CN104764727A - Fluorescence imaging analysis system and fluorescence imaging analysis method thereof - Google Patents
Fluorescence imaging analysis system and fluorescence imaging analysis method thereof Download PDFInfo
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- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
Abstract
The invention discloses a fluorescence imaging analysis system and a fluorescence imaging analysis method. The fluorescence imaging analysis system comprises a shell, and at least one excitation device and an imaging device, which are arranged inside the shell; the excitation device comprises an excitation light source and an excitation filter; light rays emitted by the excitation light source pass through the excitation filter to form monochromatic excitation light and the monochromatic excitation light is irradiated to a sample stage, and a detected substance emits fluorescent light after being excited; and the imaging device is used for capturing fluorescent light passing through a piece of filter glass and analyzing and determining the concentration of the fluorescent substance. Compared with the prior art, the system can acquire the reliable detection result without arranging an expensive optical grating, an interference color filter and a photomultiplier and determining an electronic circuit, the structure is skillful and simple, the preparation cost is low and the system and the method can be widely applied.
Description
Technical Field
The invention belongs to the field of biology, and particularly relates to a fluorescence imaging analysis system and a fluorescence imaging analysis method thereof.
Background
Fluorescent dyes broadly refer to substances that absorb light of one wavelength and emit light of another wavelength that is greater than the light absorbed. Because of high sensitivity and convenient operation, the fluorescent dye gradually replaces radioactive isotopes as scientific research detection markers, and is widely applied to fluorescence immunization, fluorescent probes, cell dyeing and the like. Including specific DNA staining, and is used for chromosome analysis, cell cycle, apoptosis and other related research. In addition, many nucleic acid dyes are very useful counterstains in multicolor staining systems, and can be used as background control to mark cell nucleus so as to make the spatial relationship of the intracellular structures clear. However, most of the existing fluorescent dye detection devices tend to be high in sensitivity, accuracy and automation, so that the detection instrument has a complex structure and high cost, and meanwhile, the instrument has high use and maintenance requirements, and the application of fluorescent dye in detection is limited. However, the results of the application of fluorescent dyes in biological research are being generalized to a wider range of daily applications such as medical examinations, food tests, and the like. Therefore, it is necessary to develop a general-purpose and inexpensive detection device having sufficient detection accuracy and sensitivity.
Disclosure of Invention
The invention provides a fluorescence imaging analysis system, which comprises a shell, and at least one excitation device and an imaging device which are arranged in the shell; wherein,
the shell is a closed hollow shell, a darkroom is formed in the shell, and a sample stage is arranged in the shell and used for placing a detected object carrying a fluorescent substance; the shell is provided with a light hole, the light hole is just corresponding to the position of the sample table, and the sample table can be observed through the light hole;
the excitation device is arranged in the shell and comprises an excitation light source and an excitation filter; light rays emitted by the excitation light source pass through the excitation filter to form monochromatic excitation light to irradiate the sample table; the detected object placed on the sample stage emits fluorescence after being excited;
the imaging device comprises a filter lens and a camera unit; the light filtering lens is arranged right opposite to the light hole and used for filtering stray light penetrating through the light hole except for fluorescence, and the camera shooting unit is arranged right opposite to the light filtering lens and used for capturing the fluorescence penetrating through the light filtering lens. The imaging device can be arranged at the position of a light-transmitting hole on the shell and is right opposite to the sample stage.
In the present invention, the fluorescent substance includes, but is not limited to, a fluorescent dye, a fluorescent protein, or any other substance that fluoresces when excited.
In the fluorescence imaging analysis system of the present invention, the excitation light source, the excitation filter, or the filter lens may be selectively and selectively provided according to the optical properties of a fluorescent substance contained in the object to be detected.
Different excitation light sources are selected according to different excitation requirements, and meanwhile, the cut-off wavelength of the excitation filter is considered. In the fluorescence imaging analysis system of the present invention, the cutoff wavelength of the excitation filter is a wavelength numerical range between the excitation wavelength and the emission wavelength corresponding to the fluorescent substance, thereby ensuring that only the excitation light is allowed to pass through the excitation filter, and the emission light generated by the fluorescent substance cannot pass through the excitation filter. For example, a fluorescent substance containing SYBR Green I + DNA solution was not detected, the excitation wavelength of SYBR Green I was 497nm, and the emission wavelength of the generated excitation light was 520 nm. In the fluorescence imaging analysis system, the excitation light source is a blue LED lamp, and the wavelength of the blue light is 476-495 nm; the excitation filter is ZB1 type, and the light transmission spectrum range of the excitation filter is 300-500 nm. The cut-off wavelength of the excitation filter is between the excitation wavelength and the emission wavelength.
In the fluorescence imaging analysis system of the present invention, the excitation light source includes, but is not limited to, an LED light source, any suitable light source, and the like.
In one specific embodiment, when the fluorescent substance is a green fluorescent substance, the spectral range of the excitation light source is 445-475 nm; the light transmission spectrum range of the excitation filter is 325 nm-500 nm; the light transmission spectrum range of the filter lens is 500 nm-2500 nm; or,
in another embodiment, when the phosphor is a red phosphor, the spectral range of the excitation light source is 585nm ± 29 nm; the light transmission spectrum of the excitation filter is 500 nm-620 nm; the light transmission spectrum range of the filter lens is 620 nm-2500 nm; or,
in another embodiment, when the fluorescent substance is a yellow fluorescent substance, the spectral range of the excitation light source is 531nm ± 40 nm; the light transmission spectrum of the excitation filter is 350 nm-580 nm; the light transmission spectrum range of the filter lens is 580 nm-2500 nm; or,
in another embodiment, when the phosphor is a blue phosphor, the spectral range of the excitation light source is 357nm ± 44 nm; the light transmission spectrum of the excitation filter is 280 nm-410 nm; the light transmission spectrum range of the filter lens is 410 nm-2500 nm; or,
in other embodiments, the excitation light source, excitation filter or filter lens may be selected to have a suitable spectral range according to the optical properties of any of the fluorochromes/fluorescent proteins/or other fluorescent substances.
In one embodiment, an LED lamp with a peak at 453nm is used as the excitation light source.
In the fluorescence imaging analysis system of the present invention, the excitation device is further provided with a scattering member disposed between the excitation light source and the excitation filter. And light rays emitted by the excitation light source form uniform light beams after passing through the scattering component and irradiate the excitation filter.
In the fluorescence imaging analysis system, the scattering component is a plane scattering mirror, and light rays emitted by the excitation light source form uniform light beams through the plane scattering mirror to irradiate the excitation light filter and then irradiate the sample stage.
In the fluorescence imaging analysis system of the invention, the scattering component may further include a plane mirror and an arc mirror, the plane mirror is used for reflecting the light of the excitation light source to the arc mirror, and the arc mirror forms a uniform light beam to be reflected to the excitation filter and then to irradiate on the sample stage.
In the fluorescence imaging analysis system, the bottom of the sample stage is provided with a slide rail for moving the sample stage out of the shell.
In the fluorescence imaging analysis system of the present invention, the "camera unit" refers to a device capable of capturing excitation light of a sample, for example, a digital camera.
In the fluorescence imaging analysis system of the present invention, the imaging device further includes a camera analysis unit connected to the camera unit for measuring and analyzing the concentration of the fluorescent substance. In a specific embodiment, the camera unit and the camera analysis unit may also be integrated for capturing fluorescence and determining and analyzing the concentration of the captured fluorescent substance.
In the fluorescence imaging analysis system of the present invention, the imaging analysis unit includes:
the digital image acquisition unit is used for capturing the fluorescence which penetrates through the filter lens to obtain a digital image;
the image processing unit is connected with the digital image acquisition unit and is used for analyzing the quantity of fluorescence and the like in the digital image;
the image display unit is connected with the image processing unit and is used for displaying the digital image and the fluorescence quantity; and the image display unit and the image processing unit perform interactive operation and the like.
The camera shooting analysis unit further comprises a data storage unit which is connected with the image processing unit and is used for storing the digital image, the fluorescence quantity and other information.
In the fluorescence imaging analysis system, the excitation light source is an adjustable excitation light source and comprises at least more than one excitation light source with different spectral ranges; the excitation filter is an adjustable excitation filter, and comprises at least more than one excitation filter with different spectral ranges. In a specific embodiment, a plurality of filters with different spectral ranges are arranged on the card slot or the bracket, and the filters with different spectral ranges can be selected by rotating the card slot or the bracket according to actual needs. The installation of the adjustable light source in a fluorescence imaging analysis system can refer to the common method in the prior art.
Similarly, the filter lens is an adjustable filter lens, which includes at least one filter lens with different spectral ranges. In a specific embodiment, a plurality of filter lenses with different spectral ranges are arranged on the clamping groove or the bracket, and the different spectral ranges can be selected by rotating the clamping groove or the bracket according to actual needs.
In the invention, mechanical structures such as the adjustable excitation filter, the adjustable filter lens and the like can be designed according to the prior common technology and are arranged in the fluorescence imaging analysis system by referring to the common method.
The invention also provides a fluorescence imaging analysis method, which comprises the following steps:
step 1: placing the detected object carrying the fluorescent substance on a sample stage, and placing the sample stage in the shell;
step 2: selecting and determining an excitation light source, an excitation filter and a filter lens according to the optical characteristics of the fluorescent substances, arranging the excitation light source and the excitation filter in an excitation device, and arranging the filter lens at the position of a through hole on the shell;
and step 3: starting the excitation light source, wherein light of the excitation light source penetrates through the excitation filter to form monochromatic excitation light, and the excitation light irradiates the detected object on the sample table; the fluorescent substance in the detected object is excited by the exciting light to generate fluorescence;
and 4, step 4: the filter lens filters stray light except fluorescence, and the camera shooting unit captures the fluorescence penetrating through the filter lens to obtain a fluorescence image and a fluorescence numerical value.
In the fluorescence imaging analysis method of the present invention, the fluorescence imaging analysis system is used to detect the actual concentration of the detected object, and the method further comprises the following steps after the step 4:
and 5: drawing a standard curve by using the fluorescence value obtained by detection and the concentration of the detected object to obtain a linear regression equation;
step 6: placing a detected substance which carries a fluorescent substance and has unknown concentration on the sample stage, placing the sample stage in the shell, and repeating the steps 2-4 to obtain a fluorescence image and a fluorescence value of the detected substance;
and 7: and substituting the obtained fluorescence value into the linear regression equation by using the camera analysis unit to calculate the actual concentration of the detected object.
The beneficial effects of the invention include:
the excitation light source adopted in the invention has various advantages: the LED lamp has good monochromaticity, narrow wavelength and large selectivity. For example, the peak of excitation light of green fluorescent protein is 470nm, and if an LED lamp with a peak of 475nm is selected, a small portion of light with a wavelength of 500nm may leak through the color filter. To avoid affecting the measurement, LED lamps with a peak of 453nm were selected to have no light that could leak through the filter. The LED lamp emits light stably, and a stable light source is very important for measurement. And the LED lamp has the advantages of electricity saving, low price and the like.
The present invention employs an excellent optical design, first, monochromaticity of the excitation light, reducing the excitation of other fluorescence of the sample, e.g., with ultraviolet light. Secondly, the application of the scattering light-transmitting sheet enables the illumination on the sample table to be uniform. The black box design inside the system of the present invention excludes the entrance of stray light, all of which provide a very good optical environment for the assay.
In the invention, in the camera shooting analysis component, the digital image acquired by the digital image acquisition unit can be read and stored in real time by the image processing unit. The fluorescence image and quantitative information of the fluorescent substance (e.g., the concentration of the fluorescent substance) can be obtained simultaneously using the imaging unit/component and the imaging analysis unit/component, as compared to a single-function imaging unit (e.g., camera). In addition, dynamic images can be recorded by using the digital image acquisition unit.
In the invention, the power supply voltage required by the LED light source and the camera shooting identification system is low, the energy consumption is low, and the LED light source and the camera shooting identification system can work outdoors and other places without power supplies for a long time by being matched with a lithium battery arranged in an instrument.
Compared with expensive fluorescent instruments in the prior art, the invention does not need to adopt expensive gratings, interference filters and the like, does not need to adopt photomultiplier tubes, measuring electronic circuits and the like, and can realize excellent qualitative and quantitative detection effects. The fluorescence imaging analysis system has the advantages of delicate structural design, simple structure, easy maintenance, easy replacement of accessories and low cost, and is suitable for wide application.
Drawings
Fig. 1 is a schematic configuration diagram of an imaging analysis system.
Fig. 2 is a schematic structural diagram of an imaging analysis system in another embodiment.
Fig. 3 is a schematic structural diagram of an imaging analysis system in another embodiment.
Fig. 4(a) is a sectional view of the housing, and fig. 4(b) is an external view of the housing.
Fig. 5 is a schematic diagram of the imaging analysis unit.
Fig. 6 is a schematic diagram of a hardware circuit in the imaging analysis unit.
Fig. 7 is an image displayed by the image display unit.
Fig. 8(a) is a schematic view of the card slot, and fig. 8(b) is a schematic view of the dial.
FIG. 9 is a line graph of data obtained in example 1 according to the present invention and the prior art.
FIG. 10 is a scatter plot of corresponding readings for different concentrations in example 3.
FIG. 11 is an image displayed by the image display unit in one embodiment.
FIG. 12 is an image displayed by the image display unit in one embodiment.
FIG. 13 shows the mean and standard error of 5 readings at a tube position in an octal tube in this embodiment.
Detailed Description
The present invention will be described in further detail with reference to the following specific examples and the accompanying drawings. The procedures, conditions, experimental methods and the like for carrying out the present invention are general knowledge and common general knowledge in the art except for the contents specifically mentioned below, and the present invention is not particularly limited.
As shown in fig. 1 to 10, 10-shell, 11-sample stage, 12-light hole, 13-pulley, 14-door, 15-card slot, 16-dial 20-excitation device, 21-excitation light source, 22-scattering component, 221-plane mirror, 222-cambered surface mirror, 23-excitation filter, 30-imaging device, 31-filter lens, 32-camera unit, 33-camera analysis unit, 331-digital image acquisition unit, 332-image processing unit, 333-image display unit, 334-data storage unit.
As shown in fig. 1 and 2, the fluorescence imaging analysis system of the present invention includes a housing 10, at least one excitation device 20 and an imaging device 30 disposed inside the housing 10.
The shell 10 is a closed hollow shell, a darkroom is formed in the shell, and a sample table 11 is arranged in the shell 10 and used for placing a detected object carrying fluorescent dye/fluorescent protein; the surface of the shell 10 is provided with a light hole 12, and the light hole 12 is opposite to the sample stage 11 and is used for observing the sample stage 11.
Referring to fig. 1, the number of the excitation devices 20 in this embodiment is one, and the excitation devices 20 are disposed inside the housing 10 and include excitation light sources 21 and excitation filters 23; the light emitted from the excitation light source 21 passes through the excitation filter 23 to form monochromatic excitation light, and the excitation light irradiates the sample stage 11, and the object to be detected emits fluorescence after being excited. In this embodiment, the excitation device 20 is disposed inside the housing 10, the excitation devices 20 are disposed on two sides of the light-transmitting hole 12, and the monochromatic excitation light output by the excitation devices is obliquely irradiated on the object to be detected on the sample stage 11. Referring to fig. 2, the number of the excitation devices 20 is two, and the two excitation devices 20 are symmetrically disposed inside the housing 10. The number of the excitation devices 20 in the invention is not limited, including 1, 2 or more, and the arrangement position of the excitation devices 20 is not limited, and can be adjusted according to the actual installation condition.
The imaging device 30 is arranged outside the housing 10 with respect to the sample stage 11, and includes a filter lens 31 and an imaging unit 32; the filter lens 31 is disposed opposite to the light transmission hole 12 for filtering stray light other than the fluorescent light transmitted through the light transmission hole 12, and the camera unit 32 is disposed opposite to the filter lens 31 for capturing the fluorescent light transmitted through the filter lens 31. The excitation wavelength and the emission wavelength of fluorescence of common green fluorescent dyes are seen in table 1 below.
TABLE 1 common Green fluorescent dye excitation and emission wavelengths
| Name of dye | Excitation wavelength (nm) | Emission wavelength (nm) |
| SYBR Green I/II | 497 | 520 |
| Gel Green | 500 | 520 |
| SYTO 13 | 488 | 509 |
| SYTO 16 | 488 | 518 |
| FITC(Cy3) | 490-495 | 525-530 |
| AlexaFluor | 495 | 519 |
| GFP | 483 | 509 |
| Calcein | 486 | 509 |
In order to form the excitation light of the corresponding wavelength, the excitation light source 21 and the excitation filter 23 are used to form monochromatic excitation light of the corresponding wavelength range. The excitation light source 21 adopts an LED light source, and the spectral range of the LED light source is 445-475 nm. Multiple sets of LEDs, including blue, red, and white LEDs, may be disposed inside the excitation light source 21, and the light sources of various colors are independently controlled by different switches, so as to select light sources in different wavelength ranges. For example, blue LED lamps centered at 453nm can be used to excite green fluorochromes/fluorescent proteins GFP such as SRBY Breen I/II, SYTO 13, FITC (Cy3), Calcein, etc.
Before the detection of the analyte containing the above-mentioned fluorescent dye/fluorescent protein, the types of the excitation light source 21, the excitation filter 23 and the filter lens 31 are determined according to the fluorescent dye/fluorescent protein or other fluorescent substances. Specifically, the excitation filter 23 has a light transmission spectrum range of 325nm to 500nm and the filter lens 31 has a light transmission spectrum range of 500nm to 2500nm for the green fluorescent substance. For example, if the excitation light is blue excitation light, and the blue wavelength is 476-; if the excitation light is green excitation light, the wavelength of the green light is generally 495-570nm, and the selectable model of the corresponding filter lens 31 is golden yellow cut-off glass (JB510, Shanghai organic optical glass factory, lambda tj510 +/-10 nm) or JB490 (lambda tj490 +/-10 nm), the stray light with the wavelength below 500nm can be filtered, and the filter lens can be used for detecting green fluorescent dye/fluorescent protein by matching with a blue LED lamp; if the excitation light is yellow excitation light, the yellow light wavelength is generally 570-590nm, and the selectable type of the corresponding filter lens 31 is CB565(λ tj565 + -10 nm) or CB580(λ tj580 + -10 nm); the red light wavelength is 620-750nm, and HB610/620/630 can be selected as the corresponding optical filter.
In the present invention, the relationship between the optical properties of the fluorescent substance, the excitation light source, the excitation filter, and the filter is shown in table 2 below.
TABLE 2 relationship of optical Properties among fluorescent substance, excitation light source, excitation filter, and filter
As shown in fig. 2, a scattering member 22 is further disposed between the excitation light source 21 and the excitation filter 23, the scattering member 22 is a plane mirror, light emitted from the excitation light source 21 passes through the scattering member 22 to form a uniform light beam to irradiate the excitation filter 23, so that uniform excitation light is generated to irradiate the sample stage 11, and the object to be detected placed on the sample stage is irradiated with uniform illumination intensity.
In another preferred embodiment of the present invention, the scattering component 22 is a plane mirror 221 and a cambered mirror 222, as shown in fig. 3, the plane mirror 221 is used for reflecting the light of the excitation light source 21 to the cambered mirror 222, and the cambered mirror 222 forms a uniform light beam to be reflected to the excitation filter 23.
As shown in fig. 4(a) and 4(b), the housing 10 is a closed darkroom, and for conveniently placing the object to be detected on the sample stage 11, an openable and closable door 14 is opened at a side of the housing 10, and a sliding groove (not shown) is provided at the bottom of the sample stage 11 and is matched with the sliding wheel 13. The sample stage 11 is moved along the slide groove by the pulley 13, and the housing 10 is removed from the door 14 to facilitate placing of the object to be detected. After the object is placed, the sliding slot is slid into the interior of the housing 10, and the door 14 is closed to keep the housing 10 in a closed dark room.
As shown in fig. 5, the imaging device 30 further includes a camera analysis unit 33. The camera unit 32 is connected to a camera analysis unit 33, and the camera analysis unit 33 is used for capturing fluorescence and measuring and analyzing the concentration of fluorescent substances including fluorescent dyes or fluorescent proteins. Referring to fig. 5, the camera analysis unit 33 includes a digital image acquisition unit 331, an image processing unit 332, and an image display unit 333. The digital image acquisition unit 331 is used to capture fluorescence to obtain a digital image. The image processing unit 332 is connected to the digital image acquisition unit 331 for analyzing the amount of fluorescence in the digital image. The image display unit 333 is connected to the image processing unit 332, and the image display unit 333 can simultaneously display the digital image and the amount of fluorescence thereof. The camera analysis unit 33 is further provided with a data storage unit 334, and the data storage unit 334 is connected to the image processing unit 332 for storing the acquired digital image and the amount of fluorescence.
In the preferred embodiment of the present invention, the digital image acquisition unit 331 is OV7670 manufactured by OV (Omni Vision) Inc. Referring to fig. 6, the image processing unit 332 adopts a micro-processing chip of STM32F103RCT6 model with ARM V7 architecture, the hardware platform is an ALIENTEK MiniSTM32 development board, the image processing unit 332 controls the OV7670 to acquire digital images, and the acquired images are processed to obtain the fluorescence quantity of fluorescent substances in the digital images. The image display unit 333 is an ALIENTEK3.5 inch TFT LCD module. Referring to fig. 7, the image display unit 333 displays the processed digital image and the fluorescence quantity (RGB value), and finally displays negative/positive with reference to the previously collected sample reading. Data storage unit 334 is a Kingston 4GB SD card.
The imaging analysis unit 33 is provided integrally with the imaging unit 32 and is attached to the outside of the housing 10. The shooting analysis unit 33 is started, then the measurement waiting state can be entered, the automatic measurement can be carried out by pressing a 'start' button, the measurement takes about 3 seconds, whether the measurement result is stored in the data storage unit 334 or not can be selected after the measurement is finished, if yes, the measurement result is stored in the Kingston 4GB SD card, and then the next measurement waiting state is entered; if the measurement result is not satisfactory, if "no" is selected, the measurement result is discarded, and the imaging and analyzing unit 33 automatically enters the next measurement waiting state. After the measurement is finished, the stored measurement result can be searched from the Kingston 4GB SD card of the device and can be imported to any other storage equipment.
The adjustable excitation light source comprises at least one LED light source with different spectral properties, and the LED light source is integrated in the excitation device 20, and the on and off of the LED light source with corresponding color is controlled by a switch.
The adjustable excitation filter is adjusted by means of a card slot 15 or a dial 16. Fig. 8(a) shows a clamping groove type adjustable excitation filter, and when the excitation filter is replaced, the excitation filter is pulled out from the clamping groove 15, and the excitation filter in the required spectral range is inserted into the clamping groove 15 and fixed. Fig. 8(b) shows a dial-type adjustable excitation filter, in which a plurality of excitation filters are disposed on a circular dial plate 16, and the dial plate 16 is rotated by a rotating shaft or the like to rotate the excitation filters in the corresponding spectral ranges below the excitation light sources 21. The adjustable filter lens is also realized by the clamping groove 15 or the dial 16. The structures of the adjustable excitation filter and the adjustable filter lens can be designed according to the prior art.
Example 1
1. Constructing and culturing Escherichia coli carrying fluorescent protein expression vector, wherein the fluorescent protein is named as GFP (green fluorescent protein), and 395nm and 475nm are maximum and second maximum excitation wavelengths respectively.
2. Quantitative measurement of excitation light:
step 1: a series of E.coli carrying fluorescent protein expression vectors of known concentration and diluted in a gradient were sequentially placed on a sample stage, and one sample was tested at a time. The detected object carries fluorescent substance, and the sample stage is placed in the shell;
step 2: the excitation light source 21 is a blue LED lamp; the excitation filter 23 is ZB1 type, and the light transmission spectrum range is 300-500 nm; the filter lens 31 is JB510 type, and has a light transmission spectrum range of 510nm +/-10 nm-2500 nm. Selecting and determining an excitation light source, an excitation filter and a filter lens according to the optical characteristics of the fluorescent substance, arranging the excitation light source and the excitation filter in an excitation device, and arranging the filter lens at the position of a through hole on the shell;
and step 3: starting an excitation light source, wherein light rays of the excitation light source penetrate through an excitation filter to form monochromatic excitation light, and the excitation light irradiates the detected object on the sample table; the fluorescent substance in the detected object is excited by the exciting light to generate fluorescence;
and 4, step 4: the filter lens filters stray light except fluorescence, and the camera shooting unit captures the fluorescence penetrating through the filter lens to obtain a fluorescence image and a fluorescence numerical value;
meanwhile, the Escherichia coli carrying the fluorescent protein expression vector is placed in a fluorescence spectrophotometer in the prior art; the selected fluorescence spectrophotometer is Varian Cary Eclipse, the adopted exciting light is blue, and the wavelength range is 485nm +/-2.5 nm.
3. And (3) correlation analysis of quantitative data and a fluorescence spectrophotometer detection result:
referring to Table 3 below and FIG. 9, the dots in FIG. 9 represent different concentrations of green fluorescent protein, the lines represent linear regression between the readings of the two instruments, and the two detection methods show good correlation (R)20.98). Therefore, the detection result of the fluorescence imaging analysis system has the same linear relation with the existing device, and the verification can be effectively applied to the fluorescence detection experiment.
Table 3: detection value of two detection devices on fluorescence signal
Example 2
In the present embodiment, the imaging unit 32 and the imaging analysis unit 33 are used. The camera unit 32 is a digital camera, and real-time measurement and analysis are performed by a camera analysis unit 33 connected to the digital camera. The purpose of this example was to determine the relationship between green fluorescent protein concentration and reading over a relatively large concentration range. The measurement conditions of this example, the shooting conditions of the digital camera (including aperture and exposure time), the kind of green fluorescent protein and its corresponding filter were the same as in example 1, wherein the linear regression equation for determining the actual concentration was determined from example 1 above, and the linear regression equation was: 0.0532x +16.628, R20.9803. And if the linear regression equation is unknown, repeating the steps 2-4 to draw a standard curve to obtain the linear regression equation.
In this example, the relationship between the concentration and the reading is verified by adjusting the concentration of green fluorescent protein and measuring the reading of green fluorescent wearing white at different concentrations, and the specific process is as follows:
and 5: drawing a standard curve by using the fluorescence value detected in the above example 1 and the concentration of the escherichia coli to obtain a linear regression equation;
step 6: and (3) placing the colibacillus substance carrying the fluorescent substance and having unknown concentration on a sample table, placing the sample table in the shell, and repeating the steps 2-4 to obtain a fluorescence image and a fluorescence value of the colibacillus.
And 7: the fluorescence value obtained is substituted into the linear regression equation by the camera analysis unit 33, and the actual concentration of the Escherichia coli is calculated.
Through the above steps 5 to 7, the test objects with different concentrations (GFP amounts) were subjected to imaging analysis, and the obtained data are shown in table 4 below and fig. 10. The results show that the green fluorescent protein concentration is in a good linear relationship with the reading. The fluorescence imaging analysis system of the present invention requires a linear assay to determine the linear range of different fluorochromes before each fluorochrome is measured. If the conventional detection is performed on the fluorescent substance under the fixed detection condition, the steps 2-4 are not required to be adjusted after the initial setting, and the linear regression equation obtained does not need to be adjusted.
TABLE 4 Green fluorescent protein reading and concentration relationship under the same photographing conditions
| Reading number | Concentration of |
| 32 | 1 |
| 62 | 2 |
| 84 | 3 |
| 102 | 4 |
| 116 | 5 |
| 130 | 6 |
| 139 | 7 |
Example 3
In this embodiment, the imaging unit 32 and the highly integrated imaging and analyzing unit 33 are used to capture fluorescence and measure and analyze the fluorescent dye/fluorescent protein concentration. The camera unit 32 and the camera analysis unit 33 are integrally arranged, and the working mode of combining a traditional digital camera and a computer is replaced. The camera unit 32 and the camera analysis unit 33 which are integrally arranged are integrated, so that the volume and the weight of the system are greatly reduced, the manufacturing cost is reduced on the premise of meeting the performance of the system, and the system is convenient to use.
In the embodiment, SYBR Green I is combined with DNA to emit fluorescence for analysis, and an eppendorf tube containing SYBR Green I + DNA solution is placed in the fluorescence imaging analysis system. Wherein the excitation light source 21 is a blue LED lamp; the excitation filter 23 is ZB1 type, and the light transmission spectrum range is 300-500 nm; the filter lens 31 is JB510 type, and the light transmission spectrum range thereof is 510nm +/-10 nm-2500 nm; the digital image acquisition unit 331 employs OV7670 of OV (omni vision) corporation. The image processing unit 332 adopts a microprocessor chip of an ARM V7-structured STM32F103RCT6 model, a hardware platform is an ALIENTEK MiniSTM32 development board, the image processing unit 332 controls OV7670 to collect digital images and process the collected images to obtain an average RGB value of the digital images, and finally, the average RGB value is 149 by referring to a set sample reading MARK, and the RGB value is higher than MARK and shows positive. The image display unit 333 is an ALIENTEK3.5 inch TFT LCD module. The image display unit 333 displays the processed digital image with RGB values. As shown in FIG. 11, the highlighted area in the figure is the excitation light generated by the green fluorescent substance in the test object, and it is shown that the negative/positive result is positive with reference to the set sample reading "149" as MARK (indicated by yang! in FIG. 11).
Example 4
In this example, the detection hardware platform was the same as that of example 3, and various analyses were performed by fluorescence emitted from SYBR Green I in combination with DNA. An Eppendorf octaplex containing a SYBR Green I + DNA solution was placed in the fluorescence imaging assay system of the invention. The image processing unit 332 automatically divides the acquired image equally into 8 parts, calculates the fluorescence signal of each part, and gives a fluorescence value. As shown in FIG. 12, the highlighted area in the graph is the excitation light generated by the green fluorescent substance in the test object, and the negative/positive result is shown with reference to the set sample reading as MARK. The same octant was read in 5 replicates and the results are shown in table 5 and fig. 13, where the dots in fig. 13 represent the average of 5 readings at one tube position in the octant and the score lines on the dots represent the standard error of the 5 readings. The results of 5 times of each tube position are positive, and good repeatability is shown in qualitative analysis.
TABLE 5 statistics of 5 readings from the same octal tube
The protection of the present invention is not limited to the above embodiments. Variations and advantages that may occur to those skilled in the art may be incorporated into the invention without departing from the spirit and scope of the inventive concept, and the scope of the appended claims is intended to be protected.
Claims (14)
1. A fluorescence imaging analysis system, comprising a housing (10), at least one excitation device (20) and an imaging device (30);
the shell (10) is a closed hollow shell, a darkroom is formed in the shell, and a sample table (11) is arranged in the shell (10) and used for placing a detected object carrying fluorescent substances; a light-transmitting hole (12) is formed in the shell (10), and the light-transmitting hole (12) is right opposite to the sample table (11) and is used for observing the sample table (11);
the excitation device (20) is arranged inside the shell (10) and comprises an excitation light source (21) and an excitation filter (23); light rays emitted by the excitation light source (21) form monochromatic excitation light through the excitation filter (23) and irradiate the sample table (11); the detected object emits fluorescence after being excited;
the imaging device (30) comprises a filter lens (31) and an image pickup unit (32); wherein the filter lens (31) is arranged right opposite to the light transmission hole (12) and is used for filtering stray light except fluorescent light which penetrates through the light transmission hole (12); the imaging unit (32) is disposed right opposite to the filter lens (31) and captures fluorescence transmitted through the filter lens (31).
2. The fluorescence imaging analysis system of claim 1, wherein said fluorescent species comprises a fluorescent dye, a fluorescent protein.
3. The fluorescence imaging analysis system according to claim 1, wherein the selection of the corresponding excitation light source (21), excitation filter (23), and filter lens (31) is determined according to the optical properties of the fluorescent substance; wherein the cut-off wavelength of the excitation filter (23) is between the excitation wavelength and the emission wavelength of the fluorescent substance.
4. The fluorescence imaging analysis system according to claim 1, wherein when the fluorescent substance is a green fluorescent substance, the spectral range of the excitation light source (21) is 445 to 475 nm; the light transmission spectrum range of the excitation filter (23) is 325 nm-500 nm; the light transmission spectrum range of the filter lens (31) is 500 nm-2500 nm; or,
when the fluorescent substance is a red fluorescent substance, the spectral range of the excitation light source (21) is 585nm +/-29 nm; the light transmission spectrum of the excitation filter (23) is 500 nm-620 nm; the light-transmitting spectrum range of the filter lens (31) is 620 nm-2500 nm; or,
when the fluorescent substance is a yellow fluorescent substance, the spectral range of the excitation light source (21) is 531nm +/-40 nm; the light transmission spectrum of the excitation filter (23) is 350 nm-580 nm; the light-transmitting spectrum range of the filter lens (31) is 580 nm-2500 nm; or,
when the fluorescent substance is a blue fluorescent substance, the spectral range of the excitation light source (21) is 357nm +/-44 nm; the light transmission spectrum of the excitation filter (23) is 280 nm-410 nm; the light transmission spectrum range of the filter lens (31) is 410 nm-2500 nm.
5. The fluorescence imaging analysis system of claim 1, wherein a scattering member (22) is further disposed in the excitation device (20); light rays emitted by the excitation light source (21) form uniform light beams after passing through the scattering component (22) and irradiate the uniform light beams to the excitation filter (23).
6. The fluorescence imaging analysis system of claim 5, wherein the scattering component (22) is a planar scattering mirror.
7. The fluorescence imaging analysis system according to claim 5, wherein the scattering component (22) comprises a plane mirror (221) and a cambered mirror (222), the plane mirror (221) is used for reflecting the light of the excitation light source (21) to the cambered mirror (222), and the cambered mirror (222) forms a uniform light beam to be reflected to the excitation filter (23).
8. The fluorescence imaging analysis system according to claim 1, wherein the bottom of the sample stage (11) is provided with a slide rail for moving the sample stage (11) out of the housing (10).
9. The fluorescence imaging analysis system according to claim 1, wherein the imaging device (30) further comprises a camera analysis unit (33), the camera analysis unit (33) being connected to the camera unit (32) for measuring and analyzing the concentration of the fluorescent substance.
10. The fluorescence imaging analysis system according to claim 9, wherein the camera analysis unit (33) comprises:
a digital image acquisition unit (331) for capturing the fluorescence transmitted through the filter lens (31) to obtain a digital image;
an image processing unit (332), connected to the digital image acquisition unit (331), for analyzing the amount of fluorescence in the digital image;
and the image display unit (333) is connected with the image processing unit (332) and is used for displaying the digital image and the fluorescence quantity.
11. The fluorescence imaging analysis system of claim 10, wherein the camera analysis unit (33) further comprises a data storage unit (334) connected to the image processing unit (332) for storing the digital image, the amount of fluorescence.
12. The fluorescence imaging analysis system according to claim 1, wherein the excitation light source (21) is an adjustable excitation light source, which includes at least one excitation light source with different spectral ranges; the excitation filter (23) is an adjustable excitation filter, and comprises at least one excitation filter with different spectral ranges; the filter lens (31) is an adjustable filter lens, and comprises at least one filter lens with different spectral ranges.
13. A fluorescence imaging analysis method using the fluorescence imaging analysis system according to any one of claims 1 to 12, comprising the steps of:
step 1: placing the detected object carrying the fluorescent substance on a sample table (11);
step 2: determining a corresponding excitation light source (21), an excitation filter (23) and a filter lens (31) according to the fluorescent substances;
and step 3: the excitation light source (21) is started, light of the excitation light source (21) penetrates through the excitation filter (23) to form monochromatic excitation light, and the excitation light irradiates on the detected object; exciting the fluorescent substance in the detected object to generate fluorescence;
and 4, step 4: the filter lens (31) filters stray light except the fluorescence, and the fluorescence transmitted through the filter lens (31) is captured by the camera unit (32) to obtain a fluorescence image and a fluorescence numerical value.
14. The fluorescence imaging analysis method of claim 13, wherein the method is used for detecting the concentration of the detected object, and the step 4 further comprises the following steps:
and 5: drawing a standard curve by using the fluorescence value obtained by detection and the concentration of the detected object to obtain a linear regression equation;
step 6: placing a detected substance which carries a fluorescent substance and has unknown concentration on the sample stage (11), placing the sample stage (11) in the shell (10), and repeating the steps 2-4 to obtain a fluorescence image and a fluorescence value of the detected substance;
and 7: and calculating the actual concentration of the detected object by substituting the obtained fluorescence value into the linear regression equation by the image pickup analysis unit (33).
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108444878A (en) * | 2018-04-20 | 2018-08-24 | 浙江大学 | A kind of portable aviation sprays the mist droplet deposition measurement of effectiveness device and method of operation |
| CN108535221A (en) * | 2017-03-03 | 2018-09-14 | 广州博鹭腾仪器仪表有限公司 | A kind of new type gel imaging system |
| CN108627482A (en) * | 2017-03-20 | 2018-10-09 | 上海溯源生物技术有限公司 | Fluorescence detection device and its fluorimetric analysis method based on mobile terminal |
| CN110823856A (en) * | 2019-12-19 | 2020-02-21 | 北京永安多谱检测科技有限公司 | Excited fluorescence peroxide detection device based on image analysis and detection method thereof |
| CN111426671A (en) * | 2020-05-22 | 2020-07-17 | 南京诺源医疗器械有限公司 | Imaging system based on fluorescence analysis |
| CN111487037A (en) * | 2020-04-16 | 2020-08-04 | 中国科学院微电子研究所 | Light source uniformity detection system and method |
| CN113702397A (en) * | 2020-05-20 | 2021-11-26 | 深圳中科飞测科技股份有限公司 | Optical detection system and optical detection method |
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-
2015
- 2015-04-17 CN CN201510186113.XA patent/CN104764727A/en active Pending
Non-Patent Citations (3)
| Title |
|---|
| 姜淑媛等: "茶树荧光性绿斑病叶黄绿色自发荧光的显微分析", 《茶叶科学》 * |
| 李晨: "悬浮式生物芯片检测系统中光学系统的研究", 《万方学位论文》 * |
| 马钰慧: "紫外荧光成像系统的研制", 《中国优秀硕士学位论文全文数据库 基础科学辑》 * |
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