JP6827201B2 - Fluorescence detector - Google Patents

Description

The present invention relates to a fluorescence detector that detects fluorescence generated from a sample by irradiating the sample with excitation light.

In the field of biotechnology, a fluorescence detection method for detecting a biomolecule such as DNA (deoxyribonucleic acid), RNA (ribonucleic acid) or protein present in a sample is known. In this fluorescence detection method, a fluorescence detector that detects fluorescence generated from a sample by irradiating a sample labeled with a fluorescent molecule with excitation light to excite it is used.

As the above-mentioned fluorescence detector, a so-called multicolor type fluorescence detector that collectively detects a plurality of types of biomolecules labeled with a plurality of types of fluorescent molecules having different central wavelengths is known (for example, a patent). References 1 and 2). This type of fluorescence detector includes a white light source, multiple fluorescence cubes, a turret, and a motor. Each of the plurality of fluorescent cubes is a unitization of an excitation filter, a dichroic mirror, and an absorption filter. The turret is rotated in a predetermined direction around a rotation axis by a motor. Multiple fluorescent cubes are attached to the turret.

In this type of fluorescence detector, the optimum center wavelength of the excitation light to be irradiated to the sample differs depending on the type of fluorescent molecules labeled on the sample. Therefore, by appropriately rotating the turret, the fluorescent cube most suitable for the type of fluorescent molecule labeled on the sample is arranged at a predetermined position from among the plurality of fluorescent cubes. As a result, the white light from the white light source is converted into excitation light having the optimum center wavelength to be irradiated on the sample by passing through the fluorescent cubes arranged at the predetermined positions, and is irradiated on the sample.

Japanese Unexamined Patent Publication No. 8-94940 JP 2011-164409

However, since the above-mentioned conventional fluorescence detector requires a turret and a motor for rotating the turret, there arises a problem that the fluorescence detector becomes large in size.

Therefore, the present invention provides a fluorescence detector that can be miniaturized.

The fluorescence detector according to one aspect of the present invention is a fluorescence detector that detects fluorescence generated from the sample by irradiating the sample with excitation light, and is a first light emitting element that emits the first excitation light. A second light emitting element that emits a second excitation light having a center wavelength different from that of the first excitation light, and when the first light emitting element emits light, the first light emitting element from the first light emitting element. When the excitation light of 1 is reflected toward the sample and the first fluorescence from the sample is transmitted and the second light emitting element emits light, the second light emitting element from the second light emitting element emits light. When the dichroic mirror that reflects the excitation light of 2 toward the sample and transmits the second fluorescence from the sample having a center wavelength different from that of the first fluorescence and the first light emitting element emit light. , said dichroic receives dichroic mirror and the first transmission fluorescence, wherein when the second light emitting element emits light, a light emitting element for receiving the dichroic said second fluorescence transmitted through the dichroic mirror, the The first emission light is emitted so that the first excitation light and the second excitation light are sequentially emitted from the first light emitting element and the second light emitting element at the first rated output and the second rated output, respectively. The control unit includes an element and a control unit that controls the second light emitting element, and when the control unit emits the first excitation light at the first rated output from the first light emitting element. The second light emitting element emits the second excitation light at an output lower than the second rated output, and the second light emitting element emits the second excitation light at the second rated output. At that time, the first light emitting element and the second light emitting element are controlled so that the first light emitting element emits the first excitation light at an output lower than the first rated output. To do .

It should be noted that these comprehensive or specific embodiments may be realized in a recording medium such as a system, method, integrated circuit, computer program or computer-readable CD-ROM, and the system, method, integrated circuit, computer program. And any combination of recording media may be realized.

The fluorescence detector according to one aspect of the present invention can be miniaturized.

It is a figure which shows the appearance of the fluorescence detector which concerns on Embodiment 1. FIG. It is a figure which shows the structure of the fluorescence detector which concerns on Embodiment 1 simply. It is a figure which shows the transmission characteristic of the excitation filter which concerns on Embodiment 1. FIG. It is a figure which shows the transmission characteristic of the dichroic mirror which concerns on Embodiment 1. FIG. It is a figure which shows the transmission characteristic of the absorption filter which concerns on Embodiment 1. FIG. It is a flowchart which shows the operation of the fluorescence detector which concerns on Embodiment 1. FIG. It is a figure which shows the light emission sequence of each light emitting element of the fluorescence detector which concerns on Example. It is a figure which shows the light emitting condition of each light emitting element of the fluorescence detector which concerns on Example. It is a figure which shows the transmission characteristic of the dichroic mirror of the fluorescence detector which concerns on Example. It is a figure which shows the light emission sequence of each light emitting element of the fluorescence detector which concerns on a comparative example. It is a figure which shows the detection result of the fluorescence signal by the fluorescence detector which concerns on Example. It is a figure which shows the crosstalk ratio of the fluorescent signal of FIG. It is a figure which shows the detection result of the fluorescence intensity by the fluorescence detector which concerns on Example and the comparative example. It is a figure which shows the structure of the fluorescence detector which concerns on Embodiment 2 simply. It is a top view which shows the light receiving element of the fluorescence detector which concerns on Embodiment 2. FIG.

The fluorescence detector according to one aspect of the present invention is a fluorescence detector that detects fluorescence generated from the sample by irradiating the sample with excitation light, and is a first light emitting element that emits the first excitation light. A second light emitting element that emits a second excitation light having a center wavelength different from that of the first excitation light, and when the first light emitting element emits light, the first light emitting element from the first light emitting element. When the excitation light of 1 is reflected toward the sample and the first fluorescence from the sample is transmitted and the second light emitting element emits light, the second light emitting element from the second light emitting element emits light. When the dichroic mirror that reflects the excitation light of 2 toward the sample and transmits the second fluorescence from the sample having a center wavelength different from that of the first fluorescence and the first light emitting element emit light. , said dichroic receives dichroic mirror and the first transmission fluorescence, wherein when the second light emitting element emits light, a light receiving element for receiving the dichroic said second fluorescence transmitted through the dichroic mirror, the The first emission light is emitted so that the first excitation light and the second excitation light are sequentially emitted from the first light emitting element and the second light emitting element at the first rated output and the second rated output, respectively. The control unit includes an element and a control unit that controls the second light emitting element, and when the control unit emits the first excitation light at the first rated output from the first light emitting element. The second light emitting element emits the second excitation light at an output lower than the second rated output, and the second light emitting element emits the second excitation light at the second rated output. At that time, the first light emitting element and the second light emitting element are controlled so that the first light emitting element emits the first excitation light at an output lower than the first rated output. To do .

According to this aspect, since a so-called multi-band pass type dichroic mirror is used, for example, even when the first fluorescent molecule and the second fluorescent molecule having different center wavelengths are labeled on the sample, one is used. The dichroic mirror can reflect the first excitation light and the second excitation light for exciting the first fluorescent molecule and the second fluorescent molecule, respectively, and can reflect the first fluorescent molecule and the second fluorescent molecule, respectively. It is capable of transmitting the first fluorescence and the second fluorescence generated from the fluorescent molecules, respectively. As a result, the turret described in the background technology column, the motor for rotating the turret, and the like can be omitted, and the fluorescence detector can be miniaturized. Further, for example, when the first light emitting element emits the first excitation light at the first rated output, the second light emitting element is excited at a power lower than the second rated output. It emits light. As a result, when the second light emitting element emits the second excitation light at the second rated output in the next sequence, the light emission of the second light emitting element is completely turned off in the immediately preceding sequence. In comparison, the output of the second light emitting element can be stabilized in a short time, and stable second fluorescence can be generated from the sample. As a result, the light emission of the first light emitting element and the light emission of the second light emitting element can be switched at high speed.

For example, each of the first light emitting element and the second light emitting element may be configured to be a laser diode.

According to this aspect, since each of the first light emitting element and the second light emitting element is a laser diode, the wavelength band (spectral width) of each of the first excitation light and the second excitation light is suppressed to be small. Can be done. As a result, it is possible to suppress the mixing of the first excitation light with the first fluorescence and the mixing of the second excitation light with the second fluorescence.

For example, the fluorescence detector further includes an excitation filter arranged between the first light emitting element and the second light emitting element and the dichroic mirror, and the excitation filter is the first light emitting element. When the light is emitted, the wavelength band of the first excitation light from the first light emitting element is limited, and when the second light emitting element emits light, the said from the second light emitting element. It may be configured to limit the wavelength band of the second excitation light.

According to this aspect, since the excitation filter is arranged between the first light emitting element and the second light emitting element and the dichroic mirror, the wavelength bands of the first excitation light and the second excitation light are set. It can be kept small. As a result, it is possible to suppress the mixing of the first excitation light with the first fluorescence and the mixing of the second excitation light with the second fluorescence.

For example, the fluorescence detector further includes an absorption filter arranged between the dichroic mirror and the light receiving element, and the absorption filter comprises the dichroic mirror when the first light emitting element emits light. The wavelength band of the first fluorescence transmitted through the dichroic mirror is limited, and when the second light emitting element emits light, the wavelength band of the second fluorescence transmitted through the dichroic mirror is limited. May be good.

According to this aspect, since the absorption filter is arranged between the dichroic mirror and the light receiving element, the wavelength bands of the first fluorescence and the second fluorescence received by the light receiving element can be suppressed to be small. .. As a result, the excitation of other fluorescent molecules due to the fluorescence generated from the specific fluorescent molecule can be suppressed.

For example, the light receiving element has a light receiving surface on which pixels are arranged, and a filter array having a first filter and a second filter arranged corresponding to the pixels, and the first filter is When the first light emitting element emits light, the wavelength band of the first fluorescence transmitted through the dichroic mirror is limited, and the second filter emits light when the second light emitting element emits light. May be configured to limit the wavelength band of the second fluorescence transmitted through the dichroic mirror.

According to this aspect, since the first filter and the second filter are arranged corresponding to one pixel, each wavelength band of the first fluorescence and the second fluorescence received by one pixel can be set. It can be kept small. Therefore, it is possible to suppress the excitation of other fluorescent molecules by the fluorescence generated from a specific fluorescent molecule.

For example, the first light emitting element, the second light emitting element, the dichroic mirror, and the light receiving element include the first excitation light and the second excitation light reflected by the dichroic mirror, and the dichroic mirror. The transmitted first fluorescence and the second fluorescence may be arranged so as to be located on the same plane.

According to this aspect, the fluorescence detector can be made thinner.

It should be noted that these comprehensive or specific embodiments may be realized in a recording medium such as a system, method, integrated circuit, computer program or computer-readable CD-ROM, and the system, method, integrated circuit, computer program. Alternatively, it may be realized by any combination of recording media.

Hereinafter, embodiments will be specifically described with reference to the drawings.

It should be noted that all of the embodiments described below show comprehensive or specific examples. The numerical values, shapes, materials, components, arrangement positions and connection forms of the components, steps, the order of steps, and the like shown in the following embodiments are examples, and are not intended to limit the present invention. Further, among the components in the following embodiments, the components not described in the independent claims indicating the highest level concept are described as arbitrary components.

(Embodiment 1)
[1-1. Fluorescence detector configuration]
First, the configuration of the fluorescence detector 2 according to the first embodiment will be described with reference to FIGS. 1 and 2. FIG. 1 is a diagram showing the appearance of the fluorescence detector 2 according to the first embodiment. FIG. 1 (a) is a view showing the front side of the fluorescence detector 2, and FIG. 1 (b) is a sectional view taken along line II of FIG. 1 (a). FIG. 2 is a diagram simply showing the configuration of the fluorescence detector 2 according to the first embodiment.

As shown in FIG. 1, the fluorescence detector 2 is a so-called multi-color type fluorescence detector that detects the fluorescence generated from the sample 4 by irradiating the sample 4 with excitation light to excite the sample 4. The fluorescence detector 2 includes a housing 6 and an optical system 8 arranged inside the housing 6.

The sample 4 is a sample to be detected by the fluorescence detector 2, and is, for example, a micro TAS (Total Analysis Systems) chip. Sample 4 contains a plurality of types of biomolecules labeled with a plurality of types of fluorescent molecules having different center wavelengths. Specifically, the sample 4 is a) a first fluorescent molecule (for example, a center wavelength of 455 nm) that is excited by being irradiated with a first excitation light (described later) to generate a first fluorescence (for example, a center wavelength of 455 nm). A biomolecule labeled with Pacific Blue) and a second fluorescent molecule (for example, a center wavelength of 512 nm) that is excited by irradiation with a second excitation light (described later) to generate a second fluorescence (for example, a center wavelength of 512 nm). For example, a biomolecule labeled with BODIPY-FL) and c) a third fluorescence that is excited by irradiation with a third excitation light (described later) to generate a third fluorescence (for example, a center wavelength of 580 nm). A biomolecule labeled with a molecule (eg, TAMRA) and d) a fourth fluorescence that is excited by irradiation with a fourth excitation light (described later) to generate a fourth fluorescence (eg, center wavelength 680 nm). Biomolecules labeled with a molecule (eg ATTO665) are included.

As shown in FIG. 2, the optical system 8 of the fluorescence detector 2 includes a first light emitting element 10, a second light emitting element 12, a third light emitting element 14, a fourth light emitting element 16, and a second light emitting element. 1 lens 18, 2nd lens 20, 3rd lens 22, 4th lens 24, 1st dichroic mirror 26, 2nd dichroic mirror 28, 3rd dichroic mirror 30 A fifth lens 32, an excitation filter 34, a dichroic mirror 36, a sixth lens 38, an absorption filter 40, a seventh lens 42, and a light receiving element 44 are provided.

The first light emitting element 10, the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 are laser diodes that emit monochromatic laser light having different center wavelengths as excitation light. Specifically, the first light emitting element 10 emits a purple laser light having a first central wavelength λ1 (for example, 405 nm) as the first excitation light. The second light emitting element 12 emits a blue laser light having a second central wavelength λ2 (for example, 470 nm) as the second excitation light. The third light emitting element 14 emits a green laser light having a third central wavelength of λ3 (for example, 528 nm) as a third excitation light. The fourth light emitting element 16 emits a red laser light having a fourth central wavelength λ4 (for example, 632 nm) as the fourth excitation light.

The first lens 18, the second lens 20, the third lens 22, and the fourth lens 24 are the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, and the fourth, respectively. This is a collimating lens that converts the laser light from the light emitting element 16 from divergent light to parallel light.

Each of the first dichroic mirror 26, the second dichroic mirror 28, and the third dichroic mirror 30 is an optical element that reflects only light having a specific central wavelength and transmits light having other central wavelengths. Specifically, the first dichroic mirror 26 reflects only the green laser light and transmits the laser light of a color other than green. The second dichroic mirror 28 reflects only the blue laser light and transmits the laser light of a color other than blue. The third dichroic mirror 30 reflects only the purple laser light and transmits the laser light of a color other than purple.

The red laser light (fourth excitation light) from the fourth light emitting element 16 passes through each of the first dichroic mirror 26, the second dichroic mirror 28, and the third dichroic mirror 30. The green laser light (third excitation light) from the third light emitting element 14 is reflected by the first dichroic mirror 26 and then passes through the second dichroic mirror 28 and the third dichroic mirror 30. The blue laser light (second excitation light) from the second light emitting element 12 is reflected by the second dichroic mirror 28 and then passes through the third dichroic mirror 30. The purple laser light (first excitation light) from the first light emitting element 10 is reflected by the third dichroic mirror 30.

The fifth lens 32 is a condensing lens for condensing the laser light emitted from the third dichroic mirror 30.

The excitation filter 34 is a so-called multi-bandpass type filter that transmits only light in a plurality of wavelength bands. The excitation filter 34 is arranged between the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, the fourth light emitting element 16, and the dichroic mirror 36. FIG. 3 is a diagram showing the transmission characteristics of the excitation filter 34 according to the first embodiment. FIG. 3A is a graph showing the transmission characteristics of the excitation filter 34, and FIG. 3B is a first excitation light, a second excitation light, and a third excitation light limited by the excitation filter 34. It is a graph which shows each wavelength band of the excitation light and the 4th excitation light of FIG. 3, and FIG. 3C is a graph which shows the absorption / fluorescence intensity in sample 4.

In the example shown in FIG. 3, the excitation filter 34 is a laser having a first wavelength band B1, a second wavelength band B2, a third wavelength band B3, and a fourth wavelength band B4 (B1 <B2 <B3 <B4). Only light is transmitted. As a result, the excitation filter 34 limits the wavelength band of the first excitation light from the first light emitting element 10 to the first wavelength band B1. Further, the excitation filter 34 limits the wavelength band of the second excitation light from the second light emitting element 12 to the second wavelength band B2. Further, the excitation filter 34 limits the wavelength band of the third excitation light from the third light emitting element 14 to the third wavelength band B3. Further, the excitation filter 34 limits the wavelength band of the fourth excitation light from the fourth light emitting element 16 to the fourth wavelength band B4.

The dichroic mirror 36 is a so-called multi-bandpass type dichroic mirror that reflects only light having a plurality of central wavelengths and transmits light having other central wavelengths. FIG. 4 is a diagram showing the transmission characteristics of the dichroic mirror 36 according to the first embodiment. FIG. 4A is a graph showing the transmission characteristics of the dichroic mirror 36, and FIG. 4B is a graph showing the absorption / fluorescence intensity in the sample 4.

In the example shown in FIG. 4, the dichroic mirror 36 has a first center wavelength λ1 laser light (first excitation light), a second center wavelength λ2 laser light (second excitation light), and a third center. Reflects only laser light of wavelength λ3 (third excitation light) and laser light of fourth center wavelength λ4 (fourth excitation light), and laser light and fluorescence of other center wavelengths (eg, first fluorescence). , 2nd fluorescence, 3rd fluorescence and 4th fluorescence).

The sixth lens 38 is a condensing lens for condensing the laser light reflected by the dichroic mirror 36 and the fluorescence generated from the sample 4.

The absorption filter 40 is a so-called multi-bandpass type filter that transmits only light in a plurality of wavelength bands. The absorption filter 40 is arranged between the dichroic mirror 36 and the light receiving element 44. FIG. 5 is a diagram showing the transmission characteristics of the absorption filter 40 according to the first embodiment. FIG. 5A is a graph showing the transmission characteristics of the absorption filter 40, and FIG. 5B is a first fluorescence, a second fluorescence, and a third fluorescence limited by the absorption filter 40. It is a graph which shows each wavelength band of the 4th fluorescence, and (c) of FIG. 5 is a graph which shows the absorption / fluorescence intensity in the sample 4.

In the example shown in FIG. 5, the absorption filter 40 has fluorescence in the fifth wavelength band B5, the sixth wavelength band B6, the seventh wavelength band B7, and the eighth wavelength band B8 (B5 <B6 <B7 <B8). Only transparent. As a result, the absorption filter 40 limits the wavelength band of the first fluorescence generated from the sample 4 to the fifth wavelength band B5. Further, the absorption filter 40 limits the wavelength band of the second fluorescence generated from the sample 4 to the sixth wavelength band B6. Further, the absorption filter 40 limits the wavelength band of the third fluorescence generated from the sample 4 to the seventh wavelength band B7. Further, the absorption filter 40 limits the wavelength band of the fourth fluorescence generated from the sample 4 to the eighth wavelength band B8.

The seventh lens 42 is a condensing lens for condensing the fluorescence transmitted through the absorption filter 40.

The light receiving element 44 receives fluorescence from the seventh lens 42 (first fluorescence, second fluorescence, third fluorescence, and fourth fluorescence) to indicate a fluorescence intensity (fluorescence signal). To generate.

As shown in FIG. 1, the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, the fourth light emitting element 16, the dichroic mirror 36, and the light receiving element 44 are formed by the dichroic mirror 36. The reflected excitation light (first excitation light, second excitation light, third excitation light and fourth excitation light) and fluorescence transmitted through the dichroic mirror 36 (first fluorescence, second fluorescence, second fluorescence). The fluorescence of No. 3 and the fluorescence of No. 4) are arranged so as to be located on the same plane (on the YZ plane).

As shown in FIG. 2, the fluorescence detector 2 further includes a control unit 46. The control unit 46 outputs the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 to a predetermined rated output (for example, 100% light intensity) (first rated output). And an example of the second rated output), and the light is repeatedly emitted in this order sequence.

At this time, when the control unit 46 causes the first light emitting element 10 to emit light at a predetermined rated output, the second light emitting element 12, the third light emitting element 14, and the third light emitting element 14 other than the first light emitting element 10 are emitted. The fourth light emitting element 16 is made to emit light at an output lower than a predetermined rated output, specifically, an output equal to or higher than a light emission threshold (for example, less than 1% of light) and 28% or less of a predetermined rated output (for example, 28% of light). .. Further, when the control unit 46 emits light from the second light emitting element 12 at a predetermined rated output, the control unit 46 emits light from the first light emitting element 10, the third light emitting element 14, and the third light emitting element 14 other than the second light emitting element 12. The light emitting element 16 of No. 4 is made to emit light at an output equal to or higher than the light emission threshold and 28% or less of the predetermined rated output.

Further, when the control unit 46 emits light from the third light emitting element 14 at a predetermined rated output, the control unit 46 emits light from the first light emitting element 10, the second light emitting element 12, and the second light emitting element 12 other than the third light emitting element 14. The light emitting element 16 of No. 4 is made to emit light at an output equal to or higher than the light emission threshold and 28% or less of the predetermined rated output. Further, when the control unit 46 causes the fourth light emitting element 16 to emit light at a predetermined rated output, the control unit 46 emits light from the first light emitting element 10, the second light emitting element 12, and the second light emitting element 12 other than the fourth light emitting element 16. The light emitting element 14 of 3 is made to emit light at an output equal to or more than the light emission threshold value and 28% or less of the predetermined rated output.

Further, the control unit 46 receives the light receiving signal generated by the light receiving element 44. At this time, when the first light emitting element 10 is made to emit light at a predetermined rated output, the control unit 46 detects the signal component corresponding to the first fluorescence of the received light signal, and the light receiving signal Of these, the signal components corresponding to the second fluorescence, the third fluorescence, and the fourth fluorescence are removed by calculation. Further, when the second light emitting element 12 is made to emit light at a predetermined rated output, the control unit 46 detects a signal component corresponding to the second fluorescence in the received light signal and among the received light signals. The signal components corresponding to the first fluorescence, the third fluorescence, and the fourth fluorescence are removed by calculation.

Further, when the third light emitting element 14 is made to emit light at a predetermined rated output, the control unit 46 detects a signal component corresponding to the third fluorescence in the received light signal and among the received light signals. The signal components corresponding to the first fluorescence, the second fluorescence, and the fourth fluorescence are removed by calculation. Further, when the fourth light emitting element 16 is made to emit light at a predetermined rated output, the control unit 46 detects a signal component corresponding to the fourth fluorescence in the received light signal and among the received light signals. The signal components corresponding to the first fluorescence, the second fluorescence, and the third fluorescence are removed by calculation.

[1-2. Fluorescence detector operation]
Next, the operation of the fluorescence detector 2 (control method of the fluorescence detector 2) will be described with reference to FIGS. 2 and 6. FIG. 6 is a flowchart showing the operation of the fluorescence detector 2 according to the first embodiment.

As shown in FIG. 6, first, the first sequence (S1 to S3) is executed. The control unit 46 causes the first light emitting element 10 to emit light at a predetermined rated output (S1). At this time, the control unit 46 sets the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 other than the first light emitting element 10 to the light emitting threshold value or more and 28% or less of the predetermined rated output. Make the output emit light.

As shown in FIG. 2, the first excitation light from the first light emitting element 10 is reflected by the third dichroic mirror 30, then passes through the fifth lens 32 and is incident on the excitation filter 34. At this time, the excitation filter 34 limits the wavelength band of the first excitation light to the first wavelength band B1 (see FIG. 3). The first excitation light transmitted through the excitation filter 34 is reflected by the dichroic mirror 36, and then passes through the sixth lens 38 to irradiate the sample 4.

When the first fluorescent molecule contained in the sample 4 is excited by the first excitation light, the first fluorescence is generated from the sample 4. The first fluorescence generated from the sample 4 passes through the sixth lens 38 and the dichroic mirror 36, and then enters the absorption filter 40. At this time, the absorption filter 40 limits the wavelength band of the first fluorescence to the fifth wavelength band B5 (see FIG. 5). The first fluorescence transmitted through the absorption filter 40 is transmitted through the seventh lens 42 and received by the light receiving element 44 (S2). As a result, the light receiving element 44 generates a light receiving signal based on the first fluorescence received.

As described above, the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 other than the first light emitting element 10 emit light at a relatively low amount of light. In addition to the signal components corresponding to the first fluorescence, a small amount of signal components corresponding to the second fluorescence, the third fluorescence, and the fourth fluorescence are included. Therefore, the control unit 46 calculates and removes the signal components corresponding to the second fluorescence, the third fluorescence, and the fourth fluorescence of the received light signal, so that the signal corresponding to the first fluorescence of the received light signal is obtained. Only components can be detected.

After that, the control unit 46 lowers the output of the first light emitting element 10 from the predetermined rated output (S3), and emits light at an output equal to or higher than the light emission threshold value and 28% or less of the predetermined rated output.

Next, the second sequence (S4 to S6) is executed. The control unit 46 causes the second light emitting element 12 to emit light at a predetermined rated output (S4). At this time, the control unit 46 sets the first light emitting element 10, the third light emitting element 14, and the fourth light emitting element 16 other than the second light emitting element 12 to the light emitting threshold value or more and 28% or less of the predetermined rated output. Make the output emit light.

As shown in FIG. 2, the second excitation light from the second light emitting element 12 is reflected by the second dichroic mirror 28 and then transmitted through the third dichroic mirror 30 and the fifth lens 32 to be excited. It is incident on the filter 34. At this time, the excitation filter 34 limits the wavelength band of the second excitation light to the second wavelength band B2 (see FIG. 3). The second excitation light that has passed through the excitation filter 34 is reflected by the dichroic mirror 36, and then passes through the sixth lens 38 to irradiate the sample 4.

When the second fluorescent molecule contained in the sample 4 is excited by the second excitation light, the second fluorescence is generated from the sample 4. The second fluorescence generated from the sample 4 passes through the sixth lens 38 and the dichroic mirror 36, and then enters the absorption filter 40. At this time, the absorption filter 40 limits the wavelength band of the second fluorescence to the sixth wavelength band B6 (see FIG. 5). The second fluorescence transmitted through the absorption filter 40 is transmitted through the seventh lens 42 and received by the light receiving element 44 (S5). As a result, the light receiving element 44 generates a light receiving signal based on the second fluorescence received.

As described above, the first light emitting element 10, the third light emitting element 14, and the fourth light emitting element 16 other than the second light emitting element 12 emit light at a relatively low amount of light. In addition to the signal components corresponding to the second fluorescence, a small amount of signal components corresponding to the first fluorescence, the third fluorescence, and the fourth fluorescence are included. Therefore, the control unit 46 calculates and removes the signal components corresponding to the first fluorescence, the third fluorescence, and the fourth fluorescence of the received light signal, so that the signal corresponding to the second fluorescence of the received light signal is obtained. Only components can be detected.

After that, the control unit 46 lowers the output of the second light emitting element 12 from the predetermined rated output (S6), and emits light at an output equal to or higher than the light emission threshold value and 28% or less of the predetermined rated output.

Next, the third sequence (S7 to S9) is executed. The control unit 46 causes the third light emitting element 14 to emit light at a predetermined rated output (S7). At this time, the control unit 46 sets the first light emitting element 10, the second light emitting element 12, and the fourth light emitting element 16 other than the third light emitting element 14 to the light emitting threshold value or more and 28% or less of the predetermined rated output. Make the output emit light.

As shown in FIG. 2, the third excitation light from the third light emitting element 14 is reflected by the first dichroic mirror 26, and then the second dichroic mirror 28, the third dichroic mirror 30, and the fifth dichroic mirror 30. It passes through the lens 32 and enters the excitation filter 34. At this time, the excitation filter 34 limits the wavelength band of the third excitation light to the third wavelength band B3 (see FIG. 3). The third excitation light that has passed through the excitation filter 34 is reflected by the dichroic mirror 36, and then passes through the sixth lens 38 to irradiate the sample 4.

When the third fluorescent molecule contained in the sample 4 is excited by the third excitation light, the third fluorescence is generated from the sample 4. The third fluorescence generated from the sample 4 passes through the sixth lens 38 and the dichroic mirror 36, and then enters the absorption filter 40. At this time, the absorption filter 40 limits the wavelength band of the third fluorescence to the seventh wavelength band B7 (see FIG. 5). The third fluorescence transmitted through the absorption filter 40 is transmitted through the seventh lens 42 and received by the light receiving element 44 (S8). As a result, the light receiving element 44 generates a light receiving signal based on the received third fluorescence.

As described above, the first light emitting element 10, the second light emitting element 12, and the fourth light emitting element 16 other than the third light emitting element 14 emit light with a relatively low amount of light. In addition to the signal components corresponding to the third fluorescence, a small amount of signal components corresponding to the first fluorescence, the second fluorescence, and the fourth fluorescence are included. Therefore, the control unit 46 calculates and removes the signal components corresponding to the first fluorescence, the second fluorescence, and the fourth fluorescence of the received light signal, so that the signal corresponding to the third fluorescence of the received light signal is obtained. Only components can be detected.

After that, the control unit 46 lowers the output of the third light emitting element 14 from the predetermined rated output (S9), and emits light at an output equal to or higher than the light emission threshold value and 28% or less of the predetermined rated output.

Next, the fourth sequence (S10 to S12) is executed. The control unit 46 causes the fourth light emitting element 16 to emit light at a predetermined rated output (S10). At this time, the control unit 46 sets the first light emitting element 10, the second light emitting element 12, and the third light emitting element 14 other than the fourth light emitting element 16 to the light emitting threshold value or more and 28% or less of the predetermined rated output. Make the output emit light.

As shown in FIG. 2, the fourth excitation light from the fourth light emitting element 16 passes through the first dichroic mirror 26, the second dichroic mirror 28, the third dichroic mirror 30, and the fifth lens 32. After that, it enters the excitation filter 34. At this time, the excitation filter 34 limits the wavelength band of the fourth excitation light to the fourth wavelength band B4 (see FIG. 3). The fourth excitation light that has passed through the excitation filter 34 is reflected by the dichroic mirror 36, and then passes through the sixth lens 38 to irradiate the sample 4.

When the fourth fluorescent molecule contained in the sample 4 is excited by the fourth excitation light, the fourth fluorescence is generated from the sample 4. The fourth fluorescence generated from the sample 4 passes through the sixth lens 38 and the dichroic mirror 36, and then enters the absorption filter 40. At this time, the absorption filter 40 limits the wavelength band of the fourth fluorescence to the eighth wavelength band B8 (see FIG. 5). The fourth fluorescence transmitted through the absorption filter 40 is transmitted through the seventh lens 42 and received by the light receiving element 44 (S11). As a result, the light receiving element 44 generates a light receiving signal based on the received fourth fluorescence.

As described above, the first light emitting element 10, the second light emitting element 12, and the third light emitting element 14 other than the fourth light emitting element 16 emit light at a relatively low amount of light. In addition to the signal component corresponding to the fourth fluorescence, a small amount of the signal component corresponding to the first fluorescence, the second fluorescence, and the third fluorescence is included. Therefore, the control unit 46 calculates and removes the signal components corresponding to the first fluorescence, the second fluorescence, and the third fluorescence of the received light signal, so that the signal corresponding to the fourth fluorescence of the received light signal is obtained. Only components can be detected.

After that, the control unit 46 lowers the output of the fourth light emitting element 16 from the predetermined rated output (S12), and causes the control unit 46 to emit light at an output equal to or higher than the light emission threshold value and 28% or less of the predetermined rated output.

If the detection of the first to fourth fluorescence is not completed (NO in S13), the above-mentioned steps S1 to S12 are executed again. When the detection of the first to fourth fluorescence is completed (YES in S13), the operation of the fluorescence detector 2 is completed.

[1-3. effect]
As described above, in the fluorescence detector 2 according to the first embodiment, a so-called multi-bandpass type dichroic mirror 36 is used. As a result, even when a plurality of types of fluorescent molecules are labeled on the sample 4, one dichroic mirror 36 can reflect a plurality of types of excitation light for exciting the plurality of types of fluorescent molecules. It is possible to transmit a plurality of types of fluorescence generated from each of a plurality of types of fluorescent molecules. As a result, the turret described in the background technology column, the motor for rotating the turret, and the like can be omitted, and the fluorescence detector 2 can be miniaturized.

As an example of the size of the housing 6, as shown in FIG. 1, the width W of the housing 6 can be about 17 mm, the depth D can be about 74 mm, and the height H can be about 39 mm.

Further, as described above, in the fluorescence detector 2 according to the first embodiment, for example, when the first light emitting element 10 emits light at a predetermined rated output, the first light emitting element other than the first light emitting element 10 is emitted. The second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 are made to emit light at an output lower than a predetermined rated output (with a light amount larger than 0%). As a result, when the second light emitting element 12 was made to emit light at a predetermined rated output in the next sequence, the light emission of the second light emitting element 12 was completely turned off (light intensity 0%) in the immediately preceding sequence. Compared with the case, the output of the second light emitting element 12 can be stabilized in a short time, and stable second fluorescence can be generated from the sample 4. As a result, the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 to be light-emitting at a predetermined rated output can be switched at high speed.

For example, when the second light emitting element 12 emits light at a predetermined rated output, the first light emitting element 10, the third light emitting element 14, and the fourth light emitting element other than the second light emitting element 12 are emitted. It is preferable to make 16 emit light at an output of 28% or less of a predetermined rated output. At this time, when the first light emitting element 10, the third light emitting element 14, and the fourth light emitting element 16 other than the second light emitting element 12 are made to emit light at an output exceeding 28% of the predetermined rated output, the light is emitted. For example, the third fluorescent molecule (TAMRA) contained in the sample 4 is excited by the third excitation light (green laser light) from the third light emitting element 14, so that it is generated from the third fluorescent molecule. The crosstalk ratio of the fluorescence signal of the third fluorescence becomes large.

[1-4. Examples and Comparative Examples]
Hereinafter, examples and comparative examples of the fluorescence detector will be described with reference to FIGS. 7 to 13.

FIG. 7 is a diagram showing light emission sequences of the light emitting elements 10, 12, 14 and 16 of the fluorescence detector 2 according to the embodiment. FIG. 8 is a diagram showing light emitting conditions of the light emitting elements 10, 12, 14 and 16 of the fluorescence detector 2 according to the embodiment. FIG. 9 is a diagram showing the transmission characteristics of the dichroic mirror 36 of the fluorescence detector 2 according to the embodiment. FIG. 10 is a diagram showing light emission sequences of the light emitting elements 10, 12, 14 and 16 of the fluorescence detector 2'according to the comparative example.

FIG. 11 is a diagram showing the detection result of the fluorescence signal by the fluorescence detector 2 according to the embodiment. FIG. 12 is a diagram showing the crosstalk ratio of the fluorescent signal of FIG. FIG. 13 is a diagram showing the results of detection of fluorescence intensity by the fluorescence detectors 2 and 2'according to Examples and Comparative Examples. FIG. 13A is a diagram showing the detection result of the fluorescence intensity by the fluorescence detector 2 according to the example, and FIG. 13B is a diagram showing the detection result of the fluorescence intensity by the fluorescence detector 2'according to the comparative example. FIG. 3C is a diagram showing the volatility of fluorescence intensity by the fluorescence detectors 2 and 2'according to Examples and Comparative Examples.

[1-4-1. Example]
As the fluorescence detector according to the example, the fluorescence detector 2 shown in FIG. 1 was used. As the housing 6, a housing 6 formed by carving out aluminum was used.

As the first light emitting element 10, an LED (Light Emitting Diode) (LZ1-00UB00, manufactured by LED ENGIN) that emits purple light having a center wavelength of 405 nm was used. As the second light emitting element 12, an LED (LBW5SN-GYHZ-25 manufactured by OSRAM) that emits blue light having a center wavelength of 470 nm was used. As the third light emitting element 14, an LED (LTW5SN-KYLY-25 manufactured by OSRAM) that emits green light having a center wavelength of 528 nm was used. As the fourth light emitting element 16, an LED (LRW5SN-JYKY-1 manufactured by OSRAM) that emits red light having a center wavelength of 632 nm was used.

Further, the fourth light emitting element 16 (red: 632 nm), the third light emitting element 14 (green: 528 nm), the second light emitting element 12 (blue: 475 nm), and the first light emitting element 10 (purple: 405 nm) are used. , The sequence “1”, “2”, “3”, “4”, ... As shown in FIG. 7 was repeatedly emitted at a light intensity of 100% (rated output). The light emitting period of each of the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 was 500 msec. In FIG. 7, “High” means that the output of the light emitting element has a light intensity of 100%, and “Low” means that the output of the light emitting element has a light intensity of 28% or less. For example, in the sequence "1" shown in FIG. 7, the fourth light emitting element 16 is made to emit light at a light amount of 100% (High), and the first light emitting element 10 and the second light emitting element 12 other than the fourth light emitting element 16 are emitted. And the third light emitting element 14 was made to emit light with a light amount of 28% or less (Low).

Further, the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 were made to emit light by the LED dimming circuit under the light emitting conditions shown in FIG. The LED dimming circuit was a circuit that controls the amount of current flowing through the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 by D / A converter control. .. For example, when the fourth light emitting element 16 was made to emit light at High, the input current was 700 mA, the voltage was 2.48 V, the power consumption was 1.74 W, and the light amount was 100%. Further, when the fourth light emitting element 16 was made to emit light at Low, the input current was 100 mA, the voltage was 1.9 V, the power consumption was 0.19 W, and the light amount was 15%.

Further, as the dichroic mirror 36, a multi-bandpass type dichroic mirror (manufactured by Semirock, FF409 / 493/573/652-Di01-25x36) having the transmission characteristics shown in FIG. 9 was used. The dichroic mirror 36 used was cut to a size of 12 × 18 mm.

As the excitation filter 34, a multi-bandpass type filter (manufactured by Semirock, FF01-392 / 474/554/635-25) was used. As the absorption filter 40, a multi-bandpass type filter (manufactured by Semirock, FF01-432 / 515/595 / 730-25) was used. As the excitation filter 34 and the absorption filter 40, those cut to a diameter of 12 mm were used.

A plano-convex lens (manufactured by Edmund, # 45-228) was used as the first lens 18, the second lens 20, the third lens 22, and the fourth lens 24. An achromat lens (manufactured by Edmund, # 65-549) was used as the fifth lens 32, the sixth lens 38, and the seventh lens 42.

As the light receiving element 44, a photodiode (manufactured by Hamamatsu, S2386-44K) was used. The fluorescence signal from the light receiving element 44 is amplified by a photosensor amplifier (manufactured by Hamamatsu, C9329) at a magnification of 1E + 7V / A and then A / D converted, and a voltage value indicating the magnitude of the fluorescence signal is taken into a personal computer. It is.

As Sample 4, a Q-Probe color compensation kit (manufactured by Nippon Steel & Sumitomo Metal Industries, Ltd.) was used. 200 μL of the fluorescent reagent was injected into the microtube, and the fluorescent reagent was irradiated with excitation light to detect fluorescence. Pacific Blue (hereinafter, also referred to as “dye B”) excited by purple light is used as the first fluorescent molecule constituting Q-Probe, and BODIPY-FL (hereinafter, also referred to as “dye B”) excited by blue light is used as the second fluorescent molecule. , "Dye G") and TAMRA (hereinafter, also referred to as "Dye R") excited by green light as the third fluorescent molecule, ATTO665 excited by red light as the fourth fluorescent molecule. (Hereinafter, also referred to as "dye S") was used.

[1-4-2. Comparative example]
As the fluorescence detector according to the comparative example, the fluorescence detector 2'shown in FIG. 1 was used.

The fourth light emitting element 16 (red: 632 nm), the third light emitting element 14 (green: 528 nm), the second light emitting element 12 (blue: 475 nm), and the first light emitting element 10 (purple: 405 nm) are shown in the figure. The sequence "1", "2", "3", "4", ... Shown in No. 10 was repeatedly emitted with a light intensity of 100% (rated output). The light emitting period of each of the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 was 500 msec. In FIG. 10, “High” means that the output of the light emitting element has a light intensity of 100% (rated output), and “Off” means that the output of the light emitting element has a light intensity of 0% (does not emit light). To do. For example, in the sequence "1" shown in FIG. 10, the fourth light emitting element 16 is made to emit light at a light amount of 100% (High), and the first light emitting element 10 and the second light emitting element 12 other than the fourth light emitting element 16 are emitted. And the light emission of the third light emitting element 14 was completely turned off.

Since the conditions other than the above in the comparative example are the same as those in the embodiment, the description thereof will be omitted.

[1-4-3. result]
The detection results of the fluorescence signal by the fluorescence detector 2 according to the embodiment are as shown in FIGS. 11 and 12.

As is clear from FIGS. 11 and 12, for example, when the fourth light emitting element 16 emits light with a light amount of 100%, the fourth fluorescent molecule (dye S) contained in the sample 4 emits the fourth light. By being excited by the fourth excitation light (red light) from the element 16, the fluorescence signal (voltage value) of the fourth fluorescence generated from the fourth fluorescent molecule became large. At this time, when the crosstalk ratio of the fluorescent signal of the fourth fluorescence is 100.0%, the first fluorescent molecule (dye B), the second fluorescent molecule (dye G) and the third fluorescent molecule (dye B) contained in the sample 4 are contained. The crosstalk ratio of the fluorescent signals of the first fluorescence, the second fluorescence, and the third fluorescence generated from the fluorescent molecule (dye R) of the above was 1.4%, 3.4%, and 1.9%, respectively. ..

Further, when the third light emitting element 14 is made to emit light with a light amount of 100%, the third fluorescent molecule (dye R) contained in the sample 4 is the third excitation light (green) from the third light emitting element 14. The fluorescence signal of the third fluorescence generated from the third fluorescent molecule became larger by being excited by the light). At this time, when the crosstalk ratio of the fluorescent signal of the third fluorescence is 100.0%, the first fluorescent molecule (dye B), the second fluorescent molecule (dye G) and the fourth fluorescent molecule (dye B) contained in the sample 4 are contained. The crosstalk ratios of the fluorescent signals of the first fluorescence, the second fluorescence, and the fourth fluorescence generated from the fluorescent molecule (dye S) of the above were 9.7%, 26.4%, and 0.7%, respectively. ..

Further, when the second light emitting element 12 is made to emit light with a light amount of 100%, the second fluorescent molecule (dye G) contained in the sample 4 is the second excitation light (blue) from the second light emitting element 12. The fluorescence signal of the second fluorescence generated from the second fluorescent molecule became larger by being excited by the light). At this time, when the crosstalk ratio of the fluorescent signal of the second fluorescence is 100.0%, the first fluorescent molecule (dye B), the third fluorescent molecule (dye R) and the fourth fluorescent molecule (dye B) contained in the sample 4 are contained. The crosstalk ratios of the fluorescent signals of the first fluorescence, the third fluorescence, and the fourth fluorescence generated from the fluorescent molecule (dye S) of the above were 3.8%, 0.2%, and 0.7%, respectively. ..

Further, when the first light emitting element 10 is made to emit light with a light amount of 100%, the first fluorescent molecule (dye B) contained in the sample 4 is the first excitation light (purple) from the first light emitting element 10. The fluorescence signal of the first fluorescence generated from the first fluorescent molecule was increased by being excited by the light). At this time, when the crosstalk ratio of the fluorescent signal of the first fluorescence is 100.0%, the second fluorescent molecule (dye G), the third fluorescent molecule (dye R) and the fourth fluorescent molecule (dye G) contained in the sample 4 are contained. The crosstalk ratios of the fluorescent signals of the second fluorescence, the third fluorescence, and the fourth fluorescence generated from the fluorescent molecule (dye S) of the above were 0.8%, 0.0%, and 1.5%, respectively. ..

Further, the results of detecting the fluorescence intensity by the fluorescence detectors 2 and 2'in the Examples and Comparative Examples are as shown in FIG.

As is clear from (a) and (c) of FIG. 13, in the fluorescence detector 2 according to the embodiment, the peak value of the detected fluorescence intensity hardly varies, and the fluctuation rate of the fluorescence intensity is 0.2. %Met.

On the other hand, as is clear from (b) and (c) of FIG. 13, in the fluorescence detector 2'according to the comparative example, the peak value of the detected fluorescence intensity varies, and the fluctuation rate of the fluorescence intensity is 4. It was .1%.

From the above results, it was confirmed that the fluorescence detector 2 of the example has an effect of generating stable fluorescence from the sample 4.

(Embodiment 2)
Next, the configuration of the fluorescence detector 2A according to the second embodiment will be described with reference to FIGS. 14 and 15. FIG. 14 is a diagram simply showing the configuration of the fluorescence detector 2A according to the second embodiment. FIG. 15 is a plan view showing a light receiving element 44A of the fluorescence detector 2A according to the second embodiment.

As shown in FIG. 14, in the fluorescence detector 2A according to the second embodiment, the light receiving element 44A has a filter array 48. As shown in FIG. 15, a plurality of pixels 50 are arranged in a matrix on the light receiving surface of the light receiving element 44A. The filter array 48 has a plurality of filter sets 52 arranged corresponding to the plurality of pixels 50, respectively.

Each of the plurality of filter sets 52 has a first filter 52a, a second filter 52b, a third filter 52c, and a fourth filter 52d. The first filter 52a is a so-called bandpass type filter that limits the wavelength band of the first fluorescence transmitted through the dichroic mirror 36 when the first light emitting element 10 emits light at a predetermined rated output. The second filter 52b is a so-called bandpass type filter that limits the wavelength band of the second fluorescence transmitted through the dichroic mirror 36 when the second light emitting element 12 emits light at a predetermined rated output. The third filter 52c is a so-called bandpass type filter that limits the wavelength band of the second fluorescence transmitted through the dichroic mirror 36 when the third light emitting element 14 emits light at a predetermined rated output. The fourth filter 52d is a so-called bandpass type filter that limits the wavelength band of the fourth fluorescence transmitted through the dichroic mirror 36 when the fourth light emitting element 16 emits light at a predetermined rated output.

As a result, the first filter 52a, the second filter 52b, the third filter 52c, and the fourth filter 52d are arranged with respect to one pixel 50, so that the fluorescence received by one pixel 50 can be received. The spectral width can be reduced. Therefore, it is possible to suppress the excitation of other fluorescent molecules by the fluorescence generated from a specific fluorescent molecule.

(Modification example)
Although the fluorescence detector according to one or more embodiments has been described above based on the above embodiment, the present invention is not limited to this embodiment. As long as it does not deviate from the gist of the present invention, there may be one or a plurality of forms in which various modifications that can be conceived by those skilled in the art are applied to the present embodiment, or forms constructed by combining components in different embodiments or modifications. It may be included within the scope of the embodiment.

In each of the above embodiments, the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 are configured by a laser diode, but the present invention is not limited to this, and for example, an LED. Etc. may be configured.

In each of the above embodiments, both the excitation filter 34 and the absorption filter 40 are arranged, but the present invention is not limited to this, and only one of the excitation filter 34 and the absorption filter 40 may be arranged, or the excitation filter may be arranged. Both 34 and the absorption filter 40 may be omitted.

In each of the above embodiments, the rated outputs of the first light emitting element 10, the second light emitting element 12, the third light emitting element 14, and the fourth light emitting element 16 are all the same output (light intensity 100%). However, the output is not limited to this, and the outputs may be different from each other. For example, the first rated output of the first light emitting element 10 and the second rated output of the second light emitting element 12 may be different.

In each of the above embodiments, each component may be configured by dedicated hardware or may be realized by executing a software program suitable for each component. Each component may be realized by a program execution unit such as a CPU or a processor reading and executing a software program recorded on a recording medium such as a hard disk or a semiconductor memory.

The following cases are also included in the present invention.

(1) Each of the above devices can be specifically realized by a computer system including a microprocessor, a ROM, a RAM, a hard disk unit, a display unit, a keyboard, a mouse, and the like. A computer program is stored in the RAM or the hard disk unit. Each device achieves its function by operating the microprocessor according to a computer program. Here, a computer program is configured by combining a plurality of instruction codes indicating commands to a computer in order to achieve a predetermined function.

(2) A part or all of the components constituting each of the above devices may be composed of one system LSI (Large Scale Integration: large-scale integrated circuit). A system LSI is an ultra-multifunctional LSI manufactured by integrating a plurality of components on a single chip, and specifically, is a computer system including a microprocessor, ROM, RAM, and the like. .. A computer program is stored in the ROM. The system LSI achieves its function by loading a computer program from the ROM into the RAM by the microprocessor and performing operations such as calculations according to the loaded computer program.

(3) A part or all of the components constituting each of the above devices may be composed of an IC card or a single module that can be attached to and detached from each device. An IC card or module is a computer system composed of a microprocessor, ROM, RAM, and the like. The IC card or module may include the above-mentioned super multifunctional LSI. When the microprocessor operates according to a computer program, the IC card or module achieves its function. This IC card or this module may have tamper resistance.

(4) The present invention may be realized by the method shown above. Further, these methods may be realized by a computer program realized by a computer, or may be realized by a digital signal composed of a computer program.

The present invention also relates to recording media capable of computer-readable computer programs or digital signals, such as flexible discs, hard disks, CD-ROMs, MOs, DVDs, DVD-ROMs, DVD-RAMs, and BDs (Blu-ray®). ) Disc), may be realized by recording in a semiconductor memory or the like. Further, it may be realized by a digital signal recorded on these recording media.

Further, the present invention may transmit a computer program or a digital signal via a telecommunication line, a wireless or wired communication line, a network typified by the Internet, data broadcasting, or the like.

Further, the present invention is a computer system including a microprocessor and a memory, in which the memory stores a computer program, and the microprocessor may operate according to the computer program.

It may also be carried out by another independent computer system by recording the program or digital signal on a recording medium and transferring it, or by transferring the program or digital signal via a network or the like.

(5) The above-described embodiment and the above-described modification may be combined.

The fluorescence detector of the present invention can be applied to a fluorescence detection method for detecting fluorescence generated from a sample, for example, by irradiating the sample with excitation light.

2,2', 2A Fluorescence detector 4 Sample 6 Housing 8 Optical system 10 First light emitting element 12 Second light emitting element 14 Third light emitting element 16 Fourth light emitting element 18 First lens 20 Second Lens 22 3rd lens 24 4th lens 26 1st dichroic mirror 28 2nd dichroic mirror 30 3rd dichroic mirror 32 5th lens 34 Excitation filter 36 Dicroic mirror 38 6th lens 40 Absorption filter 42 No. Lens 44, 44A Light receiving element 46 Control unit 48 Filter array 50 Pixels 52 Filter set 52a First filter 52b Second filter 52c Third filter 52d Fourth filter

Claims (6)

A fluorescence detector that detects fluorescence generated from a sample by irradiating the sample with excitation light.
A first light emitting element that emits a first excitation light,
A second light emitting element that emits a second excitation light having a center wavelength different from that of the first excitation light.
When the first light emitting element emits light, the first excitation light from the first light emitting element is reflected toward the sample, and the first fluorescence from the sample is transmitted. When the second light emitting element emits light, the sample that reflects the second excitation light from the second light emitting element toward the sample and has a center wavelength different from that of the first fluorescence. A dichroic mirror that transmits the second fluorescence from
When the first light emitting element emits light, the first fluorescence transmitted through the dichroic mirror is received, and when the second light emitting element emits light, the second light transmitted through the dichroic mirror is received. A light receiving element that receives the fluorescence of
The first excitation light and the second excitation light are sequentially emitted from the first light emitting element and the second light emitting element at the first rated output and the second rated output, respectively. A light emitting element and a control unit for controlling the second light emitting element are provided.
When the control unit emits the first excitation light from the first light emitting element at the first rated output, the second light emitting element has an output lower than that of the second rated output. When the second excitation light is emitted from the second light emitting element and the second excitation light is emitted from the second light emitting element at the second rated output, the first light emitting element has the first rating. A fluorescence detector that controls the first light emitting element and the second light emitting element so as to emit the first excitation light at an output lower than the output . The fluorescence detector according to claim 1, wherein each of the first light emitting element and the second light emitting element is a laser diode. The fluorescence detector further includes an excitation filter arranged between the first light emitting element and the second light emitting element and the dichroic mirror.
The excitation filter limits the wavelength band of the first excitation light from the first light emitting element when the first light emitting element emits light, and when the second light emitting element emits light. The fluorescence detector according to claim 1 or 2, which limits the wavelength band of the second excitation light from the second light emitting element. The fluorescence detector further includes an absorption filter arranged between the dichroic mirror and the light receiving element.
The absorption filter limits the wavelength band of the first fluorescence transmitted through the dichroic mirror when the first light emitting element emits light, and when the second light emitting element emits light, the absorption filter limits the wavelength band of the first fluorescence. The fluorescence detector according to any one of claims 1 to 3, which limits the wavelength band of the second fluorescence transmitted through the dichroic mirror. The light receiving element is
The light receiving surface on which the pixels are arranged and
It has a first filter and a filter array having a second filter arranged corresponding to the pixels.
The first filter limits the wavelength band of the first fluorescence transmitted through the dichroic mirror when the first light emitting element emits light.
The second filter according to any one of claims 1 to 4, which limits the wavelength band of the second fluorescence transmitted through the dichroic mirror when the second light emitting element emits light. Fluorescence detector. The first light emitting element, the second light emitting element, the dichroic mirror, and the light receiving element transmitted the first excitation light and the second excitation light reflected by the dichroic mirror and the dichroic mirror. The fluorescence detector according to any one of claims 1 to 5, wherein the first fluorescence and the second fluorescence are arranged so as to be located on the same plane.