CN107907396B - Automatic immunohistochemical staining system for biological tissues - Google Patents
Automatic immunohistochemical staining system for biological tissues Download PDFInfo
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- CN107907396B CN107907396B CN201711087020.7A CN201711087020A CN107907396B CN 107907396 B CN107907396 B CN 107907396B CN 201711087020 A CN201711087020 A CN 201711087020A CN 107907396 B CN107907396 B CN 107907396B
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- 238000011532 immunohistochemical staining Methods 0.000 title claims abstract description 11
- 239000007788 liquid Substances 0.000 claims abstract description 196
- 238000004043 dyeing Methods 0.000 claims abstract description 55
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 claims abstract description 54
- 239000011521 glass Substances 0.000 claims abstract description 50
- 239000000243 solution Substances 0.000 claims abstract description 27
- 239000007977 PBT buffer Substances 0.000 claims abstract description 21
- 238000007789 sealing Methods 0.000 claims abstract description 18
- 239000002699 waste material Substances 0.000 claims abstract description 9
- 239000000523 sample Substances 0.000 claims description 8
- 238000010186 staining Methods 0.000 claims description 2
- 230000002265 prevention Effects 0.000 claims 1
- 238000002474 experimental method Methods 0.000 abstract description 8
- 230000002055 immunohistochemical effect Effects 0.000 abstract description 8
- 238000000034 method Methods 0.000 abstract description 5
- 238000003364 immunohistochemistry Methods 0.000 abstract description 3
- 239000000427 antigen Substances 0.000 description 3
- 102000036639 antigens Human genes 0.000 description 3
- 108091007433 antigens Proteins 0.000 description 3
- 239000007853 buffer solution Substances 0.000 description 3
- 241000255588 Tephritidae Species 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 230000007547 defect Effects 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 238000011534 incubation Methods 0.000 description 2
- 229920001184 polypeptide Polymers 0.000 description 2
- 102000004196 processed proteins & peptides Human genes 0.000 description 2
- 108090000765 processed proteins & peptides Proteins 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 210000005013 brain tissue Anatomy 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 229910021645 metal ion Inorganic materials 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000008055 phosphate buffer solution Substances 0.000 description 1
- 238000009991 scouring Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
- G01N1/31—Apparatus therefor
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Physics & Mathematics (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
- Sampling And Sample Adjustment (AREA)
Abstract
An automatic immunohistochemical staining system for biological tissues relates to biological tissue culture equipment. The invention solves the technical problems of manual operation, time and labor consumption and inapplicability to immunohistochemistry of thicker biological tissues in the conventional immunohistochemical experiment. The invention comprises a glass dyeing pipe, a primary antibody liquid storage tank, a secondary antibody liquid storage tank, a PBT buffer solution liquid storage tank, a formaldehyde fixing liquid storage tank, a liquid inlet pipe, a liquid outlet pipe, a plurality of connecting pipes and a waste liquid tank; the glass dyeing pipe is characterized in that one end of the glass dyeing pipe is opened, a sealing plug is arranged at the opening, a liquid inlet hole and a liquid outlet hole are formed in the sealing plug, a liquid outlet pipe protection cavity is formed in the inner bottom side of the glass dyeing pipe, and a plurality of through holes are formed in the side wall of the liquid outlet pipe protection cavity. The invention adopts a special glass dyeing tube, can put thicker biological tissues in a dyeing cavity for an immunohistochemical dyeing experiment, and automatically finishes an immunohistochemical process.
Description
Technical Field
The invention belongs to the technical field of biological tissue culture equipment, and particularly relates to an automatic immunohistochemical staining system for biological tissues.
Background
Immunohistochemistry is the research of determining the antigen polypeptide and protein in tissue cell by applying the basic principle of immunology, namely the antigen-antibody reaction, namely the principle of specific combination of antigen and antibody, and developing the color developing agent fluorescein, enzyme, metal ion and isotope of the labeled antibody through chemical reaction, and carrying out positioning, qualitative and quantitative research on the antigen polypeptide and protein.
The whole process of the immunohistochemical experiment is complex in steps, manual sample adding and liquid changing and washing are needed at regular intervals, time and labor are consumed, and the defects of inaccurate sample adding amount and the like easily occur.
An automatic immunohistochemical device for fruit fly wing buds, which adopts a special flat space of a glass slide, can be automatically operated, but can only be used for immunohistochemical staining experiments of thinner biological tissues, but is not suitable for thicker biological tissues, such as brain tissues of fruit flies, and can not be used for immunohistochemical staining experiments of the flat space of the glass slide.
Disclosure of Invention
The invention aims to overcome the defects of the prior art, provides an automatic immunohistochemical staining system for biological tissues, and solves the technical problems that the conventional immunohistochemical experiment is manually operated, is time-consuming and labor-consuming, and is not suitable for immunohistochemistry of thicker biological tissues.
The invention is realized by the following technical scheme:
an automated immunohistochemical staining system for biological tissue, wherein: comprises a glass dyeing pipe, a primary antibody liquid storage tank, a secondary antibody liquid storage tank, a PBT buffer solution liquid storage tank, a formaldehyde fixing liquid storage tank, a liquid inlet pipe, a liquid outlet pipe, a plurality of connecting pipes and a waste liquid tank;
one end of the glass dyeing pipe is opened, a sealing plug is arranged at the opening of the glass dyeing pipe, a liquid inlet hole and a liquid outlet hole are formed in the sealing plug, and a liquid outlet pipe protection cavity is arranged at the inner bottom side of the glass dyeing pipe;
the first anti-liquid storage tank, the second anti-liquid storage tank, the PBT buffer solution liquid storage tank and the formaldehyde fixing liquid storage tank are arranged above one side of the glass dyeing pipe, the waste liquid tank is arranged below one side of the glass dyeing pipe, outlets of the first anti-liquid storage tank, the second anti-liquid storage tank, the PBT buffer solution liquid storage tank and the formaldehyde fixing liquid storage tank are respectively connected with the liquid inlet pipe through connecting pipes, the liquid inlet pipe penetrates through a liquid inlet hole in the sealing plug and extends into the glass dyeing pipe, one end of the liquid outlet pipe penetrates through a liquid outlet hole in the sealing plug and extends into the liquid outlet pipe protection cavity, and the other end of the;
the connecting pipes among the primary anti-liquid storage tank, the secondary anti-liquid storage tank, the PBT buffer solution storage tank, the formaldehyde stationary liquid storage tank and the liquid inlet pipe are all provided with miniature liquid control valves controlled by the singlechip control module, and the liquid outlet pipe is provided with a liquid pump controlled by the singlechip control module;
the first anti-liquid storage tank and the second anti-liquid storage tank are provided with low-temperature control devices controlled by a single-chip microcomputer control module, and the glass dyeing pipe is provided with an infrared temperature measuring probe.
Further, the glass dyeing tube is fixed on a rocker table, and the rocker table is rocked according to a certain frequency; the liquid inlet pipe and the liquid outlet pipe are flexible pipes.
Further, the outlet end of the liquid inlet pipe is bent towards the inner wall of the glass dyeing pipe.
Furthermore, a plurality of liquid through holes for preventing biological tissues from passing through are arranged on the side wall of the liquid outlet pipe protection cavity, and a filter screen is arranged on the liquid outlet pipe in the liquid outlet pipe protection cavity.
The invention adopts a specially-made glass dyeing tube, can put thicker biological tissues into a dyeing cavity to carry out an immunohistochemical dyeing experiment, and automatically finishes an immunohistochemical process; the arrangement of the sealing plug can realize the sealing of the dyeing tube and avoid the infection of external microorganisms on the culture solution; the first anti-liquid storage tank, the second anti-liquid storage tank, the PBT buffer solution storage tank, the formaldehyde stationary liquid storage tank, the connecting pipe, the micro liquid control valve, the liquid inlet pipe, the liquid outlet pipe and the waste liquid tank are arranged, and the culture solution in the dyeing pipe can be replaced according to the requirement; particularly, a micro liquid control valve is arranged among formaldehyde fixing liquid, primary antibody, secondary antibody, PBT buffer solution, a liquid storage tank and a liquid inlet pipe, and a liquid pump is arranged on a liquid outlet pipe, so that automatic operation can be realized; the arrangement of the low-temperature control device on the primary antibody liquid storage tank and the secondary antibody liquid storage tank ensures that the antibodies are not degraded in the dyeing process. The glass dyeing tube is provided with a temperature control system to ensure that primary antibody incubation is carried out at low temperature and other steps are carried out at about room temperature.
Drawings
FIG. 1 is a schematic diagram of an automated immunohistochemical staining system for biological tissue;
FIG. 2 is a schematic view of the structure of a glass dyeing tube;
FIG. 3 is a right side view of FIG. 2;
fig. 4 is a schematic block diagram of a single chip microcomputer control circuit.
Detailed Description
The invention is described in further detail below with reference to the figures and examples.
As shown in fig. 1 to 3, the automatic immunohistochemical staining system for biological tissue in this embodiment includes a glass staining tube 1, a primary antibody liquid storage tank 10, a secondary antibody liquid storage tank 11, a PBT buffer solution liquid storage tank 12, a formaldehyde stationary liquid storage tank 13, a liquid inlet tube 4, a liquid outlet tube 5, a plurality of connecting tubes 3, and a waste liquid tank 6;
one end of the glass dyeing tube 1 is opened, a sealing plug 7 is arranged at the opening of the glass dyeing tube 1, and a liquid inlet hole 8 and a liquid outlet hole 9 are arranged on the sealing plug 7;
in order to prevent biological tissues from being sucked into the liquid outlet pipe when liquid is discharged, a liquid outlet pipe protection cavity 2 is arranged on the bottom side in the glass dyeing pipe 1;
the first anti-liquid storage tank 10, the second anti-liquid storage tank 11, the PBT buffer solution liquid storage tank 12 and the formaldehyde fixing liquid storage tank 13 are arranged above one side of the glass dyeing pipe 1, the waste liquid tank 6 is arranged below one side of the glass dyeing pipe 1, outlets of the first anti-liquid storage tank 10, the second anti-liquid storage tank 11, the PBT buffer solution liquid storage tank 12 and the formaldehyde fixing liquid storage tank 13 are respectively connected with the liquid inlet pipe 4 through the connecting pipe 3, the liquid inlet pipe 4 penetrates through the liquid inlet hole 8 on the sealing plug 7 and extends into the glass dyeing pipe 1, one end of the liquid outlet pipe 5 penetrates through the liquid outlet hole 9 on the sealing plug 7 and extends into the liquid outlet pipe protection cavity 2, and the other end of the liquid outlet pipe;
a micro liquid control valve 14 controlled by a singlechip control module is arranged on each connecting pipe 3 between the first anti-liquid storage tank 10, the second anti-liquid storage tank 11, the PBT buffer solution storage tank 12, the formaldehyde fixing solution storage tank 13 and the liquid inlet pipe 4, and a liquid pump 15 controlled by the singlechip control module is arranged on the liquid outlet pipe 5;
the first anti-liquid storage tank 10 and the second anti-liquid storage tank 11 are provided with low-temperature control devices controlled by a single-chip microcomputer control module, and the glass dyeing pipe 1 is provided with an infrared temperature measuring probe 17.
The glass dyeing tube 1 is fixed on a warped plate shaking table 16, and the warped plate shaking table shakes according to a certain frequency; the liquid inlet pipe 4 and the liquid outlet pipe 5 are flexible pipes, and the shaking of the shaking table is not influenced.
The outlet end of the liquid inlet pipe 4 is bent towards the inner wall of the glass dyeing pipe 1, so that the liquid is prevented from directly scouring biological tissues to damage the biological tissues when the liquid is fed.
The side wall of the liquid outlet pipe protection cavity 2 is provided with a plurality of liquid through small holes for preventing biological tissues from passing through, and the liquid outlet pipe in the liquid outlet pipe protection cavity 2 is provided with a filter screen to ensure that liquid is smoothly pumped out of the glass dyeing pipe.
As shown in fig. 4, the infrared temperature measuring probe 17 is connected with the single chip microcomputer control module to measure the temperature in the glass dyeing tube 1 in real time, the low temperature control devices of the primary antibody liquid storage tank 10 and the secondary antibody liquid storage tank 11 are connected with the single chip microcomputer control module, the data measured by the infrared temperature measuring probe are fed back to the single chip microcomputer, the single chip microcomputer controls the low temperature control devices of the primary antibody liquid storage tank 10 and the secondary antibody liquid storage tank 11, so that the primary antibody incubation is ensured to be carried out at 4-6 ℃, and other steps are carried out at about 25 ℃.
The use process of the invention is as follows:
when in use, the wing buds dissected in the phosphate buffer solution are transferred into the glass dyeing tube 1, the sealing plug 7 is plugged tightly, the glass dyeing tube 1 is fixed on the rocker table 16, and the rocker table is shaken at a certain frequency;
through singlechip control circuit:
firstly, opening a micro liquid control valve 14 and a liquid drawing pump 15 on a connecting pipe below a formaldehyde stationary liquid storage tank 13 to ensure that a certain amount of stationary liquid is filled in a glass dyeing pipe 1, then closing the micro liquid control valve 14 and the liquid drawing pump 15 on the connecting pipe below the formaldehyde stationary liquid storage tank 13, and fixing for 40-60 min;
then, the micro liquid control valve 14 and the liquid pump 15 on the connecting pipe below the PBT buffer solution storage tank 12 are opened, and after the buffer solution continuously cleans the formaldehyde stationary liquid for 1-1.5 h, the micro liquid control valve 14 and the liquid pump 15 on the connecting pipe below the PBT buffer solution storage tank 12 are closed;
and simultaneously, the micro liquid control valve 14 and the liquid pump 15 on the connecting pipe below the primary anti-liquid storage tank 10 are opened to ensure that the glass dyeing pipe 1 is filled with sufficient primary anti-liquid and the biological tissue is fully soaked, and then the micro liquid control valve 14 and the liquid pump 15 on the connecting pipe below the primary anti-liquid storage tank 10 are closed to incubate for 8-12 hours. The temperature of the glass dyeing tube is controlled by the single chip microcomputer to be 4-6 ℃.
The micro liquid control valve 14 and the liquid pump 15 on the connecting pipe below the PBT buffer solution storage tank 12 are started again, and after the buffer solution continuously cleans the primary antibody solution for 1-1.5 hours, the micro liquid control valve 14 and the liquid pump 15 on the connecting pipe below the PBT buffer solution storage tank 12 are closed;
simultaneously, the micro liquid control valve 14 on the connecting pipe below the secondary antibody liquid storage tank 11 and the liquid pump 15 on the liquid outlet pipe 5 are opened to ensure that a certain amount of secondary antibody solution is filled in the glass dyeing pipe 1 and fully soak the biological tissue, and then the micro liquid control valve 14 and the liquid pump 15 on the connecting pipe below the secondary antibody liquid storage tank 10 are closed to incubate for 1-1.2 hours;
and finally, starting the micro liquid control valve 14 on the connecting pipe below the PBT buffer solution storage tank 12 and the liquid pump 15 on the liquid outlet pipe 5, continuously cleaning the secondary antibody solution by the buffer solution for 1-1.5 h, and then closing the micro liquid control valve 14 and the liquid pump 15 on the connecting pipe below the PBT buffer solution storage tank 12 to complete the experiment.
After the sealing plug 7 is used for plugging the glass dyeing tube 1, a tube cavity of the glass dyeing tube 1, the liquid inlet tube 4 and the liquid outlet tube 5 form a closed space, so that the liquid in the tube cavity of the glass dyeing tube 1 can be pumped away by the liquid pump 15.
As the present invention may be embodied in several forms without departing from the spirit or essential characteristics thereof, it should also be understood that the above-described embodiments are not limited by any of the details of the foregoing description, but rather should be construed broadly within its scope as defined in the appended claims, and therefore all changes and modifications that fall within the meets and bounds of the claims, or equivalences of such meets and bounds are therefore intended to be embraced by the appended claims.
Claims (2)
1. An automatic immunohistochemical staining system for biological tissues, which is characterized in that: comprises a glass dyeing pipe (1), a primary antibody liquid storage tank (10), a secondary antibody liquid storage tank (11), a PBT buffer solution liquid storage tank (12), a formaldehyde fixing liquid storage tank (13), a liquid inlet pipe (4), a liquid outlet pipe (5), a plurality of connecting pipes (3) and a waste liquid tank (6); one end of the glass dyeing tube (1) is opened, a sealing plug (7) is arranged at the opening of the glass dyeing tube (1), a liquid inlet hole (8) and a liquid outlet hole (9) are formed in the sealing plug (7), and a liquid outlet tube protection cavity (2) is arranged on the inner bottom side of the glass dyeing tube (1); the anti-liquid storage tank (10), the secondary antibody liquid storage tank (11), the PBT buffer solution liquid storage tank (12) and the formaldehyde fixing liquid storage tank (13) are arranged above one side of the glass dyeing pipe (1), the waste liquid tank (6) is arranged below one side of the glass dyeing pipe (1), outlets of the anti-liquid storage tank (10), the secondary antibody liquid storage tank (11), the PBT buffer solution liquid storage tank (12) and the formaldehyde fixing liquid storage tank (13) are respectively connected with the liquid inlet pipe (4) through the connecting pipe (3), the liquid inlet pipe (4) penetrates through a liquid inlet hole (8) in the sealing plug (7) and extends into the glass dyeing pipe (1), one end of the liquid outlet pipe (5) penetrates through a liquid outlet hole (9) in the sealing plug (7) and extends into the liquid outlet pipe protection cavity (2), and the other end of the liquid outlet pipe (5) is connected with the waste liquid tank; a micro liquid control valve (14) controlled by a singlechip control module is arranged on each connecting pipe (3) between the first anti-liquid storage tank (10), the second anti-liquid storage tank (11), the PBT buffer solution storage tank (12), the formaldehyde fixing liquid storage tank (13) and the liquid inlet pipe (4), and a liquid pump (15) controlled by the singlechip control module is arranged on the liquid outlet pipe (5); be equipped with the low temperature controlling means by single chip microcomputer control module control on anti liquid storage pot (10), two anti liquid storage pot (12), be equipped with infrared temperature probe (17) on glass dyeing pipe (1), infrared temperature probe (17) are connected with single chip microcomputer control module, the export end of feed liquor pipe (4) bends to dyeing pipe (1) inner wall, be equipped with the logical liquid aperture that a plurality of prevention biological tissue passed through on drain pipe protection chamber (2) lateral wall, be located be equipped with the filter screen on the drain pipe in drain pipe protection chamber (2).
2. The system of claim 1, wherein the staining system comprises: the glass dyeing tube (1) is fixed on a warped plate shaking table (16), and the warped plate shaking table shakes according to a certain frequency; the liquid inlet pipe (4) and the liquid outlet pipe (5) are flexible pipes.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201711087020.7A CN107907396B (en) | 2017-11-07 | 2017-11-07 | Automatic immunohistochemical staining system for biological tissues |
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| CN201711087020.7A CN107907396B (en) | 2017-11-07 | 2017-11-07 | Automatic immunohistochemical staining system for biological tissues |
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| CN107907396B true CN107907396B (en) | 2020-11-10 |
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| CN111983245A (en) * | 2019-05-24 | 2020-11-24 | 南京金斯瑞生物科技有限公司 | Biological reaction device and biological detection method based on same |
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