bottomed container and the contact portion. It is characterized by.

  Further, the light measurement method according to the present invention is a method of measuring light emitted by a measurement object placed in a liquid in a bottomed container through taking in a fluorescent or luminescent substance through the bottom of the bottomed container. The contact portion of the member having the hydrophobic contact portion arranged so as to surround the predetermined axis is inserted into the bottomed container along the predetermined axis and brought into contact with the liquid in the bottomed container. Thus, the background light generated in the liquid around the measurement object is reduced.

  As described above, since the inner surface of the bottomed container is subjected to hydrophilic treatment, the liquid surface is distorted into a concave shape due to the surface tension of the buffer or the like injected into the bottomed container. It is considered that the amount of background light at the bottom of the container increases due to the scattering action. In response to such a phenomenon, the present inventors have found that the shape of the liquid surface can be improved by bringing a hydrophobic member into contact with the liquid surface in the bottomed container. Then, the background light reaching the bottom of the bottomed container by making this member a specific shape, that is, a shape arranged so as to surround a predetermined axis along the insertion direction when inserted into the bottomed container. Has been found to be effectively reduced. This is because when the hydrophobic member having the above shape is brought into contact with the liquid surface, the distortion of the liquid surface in the bottomed container becomes flat or convex, and the fluorescence generated in the liquid penetrates the liquid surface. This is considered to be released to the outside.

  Thus, according to the background light reducing member and the light measurement method described above, the background light can be effectively reduced even when the inner diameter of the bottomed container is small. In addition, since the background light can be effectively reduced without the member itself coming into contact with the measurement object such as a cell, it is highly sensitive to eliminate the influence of the background light without affecting the measurement object such as the cell. Measurement is possible. And it can respond to any measurement regardless of the type (fluorescence wavelength) of the fluorescent indicator and the measurement system. In addition, by simply inserting the background light reducing member described above into the bottomed container, it is possible to reduce the background light caused by excessive fluorescent indicator in the liquid, thus eliminating the need for buffer replacement and improving throughput. Can be planned.

  The background light reducing member described above is used when using a microplate having a plurality of wells as a bottomed container, and may have a plurality of contact portions corresponding to the plurality of wells. In addition, the above-described optical measurement method may use a microplate having a plurality of wells as a bottomed container, and the member may have a plurality of contact portions corresponding to the plurality of wells. As a result, even when a measurement object is arranged in a plurality of wells of a microplate and a plurality of measurements are performed simultaneously, the background light of each well can be reduced collectively, and the throughput can be further improved. .

  Further, the background light reducing member and the light measuring method described above may be characterized in that the shape of the contact portion in the plane intersecting the predetermined axis is a shape along the inner surface of the bottomed container. When the contact portion has such a shape, the shape of the liquid surface in the bottomed container can be improved over a wide range, and the background light can be more effectively reduced.

  Further, the background light reducing member and the light measurement method described above may be characterized in that the shape of the contact portion is a cylindrical shape extending in the direction of a predetermined axis. Alternatively, the contact portion may be composed of a plurality of hydrophobic portions arranged around a predetermined axis at intervals. Even if the contact portion has any of these shapes, the above-described effects can be suitably obtained.

  In the background light reducing member and the light measurement method described above, the hydrophobic contact portion is in contact with the liquid in the bottomed container. (1) The tip of the hydrophobic contact portion is in contact with the liquid surface. (When the contact part itself is not inside the liquid) (2) The part including the contact part is inserted into the liquid, and the part in contact with the liquid surface in the part is hydrophobic (ie, the contact part) And the case where it is.

  According to the background light reducing member and the light measurement method of the present invention, background light generated in a liquid such as a buffer around the measurement object is effectively reduced without affecting the measurement object, Buffer replacement can be eliminated.

(A) It is a perspective view of the background light reduction member based on 1st Embodiment of this invention. (B) It is a longitudinal cross-sectional view of a background light reduction member. 3 is a longitudinal sectional view showing a usage state of the background light reducing member 1. FIG. It is a figure for demonstrating the effect | action of the background light reduction member 1, and has shown notionally the mode in the container in the state which does not use the background light reduction member 1. FIG. It is a figure for demonstrating the effect | action of the background light reduction member 1, and has shown notionally the mode of the change of the liquid level at the time of inserting the background light reduction member 1 in a container. 6 is a photograph showing the effect of the background light reducing member 1. (A) It is a photograph of the bottom of the container 40 when the background light reducing member 1 is not used. (B) A photograph of the bottom of the container 40 when the background light reducing member 1 is used. (A) It is a whole perspective view of the background light reduction member based on 2nd Embodiment of this invention. (B) It is an enlarged view of a background light reduction member. (C) It is an enlarged view of a contact part periphery. (D) It is a longitudinal cross-sectional view which shows a use condition. (A)-(d) It is explanatory drawing which showed the optical measurement method which concerns on 2nd Embodiment. (A)-(d) It is explanatory drawing which shows another light measurement method which concerns on 2nd Embodiment. 6 is a photograph showing the effect of the background light reducing member 2. (A) It is the photograph of the bottom part of the microplate 50 when not using the background light reduction member 2. FIG. (B) A photograph of the bottom of the microplate 50 when the background light reducing member 2 is used for each well 55 included in the region F. 6 is a graph showing temporal changes (responses) in fluorescence intensity detected through the bottom of a well 55. FIG. It is a figure which shows the example of the planar shape (cross-sectional shape of the contact part in the plane perpendicular | vertical to the predetermined axis C) of the contact part (a)-(d). It is a photograph which shows the mode of each fluorescence when using the contact part of the shape shown to Fig.11 (a) (a) before use, (b) in use. It is a photograph which shows the mode of each fluorescence when using the contact part of the shape shown in FIG.11 (b), before (a) use (b). It is a photograph which shows the mode of each fluorescence when using the contact part of the shape shown in FIG.11 (c), before (a) use (b). (A)-(c) It is a figure which shows the other example of the planar shape of a contact part.

  Hereinafter, embodiments of a background light reducing member and a light measurement method according to the present invention will be described in detail with reference to the accompanying drawings. In the description of the drawings, the same elements are denoted by the same reference numerals, and redundant description is omitted.

(First embodiment)
FIG. 1A is a perspective view of a background light reducing member according to an embodiment of the present invention, and FIG. 1B is a longitudinal sectional view thereof. The background light reducing member 1 of the present embodiment includes a contact portion 10. The contact portion 10 is a portion that is inserted into a bottomed container to be described later and is brought into contact with a liquid (buffer) in the container. The contact portion 10 has a shape like a frame in which a wall surface is disposed so as to surround a predetermined axis C along the direction of insertion into the container (arrow A in the figure). It has a cylindrical shape extending in the direction. Moreover, the shape of the contact part 10 in the plane which cross | intersects the predetermined axis C is a shape along the inner surface of a container, Preferably it is a shape which can be fitted in the inner surface of a container. The contact portion 10 has hydrophobicity and non-absorbability, and the material is preferably polystyrene, polypropylene, or polyethylene. In order to shield and extinguish the light, the background light reducing member 1 is preferably black.

  FIG. 2 is a longitudinal sectional view showing a usage state of the background light reducing member 1. As shown in FIG. 2, a measurement object S such as cultured cells or coated antibodies is arranged at the bottom of the container 40 having a transparent bottom. The container 40 is filled with a liquid (buffer) B containing a fluorescent or luminescent substance such as a fluorescent indicator, and the measurement object S takes in the substance (fluorescent indicator) in the buffer B. When the excitation light E is irradiated through the reflecting mirror 43, the fluorescent indicator in the measurement object S is excited by the excitation light E and emits fluorescence L having a predetermined wavelength. In addition, compound (reagent) D to be screened is also put into buffer B. And the contact part 10 of the background light reduction member 1 which concerns on this embodiment is inserted in the container 40. FIG. In this state, the fluorescence L emitted from the measuring object S passes through the transparent bottom of the container 40 and then passes through the reflecting mirror 43 and is imaged by an imaging device (not shown) disposed below the reflecting mirror 43. Is measured.

  Here, with reference to FIG.3 and FIG.4, the effect | action of the background light reduction member 1 of this embodiment is demonstrated. FIG. 3 conceptually shows the inside of the container in a state where the background light reducing member 1 is not used, and FIG. 4 shows how the liquid level changes when the background light reducing member 1 is inserted into the container. Is shown conceptually.

  Usually, since the location where the measuring object S is arranged on the inner surface of the container 40 needs to be hydrophilic, the inner surface of the container 40 is subjected to a hydrophilic treatment. Therefore, as shown in FIG. 3, a concave distortion (meniscus) due to surface tension is formed on the liquid surface of the buffer B in the container 40.

  On the other hand, the fluorescence emitted from the fluorescent indicator that has not been taken into the measurement object S is reflected and scattered on the concave liquid surface, and most of the fluorescence proceeds to the bottom of the container 40. In addition, many compounds (reagents) D to be screened have autofluorescence, and when a compound having such autofluorescence is dispensed and tested, the fluorescence from the compound D is also in buffer B. Reflected and scattered on the liquid surface, most of which proceeds to the bottom of the container 40. Therefore, these lights become background light and are output through the bottom of the container 40. In addition, since the distortion of the liquid level in the container 40 becomes more remarkable as the inner diameter of the container 40 is smaller, such a phenomenon appears remarkably.

  On the other hand, as shown in FIG. 4, when the hydrophobic contact portion 10 is brought into contact with the liquid surface of the buffer B, the distortion of the liquid surface of the buffer B becomes flat or convex. Such an action is considered to be caused by the shape of the contact portion 10 of the background light reducing member 1 according to the present embodiment, that is, the shape of a frame in which a wall surface is disposed so as to surround the predetermined axis C. Due to such an action, the fluorescence generated in the buffer B solution, that is, the fluorescence emitted from the fluorescent indicator that has not been taken into the measurement object S and the autofluorescence of the compound D are transmitted through the liquid surface of the buffer B to the outside. Will be released. For this reason, the fluorescence reaching the bottom of the container 40 is reduced, and the background light is effectively reduced.

  Such an effect becomes more remarkable when the inner diameter of the container 40 is small. The concave distortion of the buffer B liquid surface due to the surface tension described above is formed in the vicinity where the inner surface of the container 40 and the liquid surface of the buffer B are in contact with each other. This is because the dent in the liquid level in the vicinity of the center of the container, which is the region, becomes significant.

  Thus, according to the background light reducing member 1 of the present embodiment and the light measurement method using the same, the background light can be effectively reduced even when the inner diameter of the container 40 is small. Therefore, in the fluorescence measurement through the bottom of the container 40, only the fluorescence L from the measurement object S can be measured without being disturbed by the background light, and highly accurate fluorescence measurement with few false positives is possible. Become. In addition, since the background light reducing member 1 does not need to be in contact with the measurement object S, high-sensitivity measurement that eliminates the influence of the background light without affecting the measurement object S is possible. And it can respond to any measurement regardless of the type (fluorescence wavelength) of the fluorescent indicator and the measurement system. Further, since the background light caused by the excessive fluorescent indicator in the buffer B solution can be reduced simply by inserting the contact portion 10 of the background light reducing member 1 into the container 40, the replacement of the buffer B becomes unnecessary. Throughput can be improved.

  FIG. 5 is a photograph showing the effect of the background light reducing member 1 of the present embodiment. 5A is a photograph of the bottom of the container 40 when the background light reducing member 1 is not used (see FIG. 3), and FIG. 5B is a case where the background light reducing member 1 is used (FIG. 5). 4 is a photograph of the bottom of the container 40 in FIG. In these photographs, CHO cells are arranged as the measurement object S at the bottom of the container 40, stained with Fluo4 (Ca ion fluorescent indicator), and 1 μM FITC (Fluorescein Isothiocyanate) is used as buffer B. As a result, the background light is increased.

  As shown to Fig.5 (a), when the background light reduction member 1 is not used, the whole bottom part becomes bright with background light, and it becomes difficult to distinguish the fluorescence from a CHO cell. On the other hand, as shown in FIG. 5B, when the background light reducing member 1 is brought into contact with the liquid surface of the FITC, only the background light from the FITC is removed and the fluorescence from the CHO cells can be clearly observed. .

  As described above, the shape of the contact portion 10 in the plane intersecting the predetermined axis C is preferably a shape along the inner side surface of the container 40. When the contact portion 10 has such a shape, the shape of the liquid surface of the buffer B can be improved over a wide range, and the background light can be more effectively reduced.

(Second Embodiment)
6A is an overall perspective view of the background light reducing member according to the second embodiment of the present invention, FIG. 6B is an enlarged view thereof, and FIG. 6C is an enlarged view of the vicinity of the contact portion. FIG. 6D is a longitudinal sectional view showing the use state.

  The background light reducing member of the second embodiment is different from the first embodiment in that it is used for a microplate having a plurality of wells as a bottomed container.

  As shown in FIG. 6A to FIG. 6C, the background light reducing member 2 of the present embodiment is used for a microplate having a plurality of wells for accommodating a measurement object. A plurality of contact portions 10 corresponding to a plurality of wells of the plate are provided. The plurality of contact portions 10 are supported by a sheet-like portion 21 having an integral structure that covers the upper surface of the microplate, and are fitted into the upper surface shape of the microplate to be inserted into each well of the microplate.

  As shown in FIG. 6D, in the usage state of the background light reducing member 2 of the present embodiment, the measurement object S such as cultured cells is placed on the transparent bottom of the well 55 provided in the microplate 50. Is placed. In each well 55, a liquid (buffer) B containing a fluorescent or luminescent substance such as a fluorescent indicator is introduced, and the measurement object S takes in the substance (fluorescent indicator) in the buffer B. And when excitation light is irradiated through the reflecting mirror which is not shown in figure, the fluorescence indicator in the measuring object S is excited by this excitation light, and the fluorescence L of a predetermined wavelength is emitted. In the buffer B, a compound (reagent) to be screened is also charged. The background light reducing member 2 is inserted into the microplate 50 so that the contact portions 10 and the wells 55 are fitted to each other. The contact portion 10 is positioned at a predetermined position in the well 55 by fitting the sheet-like portion 21 having an integral structure so as to cover the upper portion of the microplate 50.

  Next, the light measurement method using the background light reducing member 2 will be specifically described. 7A to 7D are explanatory views showing a light measurement method according to the present embodiment. In this method, first, as a first step, the measuring object S is placed in the form of cultured cells or coated antibodies on the transparent bottom of each well 55 of the microplate 50 (FIG. 7A). Next, as a second step, buffer B containing a fluorescent indicator is added to each well 55 (FIG. 7B). At this time, a concave distortion due to the surface tension is generated on the liquid surface of the buffer B. Then, as a third step, the background light reducing member 2 is inserted and fitted into the microplate 50 without removing the buffer B containing the fluorescent indicator (without washing), and each contact portion 10 is connected to each well. It inserts in 55 (FIG.7 (c)). Thereby, each contact part 10 contacts the liquid level of the buffer B, the shape of the liquid level is controlled, and the fluorescence emitted from the buffer B is emitted from the liquid level. Thereafter, as a fourth step, the fluorescence L from the measuring object S is observed through the bottom of each well 55 (FIG. 7D). In addition, the reagent etc. which are the object of a screening can be thrown into the buffer B suitably.

  8A to 8D are explanatory views showing another light measurement method according to this embodiment. In this method, as a first step, the measuring object S is placed in each well 55 of the microplate 50 (FIG. 8A). As a second step, the background light reducing member 2 is inserted and fitted into the microplate 50, and each contact portion 10 is inserted into each well 55 (FIG. 8B). Then, as a third step, buffer B containing a fluorescent indicator is added to each well 55 (FIG. 8 (c)). Thereafter, as a fourth step, the fluorescence L from the measurement object S is observed through the bottom of each well 55 without removing the buffer B containing the fluorescent indicator (without washing) (FIG. 8 ( d)).

  In the present embodiment, similarly to the first embodiment, the fluorescence L from the measuring object S in each well 55 can be measured through the transparent bottom of the well 55. When the hydrophobic contact portion 10 is brought into contact with the liquid surface of the buffer B, the distortion of the liquid surface of the buffer B becomes flat or convex. Therefore, the fluorescence generated in the buffer B liquid is transmitted through the buffer B liquid surface and released to the outside. For this reason, the fluorescence reaching the bottom of the well 55 is reduced, and the background light is effectively reduced.

  Thus, according to the background light reducing member 2 of the present embodiment and the light measurement method using the same, the background light can be effectively reduced. Therefore, in the fluorescence measurement through the bottom of the well 55, only the fluorescence L from the measurement object S can be measured without being disturbed by the background light, and highly accurate fluorescence measurement with few false positives is possible. Become. In addition, since the background light reducing member 2 does not have to be in contact with the measurement object S, high-sensitivity measurement that eliminates the influence of the background light without affecting the measurement object S is possible. And it can respond to any measurement regardless of the type (fluorescence wavelength) of the fluorescent indicator and the measurement system. Further, since the background light caused by the excessive fluorescent indicator in the buffer B solution can be reduced only by inserting the contact portion 10 into the well 55, the replacement of the buffer B becomes unnecessary and the throughput is improved. be able to.

  As the microplate 50 used in the present embodiment, for example, a 96-well type in which wells 55 are arranged in an 8 × 12 grid, and a well 55 having a half size of the 96-well type is a 16 × 24 grid. There are a 384 well type arranged in a line and a 1536 well type in which wells of half the size of the 384 well type are arranged in a 32 × 48 lattice. In recent years, there is a tendency to use a microplate 50 having a larger number of wells 55 (that is, a smaller well size) in order to improve measurement efficiency.

  In particular, when a 384-well type microplate is used, the well size is as small as 3.5 mm or less, and the influence of the liquid level distortion is increased in the conventional method. Therefore, the background light reducing member according to the present embodiment 2 and the effect of the light measurement method using the same are more remarkable.

  FIG. 9 is a photograph showing the effect of the background light reducing member 2 of the present embodiment. FIG. 9A is a photograph of the bottom of the microplate 50 when the background light reducing member 2 is not used, and FIG. 9B shows the background light reducing member 2 for each well 55 included in the region F. It is the photograph of the bottom part of the microplate 50 at the time of using. In these photographs, a 384-well type is used as the microplate 50, CHO cells are placed as the measurement object S at the bottom of the well 55, this is stained with Fluo4, and 1 μM FITC is used as the buffer B It is a thing.

  As shown in FIG. 9A, when the background light reducing member 2 is not used, the entire bottom is brightened by the background light, making it difficult to distinguish the fluorescence from the CHO cells. On the other hand, when the background light reducing member 2 is brought into contact with the liquid surface of the FITC as shown in a region F of FIG. 9B, the background light from the FITC can be effectively reduced.

  FIG. 10 is a graph showing a temporal change (response) in fluorescence intensity detected through the bottom of the well 55. The vertical axis of this graph shows the relative value of light intensity, and the horizontal axis is time. In FIG. 10, the graph group G is a graph before the background light reducing member 2 is inserted into the microplate 50, and the graph group H is a graph after the background light reducing member 2 is inserted into the microplate 50. As shown in the graph group G, when the background light reducing member 2 is not used, the background light is mainly detected and the fluorescence intensity from the CHO cells cannot be measured. On the other hand, as shown in the graph group H, when the background light reducing member 2 is used, the background light is reduced and the temporal change in the fluorescence intensity from the CHO cells can be clearly detected.

(Modification)
In each said embodiment, the shape of the contact part 10 should just be a shape like the frame by which the wall surface is arrange | positioned so that the predetermined axis line C may be enclosed, and various deformation | transformation are possible. FIG. 11 is a diagram illustrating an example of a planar shape of the contact portion (a cross-sectional shape of the contact portion in a plane perpendicular to the predetermined axis C). FIGS. 12 to 14 are photographs showing the state of the fluorescence when the contact portions having the respective shapes shown in FIG. 11 are used (a) before use and (b) during use.

  FIG. 11A shows a cylindrical shape extending along a predetermined axis C, like the contact portion 10 of the above embodiment. Such a shape is the shape that is most easily created when a contact portion is created using a mold. When such a contact portion 10 is used, as can be seen from a comparison between FIG. 12 (a) and FIG. 12 (b), the background light is sufficiently reduced, and the reduction effect is different from other shapes. Highest in comparison. In addition, there is an advantage that when the reagent is dispensed using a dispenser, the reagent is uniformly poured onto the measurement object S.

  FIG. 11B shows the contact portion 11 having a shape in which a notch along a predetermined axis C is formed in two opposing portions of the cylindrical shape. The contact portion 11 includes two arc-shaped hydrophobic portions 11a and 11b arranged around a predetermined axis C with a space between each other. Even when such a contact portion 11 is used, as can be seen from the comparison between FIG. 13A and FIG. 13B, the background light is sufficiently reduced, and the reduction effect is shown in FIG. ) Shape is almost the same. However, when the reagent is dispensed using a dispenser, the reagent leaks from the gap between the two hydrophobic portions 11a and 11b, so that the reagent poured into the measuring object S may be uneven. .

  FIG. 11C shows the contact portion 12 having a shape in which notches along a predetermined axis C are formed in three portions of the cylindrical shape. The contact portion 12 includes three arc-shaped hydrophobic portions 12a to 12c arranged around a predetermined axis C with a space therebetween. Even when such a contact portion 12 is used, as can be seen from the comparison between FIG. 14A and FIG. 14B, the background light is sufficiently reduced, and the reduction effect is shown in FIG. ) Shape is almost the same. However, since there are more gaps than the contact part 11 shown in FIG. 11B, there is a possibility that the unevenness of the reagent poured into the measurement object S becomes larger.

  FIG. 11 (d) shows the contact portion 13 having a shape in which notches along the predetermined axis C are formed in four portions of the cylindrical shape. The contact portion 13 includes four arc-shaped hydrophobic portions 13a to 13d arranged around a predetermined axis C with a space between each other. Even when such a contact portion 13 is used, the background light is sufficiently reduced from the results of FIGS. 12 to 14, and the reduction effect is estimated to be almost the same as the shape of FIG. .

  FIG. 15 is a diagram illustrating another example of the planar shape of the contact portion. The contact portion 14 shown in FIG. 15A has a planar shape such as a rectangular frame shape, and has a rectangular tube shape extending along the predetermined axis C. Moreover, the contact part 15 shown in FIG.15 (b) has the shape where the notch along the predetermined axis C entered the two sides which oppose in the shape shown to Fig.15 (a), and is mutually spaced apart. It consists of two U-shaped hydrophobic portions 15a and 15b arranged around a predetermined axis C with a gap therebetween. Moreover, the contact part 16 shown in FIG.15 (c) has the shape where the notch along the predetermined axis C was entered into all the four sides in the shape shown in FIG.15 (a), and it has a space | interval mutually. It consists of four dog-shaped hydrophobic portions 16a to 16d arranged around a predetermined axis C.

  Even if the contact part in this invention is a shape as shown to Fig.15 (a)-FIG.15 (c), for example, the effect similar to the said embodiment can be show | played suitably.

  The background light reducing member and the light measurement method according to the present invention are not limited to the above-described embodiments, and various other modifications are possible. For example, the compounds (reagents) described in the above embodiments are (1) those incorporated into the measurement object, (2) those that bind directly to the measurement object, (3) measurement object and ligand, etc. It may be any of those that are indirectly bound via

  DESCRIPTION OF SYMBOLS 1, 2 ... Background light reduction member, 10-16 ... Contact part, 21 ... Sheet-like part, 40 ... Container, 50 ... Microplate, 55 ... Well, B ... Buffer, C ... Predetermined axis, D ... Compound (reagent) , E: excitation light, L: fluorescence, S: measurement object.

Claims (10)

When measuring the light emitted when the measurement object arranged in the liquid in the bottomed container takes in a fluorescent or luminescent substance through the bottom of the bottomed container, A member used to reduce background light generated in the liquid,
Having a hydrophobic contact portion that is inserted into the bottomed container and is in contact with the liquid in the bottomed container, and is disposed so as to surround a predetermined axis along the insertion direction;
A background light reducing member that reduces the background light due to the contact between the liquid in the bottomed container and the contact portion. Used when using a microplate having a plurality of wells as the bottomed container,
The background light reducing member according to claim 1, comprising a plurality of the contact portions corresponding to the plurality of wells.   The background light reducing member according to claim 1 or 2, wherein the shape of the contact portion in a plane intersecting the predetermined axis is a shape along an inner surface of the bottomed container.   The background light reducing member according to any one of claims 1 to 3, wherein a shape of the contact portion is a cylindrical shape extending in a direction of the predetermined axis.   The background light reducing member according to claim 1, wherein the contact portion includes a plurality of hydrophobic portions arranged around the predetermined axis at intervals. . A method for measuring light emitted by a measurement object disposed in a liquid in a bottomed container through taking in a fluorescent or luminescent substance through the bottom of the bottomed container,
The contact portion of a member having a hydrophobic contact portion arranged so as to surround a predetermined axis is inserted into the bottomed container along the predetermined axis and brought into contact with the liquid in the bottomed container. Then, the light measurement method characterized by reducing the background light which arises in the liquid around the said measurement object. While using a microplate having a plurality of wells as the bottomed container,
The light measurement method according to claim 6, wherein the member has a plurality of the contact portions corresponding to the plurality of wells.   The light measurement method according to claim 6 or 7, wherein a shape of the contact portion in a plane intersecting the predetermined axis is a shape along an inner surface of the bottomed container.   The optical measurement method according to claim 6, wherein the shape of the contact portion is a cylinder extending in the direction of the predetermined axis.   9. The light measurement method according to claim 6, wherein the contact portion includes a plurality of hydrophobic portions arranged around the predetermined axis at intervals.