WO2020192755A1 - 生物样品膜上的自发光物体的成像方法及装置 - Google Patents

生物样品膜上的自发光物体的成像方法及装置 Download PDF

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Publication number
WO2020192755A1
WO2020192755A1 PCT/CN2020/081617 CN2020081617W WO2020192755A1 WO 2020192755 A1 WO2020192755 A1 WO 2020192755A1 CN 2020081617 W CN2020081617 W CN 2020081617W WO 2020192755 A1 WO2020192755 A1 WO 2020192755A1
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Prior art keywords
field image
self
dark field
biological sample
luminous object
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PCT/CN2020/081617
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English (en)
French (fr)
Inventor
张英豪
奚岩
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上海易孛特光电技术有限公司
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Priority to US17/599,529 priority Critical patent/US12032149B2/en
Priority to EP20777616.2A priority patent/EP3951356A4/en
Publication of WO2020192755A1 publication Critical patent/WO2020192755A1/zh

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/36—Microscopes arranged for photographic purposes or projection purposes or digital imaging or video purposes including associated control and data processing arrangements
    • G02B21/365—Control or image processing arrangements for digital or video microscopes
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
    • G01N21/76—Chemiluminescence; Bioluminescence
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/06—Means for illuminating specimens
    • G02B21/08—Condensers
    • G02B21/10—Condensers affording dark-field illumination
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
    • G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
    • G01N21/27—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands using photo-electric detection ; circuits for computing concentration
    • G01N21/274—Calibration, base line adjustment, drift correction

Definitions

  • the invention relates to the technical field of sample analysis, in particular to a method and device for imaging a self-luminous object on a biological sample film.
  • bioluminescence detection technologies there are two types of bioluminescence detection technologies commonly used, one is the use of photosensitive film technology, and the other is the use of camera shooting technology.
  • the photosensitive film technology is similar to the traditional photographic film washing method.
  • the self-luminous object (attached to the biological sample film) is closely attached to the photosensitive film, and then the photosensitive film is rewashed with a developer.
  • the image corresponding to the self-luminous object is obtained; however, this imaging method has a complicated operation process and requires more steps for digital processing; the camera shooting technology is similar to that of a mobile phone, and the camera and the self-luminous object are placed in a dark room.
  • This method can directly store digital images.
  • the collection rate of the light signal is likely to be low. Therefore, it takes a long time for data collection and has defects such as low sensitivity.
  • both of the above-mentioned imaging systems have disadvantages such as large volume, large space occupation, high cost, and difficulty in handling.
  • the technical problem to be solved by the present invention is that the imaging method for acquiring self-luminous objects on the biological sample film in the prior art has complicated operation process, long imaging time, and the imaging system has large volume, large space occupation, high cost, and difficult to handle. Other defects, the purpose is to provide a method and device for imaging self-luminous objects on biological sample films.
  • the invention provides an imaging device for a self-luminous object on a biological sample film, the imaging device comprising a housing, a photoelectric conversion element and an image correction device;
  • the inside of the shell constitutes a darkroom space
  • the photoelectric conversion element is arranged in the housing;
  • the photoelectric conversion element is used to obtain a first dark field image inside the housing within a set collection period when the biological sample film is not placed;
  • a self-luminous object is carried on the biological sample film
  • the photoelectric conversion element is further used to obtain a second dark field image inside the housing within the set collection time;
  • the image correction device is configured to perform correction processing on the second dark field image according to the first dark field image, and obtain a target image corresponding to the self-luminous object.
  • the image correction device corrects the second dark field image according to the first dark field image, and the formula corresponding to the target image corresponding to the self-luminous object is obtained as follows:
  • I represents the pixel data corresponding to the target image
  • I 0 represents the pixel data corresponding to the second dark field image
  • I d represents the pixel data corresponding to the first dark field image
  • the imaging device further includes a light source device, and the light source device is provided in the housing.
  • the photoelectric conversion element is also used to obtain the set collection time period after the biological sample film is not put into the housing and the light source device in the darkroom space is turned on Bright field image inside the housing;
  • the image correction device is further configured to perform correction processing on the second dark field image according to the first dark field image and the bright field image to obtain a target image corresponding to the self-luminous object.
  • the photoelectric conversion element is also used to obtain the set collection when the biological sample film is not placed in the housing, the housing is in an open state, and an external light source provides a uniform light field. Corresponding bright field image within the duration;
  • the image correction device is further configured to perform correction processing on the second dark field image according to the first dark field image and the bright field image to obtain a target image corresponding to the self-luminous object.
  • the image correction device corrects the second dark field image according to the first dark field image and the bright field image, and the formula corresponding to the target image corresponding to the self-luminous object is obtained as follows:
  • I represents the pixel data corresponding to the target image
  • I 0 represents the pixel data corresponding to the second dark field image
  • I d represents the pixel data corresponding to the first dark field image
  • I f represents the bright field The pixel data corresponding to the image.
  • the housing includes a light-shielding cover and a base;
  • One side of the shading cover is hinged to one side of the base
  • the inside of the housing constitutes the dark room space.
  • the light source device is arranged at a top position inside the light shielding cover; and/or,
  • the corresponding lighting duration after the light source device is turned on is 10ms-30s; and/or,
  • the light source equipment includes several LED (light emitting diode) lamp beads arranged in a lattice, several lamps guided by optical fibers, several parallel-arranged lamp tubes or several plate-shaped lamps.
  • the imaging device further includes a diffuser
  • the light scattering plate is fixedly arranged inside the light shielding cover and directly below the light source device; and/or,
  • the photoelectric conversion element includes a CMOS (complementary metal oxide semiconductor) chip, a CCD (charge coupled device) chip or an amorphous silicon photoelectric conversion detector; and/or,
  • the biological sample membrane includes a protein membrane, agarose gel block, agarose gel strip, polyacrylamide gel block or polyacrylamide gel strip.
  • the imaging device further includes a protective film
  • the two sides of the protective film are respectively attached to the biological sample film and the photoelectric conversion element.
  • the thickness of the protective film is 0.01mm-0.2mm; and/or,
  • the material of the protective film is tempered glass film or hard plastic film.
  • the present invention also provides a method for imaging a self-luminous object on a biological sample film.
  • the imaging method is implemented by using the above-mentioned imaging device for a self-luminous object on a biological sample film.
  • the imaging method includes:
  • the photoelectric conversion element After the biological sample film is attached to the surface of the photoelectric conversion element, the photoelectric conversion element is used to obtain a second dark field image inside the housing within the set collection time;
  • a self-luminous object is carried on the biological sample film
  • the formula corresponding to the step of performing correction processing on the second dark field image according to the first dark field image and obtaining the target image corresponding to the self-luminous object is as follows:
  • I represents the pixel data corresponding to the target image
  • I 0 represents the pixel data corresponding to the second dark field image
  • I d represents the pixel data corresponding to the first dark field image
  • the imaging device further includes a light source device
  • the method further includes:
  • the photoelectric conversion element is used to obtain the bright field inside the housing within the set collection time after the biological sample film is not placed in the housing and the light source device in the darkroom space is turned on image;
  • the step of performing correction processing on the second dark field image according to the first dark field image, and obtaining a target image corresponding to the self-luminous object includes:
  • the method before the step of performing correction processing on the second dark field image according to the first dark field image, and obtaining the target image corresponding to the self-luminous object, the method further includes:
  • the photoelectric conversion element is used to obtain the corresponding bright field within the set collection time when the biological sample film is not placed in the housing, the housing is in an open state and an external light source provides a uniform light field image;
  • the step of performing correction processing on the second dark field image according to the first dark field image, and obtaining a target image corresponding to the self-luminous object includes:
  • the formula corresponding to the step of performing correction processing on the second dark field image according to the first dark field image and the bright field image, and obtaining the target image corresponding to the self-luminous object is as follows:
  • I represents the pixel data corresponding to the target image
  • I 0 represents the pixel data corresponding to the second dark field image
  • I d represents the pixel data corresponding to the first dark field image
  • I f represents the bright field The pixel data corresponding to the image.
  • the interior of the housing in the image acquisition device of the present invention constitutes a dark room space.
  • the photoelectric conversion element first obtains the first dark field image, and after the self-luminous object is placed, the photoelectric conversion element obtains the second dark field image.
  • Second dark field image and then correct the second dark field image according to the first dark field image to obtain the target image corresponding to the self-luminous object; in addition, it can be combined with the light field generated by the light source device in the housing or the light field generated by an external light source Under the bright field image, the target image of the self-luminous object with higher definition can be obtained more effectively and accurately.
  • the operation process is simple, the imaging time is shorter, and the imaging device has a small structure, low manufacturing cost, and convenient operation. And convenient to carry and other advantages.
  • FIG. 1 is a schematic diagram of a first structure of a device for imaging a self-luminous object on a biological sample film in Example 1 of the present invention.
  • FIG. 2 is a schematic diagram of a second structure of the imaging device for a self-luminous object on a biological sample film in Embodiment 1 of the present invention.
  • FIG. 3 is a schematic diagram of the structure of the first housing in the imaging device of the self-luminous object on the biological sample film in Embodiment 1 of the present invention.
  • FIG. 4 is a schematic structural diagram of a second housing in the imaging device of a self-luminous object on a biological sample film according to Embodiment 1 of the present invention.
  • FIG. 5 is a schematic diagram of the first dark field imaging in the imaging device of the self-luminous object on the biological sample film according to Embodiment 1 of the present invention.
  • FIG. 6 is a schematic diagram of the second dark field imaging in the imaging device of the self-luminous object on the biological sample film of Embodiment 1 of the present invention.
  • FIG. 7 is a schematic diagram of a first structure of an imaging device for a self-luminous object on a biological sample film according to Embodiment 2 of the present invention.
  • FIG. 8 is a schematic diagram of a second structure of an imaging device for a self-luminous object on a biological sample film according to Embodiment 2 of the present invention.
  • FIG. 9 is a schematic diagram of a third structure of an imaging device for a self-luminous object on a biological sample film according to Embodiment 2 of the present invention.
  • FIG. 10 is a schematic diagram of bright field imaging in the imaging device of a self-luminous object on a biological sample film according to Embodiment 2 of the present invention.
  • FIG. 11 is a schematic diagram of the relationship between the lane and the signal intensity in the imaging device of the self-luminous object on the biological sample film of Embodiment 2 of the present invention.
  • FIG. 12 is a schematic diagram of second dark field imaging in the imaging device for self-luminous objects on the biological sample film of Embodiment 2 of the present invention.
  • FIG. 13 is a schematic structural diagram of an imaging device for a self-luminous object on a biological sample film according to Embodiment 3 of the present invention.
  • FIG. 14 is a flowchart of a method for imaging a self-luminous object on a biological sample film according to Embodiment 4 of the present invention.
  • FIG. 15 is a flowchart of a method for imaging a self-luminous object on a biological sample film according to Embodiment 5 of the present invention.
  • FIG. 16 is a flowchart of a method for imaging a self-luminous object on a biological sample film according to Embodiment 6 of the present invention.
  • the imaging device of the self-luminous object on the biological sample film of this embodiment includes a housing 1, a photoelectric conversion element 2 and an image correction device 3.
  • the inside of the housing 1 constitutes a dark room space.
  • the housing 1 includes a light shielding cover 4 and a base 5. As shown in FIG. 3, one side of the light shielding cover 4 and one side of the base 5 are hinged, and the light shielding cover 4 is in an open state.
  • the inside of the housing 1 constitutes a dark room space to ensure that the weak signal corresponding to the self-illuminating object on the biological sample film (indicated by A in Figure 2) is effectively collection.
  • biological sample membranes include protein membranes, agarose gel blocks, agarose gel strips, polyacrylamide gel blocks, polyacrylamide gel strips, and the like.
  • the photoelectric conversion element is arranged in the casing.
  • the photoelectric conversion element includes a CMOS chip, a CCD chip, or an amorphous silicon photoelectric conversion detector.
  • the photoelectric conversion element is used to obtain the first dark field image inside the inner housing for a set collection time when the biological sample film A is not placed (as shown in Figure 5);
  • a self-luminous object is carried on the biological sample film.
  • the set collection time can be set and adjusted according to actual conditions.
  • the set collection time is 1s.
  • the biological sample membrane includes a protein membrane
  • the protein membrane sample is separated by SDS-PAGE (sodium dodecyl sulfonate-polyacrylamide gel) electrophoresis technology, and the protein is transferred to A biological sample membrane carrying protein is obtained on the PVDF (polyvinylidene fluoride) membrane.
  • the biological sample membrane is sequentially incubated with the target protein-specific primary antibody and horseradish peroxidase-coupled secondary antibody. Then use the chemiluminescent liquid for color development (that is, display as a self-luminous object), and generate a chemiluminescence signal in the region of the target protein.
  • the photoelectric conversion element After the biological sample film is attached to the surface of the photoelectric conversion element, the photoelectric conversion element is also used to obtain a second dark field image (as shown in Figure 6) inside the inner housing for a set collection time, that is, the first dark field image and The data collection time corresponding to the second dark field image is the same.
  • the image correction device is used to perform correction processing on the second dark field image according to the first dark field image to obtain a target image corresponding to the self-luminous object.
  • the photoelectric conversion element is used to collect the value I d of each pixel in the uniform dark field inside the housing, and the collection time is 1s.
  • the photoelectric conversion element is used to collect the light signal corresponding to the self-luminous object (target protein) on the biological sample membrane (at this time, the other areas except the target protein belong to the uniform dark field).
  • the value of each pixel I 0 where the acquisition time is 1s.
  • the image correction device performs correction processing on the second dark field image according to the first dark field image, and the formula corresponding to the target image corresponding to the self-luminous object is obtained as follows:
  • I represents the pixel data corresponding to the target image
  • I 0 represents the pixel data corresponding to the second dark field image
  • I d represents the pixel data corresponding to the first dark field image
  • the size of the second dark field image is the same as that of the first dark field image, and the difference of I 0 -I d represents the difference of the pixel values of the same point in the two images.
  • the housing in the image acquisition device constitutes a dark room space.
  • the photoelectric conversion element first obtains the first dark field image.
  • the photoelectric conversion element obtains the first dark field image.
  • the second dark field image, and then the second dark field image is corrected according to the first dark field image, and the target image corresponding to the self-luminous object is obtained, so as to obtain the target of the self-luminous object with higher definition through contact imaging Image, the operation process is simple, the imaging time is short, and the imaging device has the advantages of small structure, low manufacturing cost, convenient operation and convenient carrying.
  • the imaging device of the self-luminous object on the biological sample film of this embodiment is a further improvement of embodiment 1, specifically:
  • the imaging device further includes a light source device 6 which is provided in the housing 1.
  • the light source device 6 is arranged at the top position inside the light shielding cover 4;
  • the corresponding lighting duration after the light source device 6 is turned on is 10 ms-30s, and the lighting duration can also be adjusted according to actual needs.
  • the light source device 6 includes several LED lamp beads arranged in a lattice, several lamps guided by optical fibers, several parallel lamp tubes, several plate-shaped lamps, etc.
  • the imaging device also includes a diffuser plate 7, which is fixed on the inner side of the light shielding cover and directly below the light source device, so that the light source device 6 emits uniform light.
  • the imaging device further includes a protective film 8.
  • the two sides of the protective film 8 are attached to the biological sample film and the photoelectric conversion element respectively.
  • the thickness of the protective film is 0.01mm-0.2mm
  • the material of the protective film is tempered glass film or hard plastic film.
  • the outer edge of the protective film 8 is located outside the outer edge of the photoelectric conversion element 2.
  • the photoelectric conversion element 2 includes a detector body 9 and a substrate 10, and the outer edge of the substrate 10 is located outside the outer edge of the detector body 9.
  • the substrate is a metal plate, a PCBA (a circuit board manufacturing process) board, etc.
  • the top of the base 5 has a containing groove 11, and the photoelectric conversion element 2 and the protective film 8 are located in the containing groove 11.
  • the imaging device of this embodiment further includes a packaging member 12 that acts on the protective film 8 and the base 5 to restrict the movement of the photoelectric conversion element 2 relative to the base 5.
  • the package 12 includes a filler 13 and a card board 14.
  • the filler 13 is located in the area enclosed by the photoelectric conversion element 2, the protective film 8 and the surface of the base 5, and is used to fix the photoelectric conversion element 2 in the base 5.
  • the filler is gel, glass powder, plastic embryo, etc.
  • the card board 14 abuts on the peripheral edge of the protective film 8 to ensure the stability of the entire device.
  • the base 5 includes a bottom plate 15 and a base body 16.
  • the base body 16 is provided with a containing groove 11, and the containing groove 11 penetrates through the base body 16, and the bottom plate 15 is movably connected to the bottom of the base body 16 from the bottom of the base body 16. Open or close the containing tank 11.
  • the height of the photoelectric conversion element 2 is not greater than the depth of the containing groove 11.
  • the photoelectric conversion element 2 is also used to obtain the bright field image inside the casing within the set collection time after the biological sample film is not put into the casing 1 and the light source device 6 in the dark room space is turned on, that is, the first dark field image,
  • the data collection time corresponding to the second dark field image and the bright field image are the same.
  • the difference between this figure and the first dark field image in Fig. 5 is that its overall brightness is higher than the overall brightness of the first dark field image, where the smaller the number of diagonal lines, the higher the corresponding brightness.
  • the image correction device 3 is further configured to perform correction processing on the second dark field image according to the first dark field image and the bright field image, and obtain the target image corresponding to the self-luminous object.
  • the light source device is controlled to be turned on (the corresponding light duration is 10ms-30s), and the photoelectric conversion element 2 is used to synchronously collect the value I f1 of each pixel in the uniform bright field, where the collection time is 1s.
  • the image correction device performs correction processing on the second dark field image according to the first dark field image and the bright field image, and the formula corresponding to the target image corresponding to the self-luminous object is obtained as follows:
  • I represents the pixel data corresponding to the target image
  • I 0 represents the pixel data corresponding to the second dark field image
  • I d represents the pixel data corresponding to the first dark field image
  • I f1 represents the light provided by the white light lamp in the imaging device.
  • the second dark field image, the first dark field image, and the bright field image have the same size.
  • the difference of I 0 -I d and I f1 -I d are for the same pixel in the two images, and the ratio I is each pixel.
  • the abscissa represents the lanes (1-5), and the ordinate represents the signal intensity percentage (unit: %) of each lane, which decreases from left to right.
  • the darker the displayed color (that is, the clearer the display), as shown in Figure 12, from left to right (lanes 1-5) is the location of the self-luminous object, that is, the target protein.
  • the corresponding signal intensity gradually decreases, and the displayed color It gradually becomes shallower.
  • area B represents the area where the markers of the pre-stained molecular weight standards are located.
  • a more accurate target image of the target protein intensity (ie, self-luminous object) on the biological sample membrane can be obtained.
  • the housing in the image acquisition device constitutes a dark room space.
  • the photoelectric conversion element first obtains the first dark field image.
  • the photoelectric conversion element obtains the first dark field image.
  • the second dark field image is then corrected according to the first dark field image to obtain the target image corresponding to the self-luminous object.
  • the bright field image under the light field generated by the light source device in the housing can be combined,
  • the target image of the self-luminous object with higher definition can be obtained more effectively and accurately, the operation process is simple, the imaging time is shorter, and the imaging device has the advantages of small structure, low manufacturing cost, convenient operation and convenient carrying.
  • the imaging device of the self-luminous object on the biological sample film of this embodiment is a further improvement of embodiment 1, specifically:
  • This embodiment includes the light source device 6 in Embodiment 2.
  • the photoelectric conversion element 2 is also used to obtain the corresponding bright field image within the set collection time when the biological sample film is not put into the housing and the external light source provides a uniform light field, that is, the first dark field image and the second dark field image.
  • the data collection time corresponding to the image and the bright field image are the same;
  • the image correction device 3 is also used to correct the second dark field image according to the first dark field image and the bright field image, and obtain the target image corresponding to the self-luminous object, as shown in FIG. 10, which is similar to the first dark field image.
  • the difference is: its overall brightness is higher than that of the first dark field image
  • an external light source (top illuminator S) is set directly above the imaging device and is about 2 meters away from the imaging device;
  • the image correction device performs correction processing on the second dark field image according to the first dark field image and the bright field image, and the formula corresponding to the target image corresponding to the self-luminous object is obtained as follows:
  • I represents the pixel data corresponding to the target image
  • I 0 represents the pixel data corresponding to the second dark field image
  • I d represents the pixel data corresponding to the first dark field image
  • I f2 represents the acquisition in a uniform light field provided by an external light source Pixel data corresponding to the bright field image.
  • the second dark field image, the first dark field image, and the bright field image have the same size.
  • the difference of I 0 -I d and I f2 -I d are for the same pixel in the two images, and the ratio I is each pixel.
  • the abscissa represents the lanes (1-5), and the ordinate represents the signal intensity percentage (unit: %) of each lane, which decreases from left to right.
  • the darker the displayed color (that is, the clearer the display), as shown in Figure 12, from left to right (lanes 1-5) is the location of the self-luminous object, that is, the target protein.
  • the corresponding signal intensity gradually decreases, and the displayed color It also gradually becomes lighter.
  • area B represents the area where the markers of the pre-stained molecular weight standards are located.
  • a more accurate target image of the target protein intensity (ie, self-luminous object) on the biological sample membrane can be obtained.
  • the housing in the image acquisition device constitutes a dark room space.
  • the photoelectric conversion element first obtains the first dark field image.
  • the photoelectric conversion element obtains the first dark field image.
  • the second dark field image is then corrected according to the first dark field image to obtain the target image corresponding to the self-luminous object.
  • it can be combined with the bright field image under the light field generated by the external light source, which is more effective , Obtain the target image of the self-luminous object with higher definition more accurately, the operation process is simple, the imaging time is shorter, and the imaging device has the advantages of small structure, low manufacturing cost, convenient operation and convenient carrying.
  • the imaging method of the self-luminous object on the biological sample film of this embodiment is implemented by the imaging device of the self-luminous object on the biological sample film of Embodiment 1.
  • the imaging method includes:
  • the photoelectric conversion element After the biological sample film is attached to the surface of the photoelectric conversion element, the photoelectric conversion element is used to obtain a second dark field image inside the inner housing for a set collection time, that is, the corresponding first dark field image and the second dark field image
  • the data collection time is the same;
  • a self-luminous object is carried on the biological sample film
  • S103 Perform correction processing on the second dark field image according to the first dark field image, and obtain a target image corresponding to the self-luminous object.
  • step S103 The formula corresponding to step S103 is as follows:
  • I represents the pixel data corresponding to the target image
  • I 0 represents the pixel data corresponding to the second dark field image
  • I d represents the pixel data corresponding to the first dark field image
  • the size of the second dark field image is the same as that of the first dark field image, and the difference of I 0 -I d represents the difference of the pixel values of the same point in the two images.
  • the housing in the image acquisition device constitutes a dark room space.
  • the photoelectric conversion element first obtains the first dark field image.
  • the photoelectric conversion element obtains the first dark field image.
  • the second dark field image is then corrected according to the first dark field image to obtain the target image corresponding to the self-luminous object, so as to obtain the target image of the self-luminous object with higher definition.
  • the operation process is simple and The imaging time is relatively short, and the imaging device has the advantages of small structure, low manufacturing cost, convenient operation and convenient portability.
  • the imaging method of the self-luminous object on the biological sample film of this embodiment is implemented by the imaging device of the self-luminous object on the biological sample film of Embodiment 2.
  • the imaging method of the self-luminous object on the biological sample film of this embodiment is a further improvement of Embodiment 4. Specifically:
  • step S102 and before step S103 the method further includes:
  • the biological sample film is not put into the housing and the light source device in the dark room space is turned on by using a photoelectric conversion element, the bright field image inside the housing within the set collection time is acquired, that is, the first dark field image and the second dark field image.
  • the data collection time corresponding to the field image and the bright field image are the same;
  • Step S103 includes:
  • step S1031 Perform correction processing on the second dark field image according to the first dark field image and the bright field image, and obtain a target image corresponding to the self-luminous object.
  • the formula corresponding to step S1031 is as follows:
  • I represents the pixel data corresponding to the target image
  • I 0 represents the pixel data corresponding to the second dark field image
  • I d represents the pixel data corresponding to the first dark field image
  • I f represents the pixel data corresponding to the bright field image.
  • the second dark field image, the first dark field image, and the bright field image have the same size.
  • the difference of I 0 -I d and I f1 -I d are for the same pixel in the two images, and the ratio I is each pixel.
  • the abscissa represents the lanes (1-5), and the ordinate represents the signal intensity percentage (unit: %) of each lane, which decreases from left to right.
  • the darker the displayed color (that is, the clearer the display), as shown in Figure 12, from left to right (lanes 1-5) is the location of the self-luminous object, that is, the target protein.
  • the corresponding signal intensity gradually decreases, and the displayed color It gradually becomes shallower.
  • area B represents the area where the markers of the pre-stained molecular weight standards are located.
  • a more accurate target image of the target protein intensity (ie, self-luminous object) on the biological sample membrane can be obtained.
  • the housing in the image acquisition device constitutes a dark room space.
  • the photoelectric conversion element first obtains the first dark field image.
  • the photoelectric conversion element obtains the first dark field image.
  • the second dark field image is then corrected according to the first dark field image to obtain the target image corresponding to the self-luminous object.
  • the bright field image under the light field generated by the light source device in the housing can be combined,
  • the target image of the self-luminous object with higher definition can be obtained more effectively and accurately, the operation process is simple, the imaging time is shorter, and the imaging device has the advantages of small structure, low manufacturing cost, convenient operation and convenient carrying.
  • the imaging method of the self-luminous object on the biological sample film of this embodiment is implemented by the imaging device of the self-luminous object on the biological sample film of Embodiment 3, and the self-luminous object on the biological sample film of this embodiment
  • the imaging method of the object is a further improvement of Embodiment 4. Specifically:
  • step S102 and before step S103 the method further includes:
  • Step S103 includes:
  • step S1032 The formula corresponding to step S1032 is as follows:
  • I represents the pixel data corresponding to the target image
  • I 0 represents the pixel data corresponding to the second dark field image
  • I d represents the pixel data corresponding to the first dark field image
  • I f represents the pixel data corresponding to the bright field image.
  • the second dark field image, the first dark field image, and the bright field image have the same size.
  • the difference of I 0 -I d and I f2 -I d are for the same pixel in the two images, and the ratio I is each pixel.
  • the abscissa represents the lanes (1-5), and the ordinate represents the signal intensity percentage (unit: %) of each lane, which decreases from left to right.
  • the darker the displayed color (that is, the clearer the display), as shown in Figure 12, from left to right (lanes 1-5) is the location of the self-luminous object, that is, the target protein.
  • the corresponding signal intensity gradually decreases, and the displayed color It gradually becomes shallower.
  • area B represents the area where the markers of the pre-stained molecular weight standards are located.
  • a more accurate target image of the target protein intensity (ie, self-luminous object) on the biological sample membrane can be obtained.
  • the housing in the image acquisition device constitutes a dark room space.
  • the photoelectric conversion element first obtains the first dark field image.
  • the photoelectric conversion element obtains the first dark field image.
  • the second dark field image is then corrected according to the first dark field image to obtain the target image corresponding to the self-luminous object.
  • it can be combined with the bright field image under the light field generated by the external light source, which is more effective , Obtain the target image of the self-luminous object with higher definition more accurately, the operation process is simple, the imaging time is shorter, and the imaging device has the advantages of small structure, low manufacturing cost, convenient operation and convenient carrying.

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Abstract

一种生物样品膜(A)上的自发光物体的成像装置及成像方法，该成像装置包括壳体(1)、光电转换元件(2)和图像校正设备(3)；壳体(1)内部构成暗室空间；光电转换元件(2)设于壳体(1)内；光电转换元件(2)用于在未放入生物样品膜(A)时，获取壳体(1)内部的第一暗场图像；在将生物样品膜(A)贴附在光电转换元件(2)表面后，光电转换元件(2)还用于获取壳体(1)内部的第二暗场图像；图像校正设备(3)用于根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像。该成像装置及成像方法能够有效地、更准确地获取清晰度较高的自发光物体的目标图像，操作过程简单、成像耗时较短，且该成像装置具有结构小、制造成本低、操作便捷且方便携带的优点。

Description

生物样品膜上的自发光物体的成像方法及装置
本申请要求申请日为2019/03/28的中国专利申请201910244792X的优先权。本申请引用上述中国专利申请的全文。
技术领域
本发明涉及样本分析技术领域，特别涉及一种生物样品膜上的自发光物体的成像方法及装置。
背景技术
目前常用的生物自发光检测技术有两类，一类是使用感光胶片技术，一类是使用相机拍摄技术。
其中，感光胶片技术类似于传统的照相洗胶片的方法，在暗室中，将自发光物体(附着在生物样品膜上)紧密贴合感光胶片，然后再把感光后的胶片使用显影液重洗，从而获得自发光物体对应的图像；但是，该成像方法存在操作过程复杂，且需要更多的步骤进行数字化处理；相机拍摄技术类似于手机的拍摄技术，将相机和自发光物体置于暗室内，该方法能够直接地进行数字化图像存储，但是，由于相机与自发光物体的距离较远，易造成光信号的采集率较低，因此需要较长的时间进行数据采集，且存在灵敏度不高等缺陷。另外，上述两种成像系统都存在体积大、占用空间大、成本高、不易搬运等缺陷。
发明内容
本发明要解决的技术问题是现有技术中获取生物样品膜上自发光物体的成像方式存在操作过程复杂、成像耗时较久，且成像系统存在体积大、占用空间大、成本高、不易搬运等缺陷，目的在于提供一种生物样品膜上的自发光物体的成像方法及装置。
本发明是通过下述技术方案来解决上述技术问题：
本发明提供一种生物样品膜上的自发光物体的成像装置，所述成像装置包括壳体、光电转换元件和图像校正设备；
所述壳体内部构成暗室空间；
所述光电转换元件设于所述壳体内；
所述光电转换元件用于在未放入生物样品膜时，获取设定采集时长内所述壳体内部的第一暗场图像；
其中，所述生物样品膜上承载有自发光物体；
在将所述生物样品膜贴附在所述光电转换元件表面后，所述光电转换元件还用于获取所述设定采集时长内所述壳体内部的第二暗场图像；
所述图像校正设备用于根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
较佳地，所述图像校正设备根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像对应的公式如下：
I＝I 0-I d
其中，I表示所述目标图像对应的像素数据，I 0表示所述第二暗场图像对应的像素数据，I d表示所述第一暗场图像对应的像素数据。
较佳地，所述成像装置还包括光源设备，所述光源设备设于所述壳体内。
较佳地，所述光电转换元件还用于在未将所述生物样品膜放入所述壳体且开启所述暗室空间内的所述光源设备后，获取所述设定采集时长内所述壳体内部的亮场图像；
所述图像校正设备还用于根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
较佳地，所述光电转换元件还用于在未将所述生物样品膜放入所述壳体、所述壳体处于打开状态且由外部光源提供均匀光场时，获取所述设定采集时长内对应的亮场图像；
所述图像校正设备还用于根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
较佳地，所述图像校正设备根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像对应的公式如下：
Figure PCTCN2020081617-appb-000001
其中，I表示所述目标图像对应的像素数据，I 0表示所述第二暗场图像对应的像素数据，I d表示所述第一暗场图像对应的像素数据，I f表示所述亮场图像对应的像素数据。
较佳地，所述壳体包括遮光盖和底座；
所述遮光盖的一侧和所述底座的一侧铰接；
当所述遮光盖与所述底座闭合时，所述壳体内部构成所述暗室空间。
较佳地，所述光源设备设于所述遮光盖内侧的顶部位置；和/或，
所述光源设备开启后对应的光照时长为10ms-30s；和/或，
所述光源设备包括点阵状布置的若干LED(发光二极管)灯珠、若干个由光纤导入 的灯、若干根平行排列的灯管或若干个板块状的灯。
较佳地，所述成像装置还包括散光板；
所述散光板固设于所述遮光盖内侧且位于所述光源设备正下方；和/或，
所述光电转换元件包括CMOS(互补金属氧化物半导体)芯片、CCD(电荷耦合器件)芯片或非晶硅光电转换探测器；和/或，
所述生物样品膜包括蛋白膜、琼脂糖胶块、琼脂糖胶条、聚丙烯酰氨凝胶胶块或聚丙烯酰氨凝胶胶条。
较佳地，所述成像装置还包括保护膜；
所述保护膜的两侧分别与所述生物样品膜和所述光电转换元件贴合。
较佳地，所述保护膜的厚度为0.01mm-0.2mm；和/或，
所述保护膜的材质为钢化玻璃膜或硬质塑料膜。
本发明还提供一种生物样品膜上的自发光物体的成像方法，所述成像方法利用上述的生物样品膜上的自发光物体的成像装置实现，所述成像方法包括：
采用所述光电转换元件获取设定采集时长内所述壳体内部的第一暗场图像；
在将生物样品膜贴附在所述光电转换元件表面后，采用所述光电转换元件获取所述设定采集时长内所述壳体内部的第二暗场图像；
其中，所述生物样品膜上承载有自发光物体；
根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
较佳地，所述根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤对应的公式如下：
I＝I 0-I d
其中，I表示所述目标图像对应的像素数据，I 0表示所述第二暗场图像对应的像素数据，I d表示所述第一暗场图像对应的像素数据。
较佳地，所述成像装置还包括光源设备；
所述根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤之前还包括：
采用所述光电转换元件在未将所述生物样品膜放入所述壳体且开启所述暗室空间内的所述光源设备后，获取所述设定采集时长内所述壳体内部的亮场图像；
所述根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤包括：
根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
较佳地，所述根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤之前还包括：
采用所述光电转换元件在未将所述生物样品膜放入所述壳体、所述壳体处于打开状态且由外部光源提供均匀光场时，获取所述设定采集时长内对应的亮场图像；
所述根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤包括：
根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
较佳地，所述根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤对应的公式如下：
Figure PCTCN2020081617-appb-000002
其中，I表示所述目标图像对应的像素数据，I 0表示所述第二暗场图像对应的像素数据，I d表示所述第一暗场图像对应的像素数据，I f表示所述亮场图像对应的像素数据。
本发明的积极进步效果在于：
本发明中的图像采集装置中的壳体内部构成暗室空间，在没有放入自发光物体时，光电转换元件先获取第一暗场图像，在放入自发光物体后，光电转换元件再获取第二暗场图像，然后根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像；另外可以结合壳体内的光源设备产生的光场或者外部光源产生的光场下的亮场图像，更有效地、更准确地获取清晰度较高的自发光物体的目标图像，操作过程简单、成像耗时较短，且该成像装置具有结构小、制造成本低、操作便捷且方便携带等优点。
附图说明
图1为本发明实施例1的生物样品膜上的自发光物体的成像装置的第一结构示意图。
图2为本发明实施例1的生物样品膜上的自发光物体的成像装置的第二结构示意图。
图3为本发明实施例1的生物样品膜上的自发光物体的成像装置中第一壳体结构示意图。
图4为本发明实施例1的生物样品膜上的自发光物体的成像装置中第二壳体结构示意图。
图5为本发明实施例1的生物样品膜上的自发光物体的成像装置中第一暗场成像示意图。
图6为本发明实施例1的生物样品膜上的自发光物体的成像装置中第二暗场成像示意图。
图7为本发明实施例2的生物样品膜上的自发光物体的成像装置的第一结构示意图。
图8为本发明实施例2的生物样品膜上的自发光物体的成像装置的第二结构示意图。
图9为本发明实施例2的生物样品膜上的自发光物体的成像装置的第三结构示意图。
图10为本发明实施例2的生物样品膜上的自发光物体的成像装置中亮场成像示意图。
图11为本发明实施例2的生物样品膜上的自发光物体的成像装置中泳道与信号强度的关系示意图。
图12为本发明实施例2的生物样品膜上的自发光物体的成像装置中第二暗场成像示意图。
图13为本发明实施例3的生物样品膜上的自发光物体的成像装置的结构示意图。
图14为本发明实施例4的生物样品膜上的自发光物体的成像方法的流程图。
图15为本发明实施例5的生物样品膜上的自发光物体的成像方法的流程图。
图16为本发明实施例6的生物样品膜上的自发光物体的成像方法的流程图。
具体实施方式
下面通过实施例的方式进一步说明本发明，但并不因此将本发明限制在所述的实施例范围之中。
实施例1
如图1和图2所示，本实施例的生物样品膜上的自发光物体的成像装置包括壳体1、光电转换元件2和图像校正设备3。
壳体1内部构成暗室空间，其中，壳体1包括遮光盖4和底座5，如图3所示，遮光盖4的一侧和底座5的一侧铰接，此时遮光盖4处于打开状态。
如图4所示，当遮光盖4与底座5闭合时，壳体1内部构成暗室空间，以保证对生物样品膜(在图2中用A表示)上自放光物体对应的微弱信号进行有效采集。
其中，生物样品膜包括蛋白膜、琼脂糖胶块、琼脂糖胶条、聚丙烯酰氨凝胶胶块、聚丙烯酰氨凝胶胶条等。
光电转换元件设于壳体内。其中，光电转换元件包括CMOS芯片、CCD芯片或非晶硅光电转换探测器等。
光电转换元件用于在未放入生物样品膜A时，获取设定采集时长内壳体内部的第一暗场图像(如图5所示)；
其中，生物样品膜上承载有自发光物体。
设定采集时长可以根据实际情况设置与调整，优选地，该设定采集时长为1s。
具体地，当生物样品膜包括蛋白膜时，将蛋白膜样品经过SDS-PAGE(十二烷基磺酸钠-聚丙烯酰氨凝胶)电泳技术进行分离后，在缓冲液中将蛋白转移到PVDF(聚偏氟乙烯)膜上获得承载有蛋白的生物样品膜，该生物样品膜依次完成目标蛋白特异性第一级抗体孵育、辣根过氧化物酶耦联的第二级抗体孵育，然后再用化学发光液进行显色(即显示为自发光物体)，在目标蛋白的区域产生化学光信号。
在将生物样品膜贴附在光电转换元件表面后，光电转换元件还用于获取设定采集时长内壳体内部的第二暗场图像(如图6所示)，即第一暗场图像和第二暗场图像对应的数据采集时长一致。
图像校正设备用于根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像。
采用光电转换元件获取第一暗场图像的过程包括：
打开遮光盖4，清除该成像装置中载物台(即光电转换元件)上残留的污渍等；
关闭遮光盖4，使得壳体1内部构成暗室空间，确保光电转换元件不受外光信号的影响；
等待设定时间(如1s)，采用光电转换元件采集壳体内部的均匀暗场中每个像素点的数值I d，其中采集时长为1s。
采用光电转换元件获取第二暗场图像的过程包括：
打开遮光盖4，将承载有自发光物体的生物样品膜放置在载物台(即光电转换元件)上，使得生物样品膜与光电转换元件紧密贴合；
关闭遮光盖4，使得壳体1内部构成暗室空间，确保光电转换元件不受外光信号的影响；
采用光电转换元件对生物样品膜上的自放光物体(目标蛋白)对应的光信号进行采集(此时目标蛋白之外的其他区域均属于均匀暗场)，光电转换元件采集壳体内部的每个像素点的数值I 0，其中采集时长为1s。
其中，图像校正设备根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像对应的公式如下：
I＝I 0-I d
其中，I表示目标图像对应的像素数据，I 0表示第二暗场图像对应的像素数据，I d表示第一暗场图像对应的像素数据。
第二暗场图像与第一暗场图像的尺寸大小一致，I 0-I d的差值表示两张图像中同一点的像素值的差值。
本实施例中，图像采集装置中的壳体内部构成暗室空间，在没有放入自发光物体时，光电转换元件先获取第一暗场图像，在放入自发光物体后，光电转换元件再获取第二暗场图像，然后根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像，从而通过接触式成像的方式获取清晰度较高的自发光物体的目标图像，操作过程简单、成像耗时较短，且该成像装置具有结构小、制造成本低、操作便捷且方便携带等优点。
实施例2
如图7所示，本实施例的生物样品膜上的自发光物体的成像装置是对实施例1的进一步改进，具体地：
成像装置还包括光源设备6，光源设备6设于壳体1内。
光源设备6设于遮光盖4内侧的顶部位置；
光源设备6开启后对应的光照时长为10ms-30s，该光照时长也可以根据实际需求进行调整。
光源设备6包括点阵状布置的若干LED灯珠、若干个由光纤导入的灯、若干根平行排列的灯管、若干个板块状的灯等。
成像装置还包括散光板7，固设于遮光盖内侧且位于光源设备正下方，使得光源设备6的光照均匀。
如图8所示，成像装置还包括保护膜8，保护膜8的两侧分别与生物样品膜和光电转换元件贴合。
其中，保护膜的厚度为0.01mm-0.2mm，保护膜的材质为钢化玻璃膜或硬质塑料膜。
保护膜8的外边缘位于光电转换元件2的外边缘的外部。
光电转换元件2包括探测器主体9和基板10，基板10的外边缘位于探测器主体9的外边缘的外部。
其中，基板为金属板、PCBA(一种线路板制作流程)板等。
如图9所示，底座5的顶部具有容置槽11，光电转换元件2和保护膜8位于容置槽11内。
本实施例的成像装置还包括封装件12，封装件作用于保护膜8和底座5，以限制光电转换元件2相对于底座5的运动。
具体地，封装件12包括填充物13和卡板14。
填充物13位于光电转换元件2、保护膜8与底座5的表面所围成的区域内，用于固定光电转换元件2在底座5中。
其中，填充物为凝胶，玻璃粉、塑料胚等。
当光电转换元件2处于封装状态时，卡板14抵接于保护膜8的四周边缘，用以保证整个装置的稳定性。
底座5包括底板15和底座主体16，底座主体16上设有容置槽11，且容置槽11贯穿于底座主体16，底板15活动连接于底座主体16的底部，以自底座主体16的底部打开或关闭容置槽11。光电转换元件2的高度不大于容置槽11的深度。
光电转换元件2还用于在未将生物样品膜放入壳体1且开启暗室空间内的光源设备6后，获取设定采集时长内壳体内部的亮场图像，即第一暗场图像、第二暗场图像和亮场图像对应的数据采集时长均一致。
如图10所示，该图与图5中第一暗场图像的区别在于：其整体亮度高于第一暗场图像的整体亮度，其中，斜线数量越少，表示对应的亮度越高。
图像校正设备3还用于根据第一暗场图像和亮场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像。
采用光电转换元件获取亮场图像的过程包括：
打开遮光盖4，清除该成像装置中载物台(即光电转换元件)上残留的污渍等；
关闭遮光盖4，使得壳体1内部构成暗室空间，确保光电转换元件不受外光信号的影响；
控制光源设备开启(对应的光照时长为10ms-30s)，并采用光电转换元件2同步采集该均匀亮场中每个像素点的数值I f1，其中采集时长为1s。
则图像校正设备根据第一暗场图像和亮场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像对应的公式如下：
Figure PCTCN2020081617-appb-000003
其中，I表示目标图像对应的像素数据，I 0表示第二暗场图像对应的像素数据，I d表示第一暗场图像对应的像素数据，I f1表示在成像装置内的白光灯提供的光场中获取的亮场图像对应的像素数据。
第二暗场图像、第一暗场图像以及亮场图像尺寸大小一致，I 0-I d的差值以及I f1-I d均为针对两张图像中同一像素点，比值I为每个像素点对应的信号强度百分比。
具体地，如图11所示，横坐标表示泳道(1-5)，纵坐标表示每个泳道的信号强度百分比(单位：％)，从左向右依次减弱。选取生物样品膜上的每个泳道，并计算每个泳道的信号值，基于这些信号值比较每个泳道目标蛋白的信号强度，其中信号值越大，对应泳道的目标蛋白的信号强度越强，显示的颜色越深(即显示越清晰)，如图12所示，从左到右(泳道1-5)为自发光物体即目标蛋白所在位置处，其对应的信号强度逐渐递减，显示的颜色也依次逐渐变浅。另外，B区域表示预染的分子量标样的标志物所在区域。
因此，根据数据处理得到的信号值，可以获得更精确的生物样品膜上目标蛋白强度(即自发光物体)的目标图像。
本实施例中，图像采集装置中的壳体内部构成暗室空间，在没有放入自发光物体时，光电转换元件先获取第一暗场图像，在放入自发光物体后，光电转换元件再获取第二暗场图像，然后根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像，另外可以结合壳体内的光源设备产生的光场下的亮场图像，更有效地、更准确地获取清晰度较高的自发光物体的目标图像，操作过程简单、成像耗时较短，且该成像装置具有结构小、制造成本低、操作便捷且方便携带等优点。
实施例3
本实施例的生物样品膜上的自发光物体的成像装置是对实施例1的进一步改进，具体地：
本实施例包括实施例2中的光源设备6。
光电转换元件2还用于在未将生物样品膜放入壳体且由外部光源提供均匀光场时，获取设定采集时长内对应的亮场图像，即第一暗场图像、第二暗场图像和亮场图像对应的数据采集时长均一致；
图像校正设备3还用于根据第一暗场图像和亮场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像，如图10所示，该图与第一暗场图像的区别在于：其整体亮度高于第一暗场图像的整体亮度
采用光电转换元件获取亮场图像的过程包括：
打开遮光盖4，清除该成像装置中载物台(即光电转换元件)上残留的污渍等；
如图13所示，在开放的空间里，在成像装置正上方设置一外部光源(顶部照明灯S)，且距离成像装置约2米处；
等待1分钟，然后采用光电转换元件采集该均匀亮场中每个像素点的数值I f2。
则图像校正设备根据第一暗场图像和亮场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像对应的公式如下：
Figure PCTCN2020081617-appb-000004
其中，I表示目标图像对应的像素数据，I 0表示第二暗场图像对应的像素数据，I d表示第一暗场图像对应的像素数据，I f2表示在由外部光源提供均匀光场中获取的亮场图像对应的像素数据。
第二暗场图像、第一暗场图像以及亮场图像尺寸大小一致，I 0-I d的差值以及I f2-I d均为针对两张图像中同一像素点，比值I为每个像素点对应的信号强度百分比。
具体地，如图11所示，横坐标表示泳道(1-5)，纵坐标表示每个泳道的信号强度百分比(单位：％)，从左向右依次减弱。选取生物样品膜上的每个泳道，并计算每个泳道的信号值，基于这些信号值比较每个泳道目标蛋白的信号强度，其中信号值越大，对应泳道的目标蛋白的信号强度越强，显示的颜色越深(即显示越清晰)，如图12所示，从左到右(泳道1-5)为自发光物体即目标蛋白所在位置处，其对应的信号强度逐渐递减，显示的颜色也依次逐渐变浅，另外，B区域表示预染的分子量标样的标志物所在区域。
因此，根据数据处理得到的信号值，可以获得更精确的生物样品膜上目标蛋白强度(即自发光物体)的目标图像。
本实施例中，图像采集装置中的壳体内部构成暗室空间，在没有放入自发光物体时，光电转换元件先获取第一暗场图像，在放入自发光物体后，光电转换元件再获取第二暗场图像，然后根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像，另外可以结合外部光源产生的光场下的亮场图像，更有效地、更准确地获取清晰度较高的自发光物体的目标图像，操作过程简单、成像耗时较短，且该成像装置具有结构小、制造成本低、操作便捷且方便携带等优点。
实施例4
如图14所示，本实施例的生物样品膜上的自发光物体的成像方法实施例1的生物样品膜上的自发光物体的成像装置实现，该成像方法包括：
S101、采用光电转换元件获取设定采集时长内壳体内部的第一暗场图像；
设定采集时长可以根据实际情况设置与调整，优选地，该设定采集时长为1s。
S102、在将生物样品膜贴附在光电转换元件表面后，采用光电转换元件获取设定采集时长内壳体内部的第二暗场图像，即第一暗场图像和第二暗场图像对应的数据采集时长一致；
其中，生物样品膜上承载有自发光物体；
S103、根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像。
步骤S103对应的公式如下：
I＝I 0-I d
其中，I表示目标图像对应的像素数据，I 0表示第二暗场图像对应的像素数据，I d表示第一暗场图像对应的像素数据。
第二暗场图像与第一暗场图像的尺寸大小一致，I 0-I d的差值表示两张图像中同一点的像素值的差值。
本实施例中，图像采集装置中的壳体内部构成暗室空间，在没有放入自发光物体时，光电转换元件先获取第一暗场图像，在放入自发光物体后，光电转换元件再获取第二暗场图像，然后根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像，从而获取清晰度较高的自发光物体的目标图像，操作过程简单、成像耗时较短，且该成像装置具有结构小、制造成本低、操作便捷且方便携带等优点。
实施例5
如图15所示，本实施例的生物样品膜上的自发光物体的成像方法实施例2的生物样品膜上的自发光物体的成像装置实现，
本实施例的生物样品膜上的自发光物体的成像方法是对实施例4的进一步改进，具体地：
步骤S102之后、步骤S103之前还包括：
S10301、采用光电转换元件在未将生物样品膜放入壳体且开启暗室空间内的光源设备后，获取设定采集时长内壳体内部的亮场图像，即第一暗场图像、第二暗场图像和亮场图像对应的数据采集时长均一致；
步骤S103包括：
S1031、根据第一暗场图像和亮场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像。步骤S1031对应的公式如下：
Figure PCTCN2020081617-appb-000005
其中，I表示目标图像对应的像素数据，I 0表示第二暗场图像对应的像素数据，I d表示第一暗场图像对应的像素数据，I f表示亮场图像对应的像素数据。
第二暗场图像、第一暗场图像以及亮场图像尺寸大小一致，I 0-I d的差值以及I f1-I d均为针对两张图像中同一像素点，比值I为每个像素点对应的信号强度百分比。
具体地，如图11所示，横坐标表示泳道(1-5)，纵坐标表示每个泳道的信号强度百分比(单位：％)，从左向右依次减弱。选取生物样品膜上的每个泳道，并计算每个泳道的信号值，基于这些信号值比较每个泳道目标蛋白的信号强度，其中信号值越大，对应泳道的目标蛋白的信号强度越强，显示的颜色越深(即显示越清晰)，如图12所示，从左到右(泳道1-5)为自发光物体即目标蛋白所在位置处，其对应的信号强度逐渐递减，显示的颜色也依次逐渐变浅。另外，B区域表示预染的分子量标样的标志物所在区域。
因此，根据数据处理得到的信号值，可以获得更精确的生物样品膜上目标蛋白强度(即自发光物体)的目标图像。
本实施例中，图像采集装置中的壳体内部构成暗室空间，在没有放入自发光物体时，光电转换元件先获取第一暗场图像，在放入自发光物体后，光电转换元件再获取第二暗场图像，然后根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像，另外可以结合壳体内的光源设备产生的光场下的亮场图像，更有效地、更准确地获取清晰度较高的自发光物体的目标图像，操作过程简单、成像耗时较短，且该成像装置具有结构小、制造成本低、操作便捷且方便携带等优点。
实施例6
如图16所示，本实施例的生物样品膜上的自发光物体的成像方法实施例3的生物样品膜上的自发光物体的成像装置实现，且本实施例的生物样品膜上的自发光物体的成像方法是对实施例4的进一步改进，具体地：
步骤S102之后、步骤S103之前还包括：
S10302、采用光电转换元件在未将生物样品膜放入壳体且由外部光源提供均匀光场时，获取设定采集时长内对应的亮场图像，即第一暗场图像、第二暗场图像和亮场图像对应的数据采集时长均一致；
步骤S103包括：
S1032、根据第一暗场图像和亮场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像。
步骤S1032对应的公式如下：
Figure PCTCN2020081617-appb-000006
其中，I表示目标图像对应的像素数据，I 0表示第二暗场图像对应的像素数据，I d表示第一暗场图像对应的像素数据，I f表示亮场图像对应的像素数据。
第二暗场图像、第一暗场图像以及亮场图像尺寸大小一致，I 0-I d的差值以及I f2-I d均为针对两张图像中同一像素点，比值I为每个像素点对应的信号强度百分比。
具体地，如图11所示，横坐标表示泳道(1-5)，纵坐标表示每个泳道的信号强度百分比(单位：％)，从左向右依次减弱。选取生物样品膜上的每个泳道，并计算每个泳道的信号值，基于这些信号值比较每个泳道目标蛋白的信号强度，其中信号值越大，对应泳道的目标蛋白的信号强度越强，显示的颜色越深(即显示越清晰)，如图12所示，从左到右(泳道1-5)为自发光物体即目标蛋白所在位置处，其对应的信号强度逐渐递减，显示的颜色也依次逐渐变浅。另外，B区域表示预染的分子量标样的标志物所在区域。
因此，根据数据处理得到的信号值，可以获得更精确的生物样品膜上目标蛋白强度(即自发光物体)的目标图像。
本实施例中，图像采集装置中的壳体内部构成暗室空间，在没有放入自发光物体时，光电转换元件先获取第一暗场图像，在放入自发光物体后，光电转换元件再获取第二暗场图像，然后根据第一暗场图像对第二暗场图像进行校正处理，获取自发光物体对应的目标图像，另外可以结合外部光源产生的光场下的亮场图像，更有效地、更准确地获取清晰度较高的自发光物体的目标图像，操作过程简单、成像耗时较短，且该成像装置具有结构小、制造成本低、操作便捷且方便携带等优点。
虽然以上描述了本发明的具体实施方式，但是本领域的技术人员应当理解，这些仅是举例说明，在不背离本发明的原理和实质的前提下，可以对这些实施方式做出多种变更或修改。因此，本发明的保护范围由所附权利要求书限定。

Claims (16)

  1. 一种生物样品膜上的自发光物体的成像装置，其特征在于，所述成像装置包括壳体、光电转换元件和图像校正设备；
    所述壳体内部构成暗室空间；
    所述光电转换元件设于所述壳体内；
    所述光电转换元件用于在未放入生物样品膜时，获取设定采集时长内所述壳体内部的第一暗场图像；
    其中，所述生物样品膜上承载有自发光物体；
    在将所述生物样品膜贴附在所述光电转换元件表面后，所述光电转换元件还用于获取所述设定采集时长内所述壳体内部的第二暗场图像；
    所述图像校正设备用于根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
  2. 如权利要求1所述的生物样品膜上的自发光物体的成像装置，其特征在于，所述图像校正设备根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像对应的公式如下：
    I＝I 0-I d
    其中，I表示所述目标图像对应的像素数据，I 0表示所述第二暗场图像对应的像素数据，I d表示所述第一暗场图像对应的像素数据。
  3. 权利要求1或2所述的生物样品膜上的自发光物体的成像装置，其特征在于，所述成像装置还包括光源设备，所述光源设备设于所述壳体内。
  4. 如权利要求3所述的生物样品膜上的自发光物体的成像装置，其特征在于，所述光电转换元件还用于在未将所述生物样品膜放入所述壳体且开启所述暗室空间内的所述光源设备后，获取所述设定采集时长内所述壳体内部的亮场图像；
    所述图像校正设备还用于根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
  5. 如权利要求3或4所述的生物样品膜上的自发光物体的成像装置，其特征在于，所述光电转换元件还用于在未将所述生物样品膜放入所述壳体、所述壳体处于打开状态且由外部光源提供均匀光场时，获取所述设定采集时长内对应的亮场图像；
    所述图像校正设备还用于根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
  6. 如权利要求4或5所述的生物样品膜上的自发光物体的成像装置，其特征在于，所述图像校正设备根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像对应的公式如下：
    Figure PCTCN2020081617-appb-100001
    其中，I表示所述目标图像对应的像素数据，I 0表示所述第二暗场图像对应的像素数据，I d表示所述第一暗场图像对应的像素数据，I f表示所述亮场图像对应的像素数据。
  7. 如权利要求3-6中至少一项所述的生物样品膜上的自发光物体的成像装置，其特征在于，所述壳体包括遮光盖和底座；
    所述遮光盖的一侧和所述底座的一侧铰接；
    当所述遮光盖与所述底座闭合时，所述壳体内部构成所述暗室空间。
  8. 如权利要求7所述的生物样品膜上的自发光物体的成像装置，其特征在于，所述光源设备设于所述遮光盖内侧的顶部位置；和/或，
    所述光源设备开启后对应的光照时长为10ms-30s；和/或，
    所述光源设备包括点阵状布置的若干LED灯珠、若干个由光纤导入的灯、若干根平行排列的灯管或若干个板块状的灯。
  9. 如权利要求7或8所述的生物样品膜上的自发光物体的成像装置，其特征在于，所述成像装置还包括散光板；
    所述散光板固设于所述遮光盖内侧且位于所述光源设备正下方；和/或，
    所述光电转换元件包括CMOS芯片、CCD芯片或非晶硅光电转换探测器；和/或，
    所述生物样品膜包括蛋白膜、琼脂糖胶块、琼脂糖胶条、聚丙烯酰氨凝胶胶块或聚丙烯酰氨凝胶胶条。
  10. 如权利要求1-9中至少一项所述的生物样品膜上的自发光物体的成像装置，其特征在于，所述成像装置还包括保护膜；
    所述保护膜的两侧分别与所述生物样品膜和所述光电转换元件贴合。
  11. 如权利要求10所述的生物样品膜上的自发光物体的成像装置，其特征在于，所述保护膜的厚度为0.01mm-0.2mm；和/或，
    所述保护膜的材质为钢化玻璃膜或硬质塑料膜。
  12. 一种生物样品膜上的自发光物体的成像方法，其特征在于，所述成像方法利用权利要求1-11中任意一项所述的生物样品膜上的自发光物体的成像装置实现，所述成像方法包括：
    采用所述光电转换元件获取设定采集时长内所述壳体内部的第一暗场图像；
    在将生物样品膜贴附在所述光电转换元件表面后，采用所述光电转换元件获取所述设定采集时长内所述壳体内部的第二暗场图像；
    其中，所述生物样品膜上承载有自发光物体；
    根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
  13. 如权利要求12所述的生物样品膜上的自发光物体的成像方法，其特征在于，所述根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤对应的公式如下：
    I＝I 0-I d
    其中，I表示所述目标图像对应的像素数据，I 0表示所述第二暗场图像对应的像素数据，I d表示所述第一暗场图像对应的像素数据。
  14. 如权利要求12或13所述的生物样品膜上的自发光物体的成像方法，其特征在于，所述成像装置还包括光源设备；
    所述根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤之前还包括：
    采用所述光电转换元件在未将所述生物样品膜放入所述壳体且开启所述暗室空间内的所述光源设备后，获取所述设定采集时长内所述壳体内部的亮场图像；
    所述根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤包括：
    根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
  15. 如权利要求12所述的生物样品膜上的自发光物体的成像方法，其特征在于，所述根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤之前还包括：
    采用所述光电转换元件在未将所述生物样品膜放入所述壳体、所述壳体处于打开状态且由外部光源提供均匀光场时，获取所述设定采集时长内对应的亮场图像；
    所述根据所述第一暗场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤包括：
    根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像。
  16. 如权利要求14或15所述的生物样品膜上的自发光物体的成像方法，其特征在于，所述根据所述第一暗场图像和所述亮场图像对所述第二暗场图像进行校正处理，获取所述自发光物体对应的目标图像的步骤对应的公式如下：
    Figure PCTCN2020081617-appb-100002
    其中，I表示所述目标图像对应的像素数据，I 0表示所述第二暗场图像对应的像素数据，I d表示所述第一暗场图像对应的像素数据，I f表示所述亮场图像对应的像素数据。
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