CN111896746A - Western blot multi-antibody whole-membrane automatic incubation box - Google Patents
Western blot multi-antibody whole-membrane automatic incubation box Download PDFInfo
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- CN111896746A CN111896746A CN202010758239.0A CN202010758239A CN111896746A CN 111896746 A CN111896746 A CN 111896746A CN 202010758239 A CN202010758239 A CN 202010758239A CN 111896746 A CN111896746 A CN 111896746A
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- 238000011534 incubation Methods 0.000 title claims abstract description 98
- 239000012528 membrane Substances 0.000 title claims abstract description 46
- 238000001262 western blot Methods 0.000 title claims abstract description 27
- 238000005192 partition Methods 0.000 claims abstract description 51
- 239000012530 fluid Substances 0.000 claims abstract description 43
- 239000007788 liquid Substances 0.000 claims abstract description 28
- 238000000638 solvent extraction Methods 0.000 claims abstract description 5
- 238000007789 sealing Methods 0.000 claims description 6
- 230000000903 blocking effect Effects 0.000 claims description 5
- 108090000623 proteins and genes Proteins 0.000 abstract description 23
- 102000004169 proteins and genes Human genes 0.000 abstract description 23
- 238000002474 experimental method Methods 0.000 abstract description 10
- 238000000034 method Methods 0.000 abstract description 10
- 238000010008 shearing Methods 0.000 abstract description 2
- 230000002572 peristaltic effect Effects 0.000 description 17
- 239000006180 TBST buffer Substances 0.000 description 16
- 239000002699 waste material Substances 0.000 description 11
- 239000008267 milk Substances 0.000 description 10
- 210000004080 milk Anatomy 0.000 description 10
- 235000013336 milk Nutrition 0.000 description 10
- 239000002033 PVDF binder Substances 0.000 description 4
- 229920002981 polyvinylidene fluoride Polymers 0.000 description 4
- 238000011084 recovery Methods 0.000 description 3
- 230000000694 effects Effects 0.000 description 2
- 230000014509 gene expression Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 241000592183 Eidolon Species 0.000 description 1
- -1 antibody Substances 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000007599 discharging Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000001704 evaporation Methods 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 238000001502 gel electrophoresis Methods 0.000 description 1
- 230000002998 immunogenetic effect Effects 0.000 description 1
- 238000003780 insertion Methods 0.000 description 1
- 230000037431 insertion Effects 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 238000002331 protein detection Methods 0.000 description 1
- 239000012474 protein marker Substances 0.000 description 1
- 238000005057 refrigeration Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 238000010186 staining 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
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/5302—Apparatus specially adapted for immunological test procedures
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Abstract
The invention relates to a Western blot multi-antibody whole-membrane automatic incubation box, belonging to the technical field of molecular biology experiments. Comprises a separable incubation box, a partition plate and a fluid pipeline; partition boards for partitioning the inside of the incubation boxes are arranged in the separable incubation boxes; the partition plate is provided with fluid pipelines for conveying and recovering different liquids. The invention solves the problems that the incubation volume in the incubation box in the Western blot experiment can be adjusted, the automatic operation can be realized, the whole membrane can expose a plurality of different proteins, and the invention can also be suitable for the incubation of a plurality of membranes in the same incubation box after membrane shearing, saves the antibody and time, and provides a basic device for realizing the automatic control of the Western blot experiment process.
Description
Technical Field
The invention relates to a Western blot multi-antibody whole-membrane automatic incubation box, belonging to the technical field of molecular biology experiments.
Background
Western Blot, is a commonly used experimental method in molecular biology, biochemistry and immunogenetics. The basic principle is to stain a gel electrophoresis treated cell or biological tissue sample with a specific antibody. Information on the expression of a specific protein in the analyzed cell or tissue is obtained by analyzing the location and depth of staining.
The Western immunoblotting (Western Blot) is a protein detection technique in which the total protein of cells or tissues after electrophoretic separation is transferred from a gel to a solid support NC membrane or a PVDF membrane, and then a specific antibody is used to detect a specific antigen, and has been widely used in many fields such as gene expression studies at the protein level, antibody activity detection, and early disease diagnosis. In the current experimental process, technicians in the technical field often feel that the process of incubating the antibodies is complicated and the waiting time is long, and different antibodies need to be artificially cut in order to be incubated simultaneously, so that the finally exposed strips are not on the same membrane, and the sizes of all target protein strips are not easy to judge according to a protein Marker. The existing method is operated manually, so that the operation difference between people is large, antibodies are easy to waste, and the experiment cost is increased; some current automatic incubation devices can not solve the purpose of incubating a plurality of antibodies simultaneously on the same membrane, and the current incubation box often volume can not be adjusted moreover, can not reach the purpose that the experimenter practices thrift the antibody.
Disclosure of Invention
The invention aims to solve the technical problems that the incubation volume of an incubation box can be adjusted and the incubation process is automated when a plurality of target proteins are incubated on the same membrane at the same time.
In order to solve the problems, the technical scheme adopted by the invention is to provide a Westernblot multi-antibody whole-membrane automatic incubation box, wherein one end of the incubation box is provided with an opening, and the incubation box comprises an incubation box, a partition plate and a fluid pipeline which can be separated; the separable incubation box is internally provided with a partition plate for partitioning the interior of the incubation box; the partition plate is provided with a fluid pipeline.
Preferably, parallel partition plates for partition are arranged in the separable incubation boxes.
Preferably, the fluid conduit is provided as a conduit open at both ends.
Preferably, the fluid conduit is perpendicular to the lower end face of the compartmentalized incubation well.
Preferably, one end opening of the fluid pipeline is connected with the lower end face of the incubation box.
Preferably, the number of fluid conduits is greater than or equal to 1.
Preferably, the partition panel is in plug-in connection with the isolatable incubation cassette.
Preferably, the plug-in connection position of the partition board and the separable incubation box and the contact connection position of the partition board and the whole multi-antibody membrane are provided with sealing devices for blocking liquid.
Preferably, the periphery of the partition plate is provided with a sealing strip for blocking liquid at the splicing connection part with the separable incubation box and the contact connection part with the whole multi-antibody membrane.
Preferably, the partition plate is provided with a scale for displaying the height of the liquid in the incubation box; the side wall of the separable incubation box is provided with scales for measuring and displaying the distance between the partition boards.
Compared with the prior art, the invention has the following beneficial effects:
the invention solves the problems that the incubation volume in an incubation box in a Western blot experiment can be adjusted, the automatic operation can be realized, the whole membrane can expose a plurality of different proteins (or a certain protein can be incubated selectively, and all protein markers are reserved), and the invention can also be suitable for the incubation of a plurality of membranes in an incubation box after membrane shearing, thereby saving the antibody and time.
The invention realizes that different antibodies can be directly incubated at different positions without cutting PVDF or NC membranes. If the PVDF or NC membrane is cut, the effect of incubating a plurality of strip proteins by the same antibody incubation box can be realized.
Drawings
FIG. 1 is a schematic structural diagram of a Western blot multi-antibody whole membrane automatic incubation box provided by the invention;
FIG. 2 is a schematic view of a top view structure of a Western blot multi-antibody whole membrane automatic incubation box provided by the invention;
reference numerals: 1. an incubation box; 2. a partition panel; 3. a fluid conduit; 4. a film; 5. a peristaltic pump; 6. marking a Marker; 7. a protein of interest;
Detailed Description
In order to make the invention more comprehensible, preferred embodiments are described in detail below with reference to the accompanying drawings:
as shown in figures 1-2, the invention provides a Western blot multi-antibody whole membrane automatic incubation box, wherein one end of the incubation box 1 is provided with an opening, and the incubation box comprises an incubation box 1, a partition plate 2 and a fluid pipeline 3 which can be separated; the separable incubation box 1 is internally provided with a partition plate 2 for partitioning the interior of the incubation box 1; the partition plate 2 is provided with a fluid conduit 3. Partition plates 2 which are parallel and used for partitioning are arranged in the separable incubation boxes 1; the fluid pipe 3 is a pipe with openings at both ends; the fluid pipeline 3 is vertical to the lower end surface of the separable incubation box 1; one end opening of the fluid conduit 3 is provided to be connected to the lower end face of the incubation box 1. The number of fluid conduits 3 is equal to or greater than 1. The partition plate 2 is in plug-in connection with the isolatable incubation box 1. The plug-in connection of the partition board 2 and the separable incubation box 1 and the contact connection of the partition board and the whole multi-antibody membrane 4 are provided with sealing devices for separating liquid. Sealing strips for blocking liquid are arranged at the insertion connection part of the periphery of the partition board 2 and the separable incubation box 1 and the contact connection part of the periphery of the partition board and the whole multi-antibody membrane 4. The partition plate 2 is provided with scales for displaying the height of the liquid in the incubation box 1; the side wall of the separable incubation box 1 is provided with scales for measuring and displaying the distance between the partition boards.
Examples
As shown in figures 1-2, when Western blot experiment is carried out, the incubation box 1 of the invention is placed on a base with a refrigeratable base, the base can be tilted and shaken, and when liquid (milk, antibody, TBST and the like) is conveyed and recovered under the action of a peristaltic pump 5, the base can be tilted when one stage of incubation work is finished, so that the liquid can be conveniently and completely discharged by a fluid pipeline 3. A plurality of incubation boxes can be placed on the base; the base is provided with a cover, and the edge of the base is provided with a reservoir for keeping certain humidity and preventing the evaporation of the antibody.
The incubation box 1 is matched with a plurality of partition plates 2, and the partition plates 2 are marked with marks and respectively correspond to different antibody peristaltic tubes; a multi-antibody whole membrane 4 is arranged between the bottom of the incubation box 1 and the lower end of the partition plate 2; judging the approximate position of each target protein 7 on the whole multi-antibody membrane 4 according to the position of a Marker6 which represents the molecular weight of the protein and is displayed on the whole multi-antibody membrane 4, wherein each target protein 7 is positioned in a different incubation space because the partition plate 2 divides the incubation box 1 into different incubation spaces; scales are marked on the side wall of the incubation box 1 and the partition plate 2, and the volume of each incubation space can be calculated according to the scales, namely the volume of the antibody liquid required in each incubation space can be calculated; the required incubation liquid is conveyed to different incubation spaces through the fluid pipeline 3 by the peristaltic pump 5, finally, a plurality of target proteins 7 can be incubated on one membrane at the same time, and a plurality of strips of the target proteins 7 can be exposed on the whole membrane.
Each partition plate 2 is connected on one side with 5 fluid conduits 3, one for the addition and recovery of primary antibodies (primary antibody fluid conduit), one for the addition and recovery of secondary antibodies (secondary antibody fluid conduit), one for the addition of TBST (TBST fluid conduit), one for the addition of milk (milk fluid conduit), and one for the recovery of waste fluid (waste fluid conduit).
Four paths of mobile phone remote control devices are arranged to control the switching time or the forward/reverse rotation of each peristaltic pump, so that the aim of adding or sucking out liquid is fulfilled. The processes of conveying the incubation liquid and discharging the incubation liquid are automatically controlled through remote operation of a mobile phone or linkage with voice assistants such as a tianmao eidolon and the like; the purpose of automatically controlling the experimental process is achieved.
The invention relates to a method for using a Western blot multi-antibody whole-membrane automatic incubation box, which comprises the following steps:
and controlling the incubation time and the cleaning time of the milk, the TBST, the primary antibody and the secondary antibody through the corresponding App of the mobile phone. The following takes the example of multiple target proteins per membrane at a time: as shown in fig. 2;
after the membrane is transferred by Western blot, putting the PVDF/NC membrane into an incubation box 1, and inserting a corresponding partition plate 2 into the incubation box 1 according to the relative position of a protein marker6 corresponding to the molecular weight of a target protein 7;
2. TBST fluid pipeline on the partition panel 2, milk fluid pipeline, waste liquid fluid pipeline and TBST, milk, waste liquid storage container UNICOM, according to the different protein that the incubation space that every partition panel 2 cut off corresponds respectively through different fluid pipeline 3 insert different first anti liquid, with two anti liquid. The difference in the scale between the partition plates 2 and the scale on the incubation cassette indicating the height of the liquid can help to determine the volume of antibody that needs to be added.
3. And (3) setting a program (which can be stored as a scene after setting) by using a mobile phone, starting the base in advance for refrigeration, and preparing 5% milk, TBST, different primary antibodies and different secondary antibodies into corresponding storage containers.
When the experiment begins, adding milk with the volume of 5% of the incubation space through a milk fluid pipeline by a peristaltic pump 5, incubating for 1 hour, and starting a waste liquid peristaltic pump to pump out the milk; adding TBST through a TBST fluid pipeline by a peristaltic pump 5, incubating for 15 minutes, starting a waste liquid peristaltic pump to pump out the TBST, and repeating for 3 times; adding primary antibodies (according to the distance between the partition plates and the corresponding added volume; different proteins corresponding to different incubation spaces are respectively connected with different primary antibodies through the fluid pipelines 3) through the primary antibody fluid pipeline by the peristaltic pump 5, and starting the waste liquid peristaltic pump to suck out the primary antibodies after incubation for 12 hours; adding TBST through a TBST fluid pipeline by a peristaltic pump 5, starting a waste liquid peristaltic pump to pump out TBST after 15 minutes, and repeating for 3 times; adding secondary antibody through a secondary antibody fluid pipeline by a peristaltic pump 5 (preparing a corresponding adding volume according to the distance between the partition plates; respectively accessing different proteins corresponding to each incubation space into different secondary antibody fluids through a fluid pipeline 3), and starting a waste liquid peristaltic pump to pump out the secondary antibody after incubation for 1 hour; adding TBST through a TBST fluid pipeline by a peristaltic pump 5, starting a waste liquid peristaltic pump to pump out TBST after 15 minutes, and repeating for 3 times; finally, manually adding a developing solution to prepare for exposing the target protein; the experiment was completed.
While the invention has been described with respect to a preferred embodiment, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention. Those skilled in the art can make various changes, modifications and equivalent arrangements, which are equivalent to the embodiments of the present invention, without departing from the spirit and scope of the present invention, and which may be made by utilizing the techniques disclosed above; meanwhile, any changes, modifications and variations of the above-described embodiments, which are equivalent to those of the technical spirit of the present invention, are within the scope of the technical solution of the present invention.
Claims (10)
1. The utility model provides a box is hatched automatically to whole membrane of Western blot many antibodies, hatches box one end and is equipped with the opening, its characterized in that: comprises a separable incubation box, a partition plate and a fluid pipeline; the separable incubation box is internally provided with a partition plate for partitioning the interior of the incubation box; the partition plate is provided with a fluid pipeline.
2. The Western blot multi-antibody whole membrane automatic incubation box of claim 1, wherein: and parallel partition plates for partition are arranged in the separable incubation boxes.
3. The Western blot multi-antibody whole membrane automatic incubation box of claim 2, which is characterized in that: the fluid pipeline is a pipeline with two open ends.
4. The Western blot multi-antibody whole membrane automatic incubation box of claim 3, which is characterized in that: the fluid pipeline is perpendicular to the lower end face of the separable incubation box.
5. The Western blot multi-antibody whole membrane automatic incubation box of claim 4, wherein: an opening at one end of the fluid pipeline is connected with the lower end face of the incubation box.
6. The Western blot multi-antibody whole membrane automatic incubation box of claim 5, wherein: the number of the fluid pipelines is more than or equal to 1.
7. The Western blot multi-antibody whole membrane automatic incubation box of claim 6, wherein: the partition board is connected with the separable incubation box in an inserting mode.
8. The Western blot multi-antibody whole membrane automatic incubation box of claim 7, wherein: and sealing devices for blocking liquid are arranged at the splicing connection part of the partition board and the separable incubation box and the contact connection part of the partition board and the whole multi-antibody membrane.
9. The Western blot multi-antibody whole membrane automatic incubation box of claim 8, wherein: the periphery of the partition board is provided with a sealing strip for blocking liquid at the splicing joint of the partition board and the separable incubation box and the contact joint of the partition board and the whole multi-antibody membrane.
10. The Western blot multi-antibody whole membrane automatic incubation box of claim 9, wherein: scales for displaying the height of the liquid in the incubation box are arranged on the partition plate; the side wall of the separable incubation box is provided with scales for measuring and displaying the distance between the partition boards.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010758239.0A CN111896746A (en) | 2020-07-31 | 2020-07-31 | Western blot multi-antibody whole-membrane automatic incubation box |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010758239.0A CN111896746A (en) | 2020-07-31 | 2020-07-31 | Western blot multi-antibody whole-membrane automatic incubation box |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112684161A (en) * | 2020-12-11 | 2021-04-20 | 东南大学 | Device for incubation and detection of western blot of any lane |
| CN113533760A (en) * | 2021-07-14 | 2021-10-22 | 上海市第十人民医院 | Automatic incubation device for Western blot experiment |
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| CN113533760A (en) * | 2021-07-14 | 2021-10-22 | 上海市第十人民医院 | Automatic incubation device for Western blot experiment |
| CN113533760B (en) * | 2021-07-14 | 2024-02-06 | 上海市第十人民医院 | Automatic incubation device for Western blot experiment |
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Application publication date: 20201106 |