CN101153874A - Automatic protein hybridization elution instrument - Google Patents
Automatic protein hybridization elution instrument Download PDFInfo
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- CN101153874A CN101153874A CNA200710131403XA CN200710131403A CN101153874A CN 101153874 A CN101153874 A CN 101153874A CN A200710131403X A CNA200710131403X A CN A200710131403XA CN 200710131403 A CN200710131403 A CN 200710131403A CN 101153874 A CN101153874 A CN 101153874A
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Abstract
An automatic protein hybridization elution apparatus is characterized in that the apparatus consists of a sterile cleaning system, a sample injection system, a sample discharge system and a power system, wherein, the sterile cleaning system comprises a solution cylinder and a rotary shaft with the rotary shaft arranged inside the solution cylinder; the sample injection system comprises a liquid inlet tube, a constant flow pump and a reagent container; one end of the liquid inlet tube is connected with the upper part of the solution cylinder, while the other end of the liquid inlet tube is connected with the reagent container through the constant flow pump; the sample discharge system comprises a liquid outlet tube H, a liquid outlet tube L, a switch and a waste liquid collector; one end of the liquid outlet tube H and one end of the liquid outlet tube L are respectively connected with the lower part of the solution cylinder, while the other end of the liquid outlet tub H and the other end of the liquid outlet tube L are respectively connected with the waste liquid collector; moreover, the switch is arranged on the liquid outlet tubes; the power system comprises a motor, a speed regulation switch and a power source with the motor controlled by the speed regulation switch and connected with the rotary shaft. The elution apparatus does not influence result when the number of antibody is small and can realize large-scale protein hybridization.
Description
One, technical field
The invention belongs to technical field of molecular biology, particularly a kind of automatic protein hybridization elution instrument.
Two, background technology
Prior art: protein hybridization (Western blot) is a kind of experimental technique that often use in the modern life science field, be about to transfer on the pvdf membrane after the capable electrophoresis of destination protein separates, again with known protein part (as antibody etc.) hybridization, by having or not and quantitatively of enzyme system colour developing back testing goal albumen.It comprises sealing, hybridization and three processes of wash-out, and wherein sealing and crossover process can be collectively referred to as the process of hatching.At present, the way of protein hybridization routine is the shaking table method, and the pvdf membrane behind the film of electrophoresis commentaries on classics soon is put in the glass dish, then plate is placed on the decolorization swinging table.Shake sealing at a slow speed with the confining liquid that contains bovine serum albumin(BSA) (BSA) or skimmed milk power, shake at a slow speed with the Incubating Solution that contains antibody then and hatch.Shake wash-out 4-6 time fast with eluent, each 5-10 minute.If use enzyme target two anti-, an anti-Incubating Solution of the anti-target protein of Ying Zaihan hatch and wash-out after, change and contain enzyme and mark two anti-Incubating Solutions of anti-first antibody and shake at a slow speed and hatch.Shake wash-out 4-6 time fast with eluent, each 5-10 minute.Develop the color having or not with DAB or chemoluminescence method at last with testing goal albumen with quantitative.The shortcoming of conventional way is: 1. waste time and energy, operate loaded down with trivial details; 2. the antibody amount is not easy to experiment more after a little while; 3. be not easy to the scale operation.
Three, summary of the invention
The present invention is directed to above-mentioned technical matters, provide a kind of simple to operate, also do not influence the result after a little while, can accomplish the automatic protein hybridization elution instrument of scale protein hybridization in enormous quantities in the antibody amount.
Technical solution of the present invention is: a kind of automatic protein hybridization elution instrument, and by incubating the system of washing, sampling system, going out the sample system and power system is formed, wherein to incubate the system of washing and comprise solution tube and rotating shaft, rotating shaft is located in the solution tube; Sampling system comprises feed tube, constant flow pump and reagent container, and feed tube one end links to each other with solution tube top, and the other end links to each other with reagent container by constant flow pump; Go out the sample system and comprise drain pipe H, drain pipe L, switch and waste collection device, the end of drain pipe H and drain pipe L links to each other with solution tube bottom respectively, and the other end links to each other with the waste collection device, and switch is located on the drain pipe; Power system comprises motor, speed-regulating switch and power supply, and motor is controlled by speed-regulating switch, and links to each other with rotating shaft.Incubating the system of washing can be connected with some sampling systems.Be arranged with into film mouth and film outlet port on the solution tube.The solution tube can link to each other with 4 feed tubes.Rotating shaft is provided with the film stationary installation, and the film stationary installation is provided with magnetic film folder and diaphragm holder.The pvdf membrane that electrophoresis is changeed behind the film is fixed in the rotatable rotating shaft with adhesive tape, and rotating shaft is put in the solution tube, by sampling system confining liquid is joined in the solution tube, and rotating shaft is with 2-10 rev/min low speed rotation sealing.By going out the sample system confining liquid is emitted, by sampling system antibody-solutions is joined in the solution tube then, rotating shaft is hatched with 2-10 rev/min low speed rotation.Hatch and antibody-solutions is emitted by going out the sample system after finishing, by sampling system eluent is joined in the solution tube constantly then, rotating shaft is with 100-200 rev/min high speed rotating wash-out.When the eluent that adds reached the top of drain pipe H, unnecessary eluent was discharged in the waste liquid bottle by drain pipe H, and new like this eluent constantly enters, and unnecessary eluent is constantly discharged, thereby has realized continuous wash-out.Wash-out has saved the step of constantly changing eluent in the conventional method continuously, has saved time and manpower.Because the interstitial cavities between rotating shaft and the solution tube can be very little, therefore can under the less situation of antibody amount, experimentize, saved antibody.Same rotating shaft can many pvdf membranes of parallel attached subsides, thereby can realize scale operations.Hatch with the process of wash-out and be convenient to control, experiment condition is consistent, so result's good reproducibility.The method that rotation is hatched can increase the chance of antigen-antibody contact, therefore can improve hybridization efficiency.In conjunction with automatic control system, can realize that sealing is hatched and wash-out automatically, thereby realize that experiment automatically.This protein hybridization elution instrument comprises four systems, promptly sampling system, power system, go out the sample system and incubate the system of washing.Sampling system comprises feed tube, constant flow pump and reagent bottle three parts.Reagent bottle is the container of splendid attire confining liquid, antibody-solutions and washing lotion, an a kind of solution of reagent bottle splendid attire.Under the promotion of constant flow pump, solution enters into interstitial cavities between solution tube and the rotating shaft via feed tube.Power system comprises power supply, motor and speed-regulating switch three parts.Speed-regulating switch can be regulated the rotating speed of motor, and motor speed is lower when hatching, and motor speed is higher during wash-out.Motor links to each other with the rotating shaft of incubating the system of washing, and rotates thereby drive rotating shaft.Go out the sample system and comprise drain pipe H, drain pipe L, switch and waste liquid bottle four parts.Liquid level of solution is lower than the top of drain pipe H when hatching, and the switch of drain pipe L is in closed condition; Liquid level of solution is equal with the top of drain pipe H during wash-out, and the switch of drain pipe L is in closed condition, and the feed liquor system is in running order simultaneously; Open the switch of drain pipe L during the discharging waste liquid, waste liquid is discharged in the waste liquid bottle by drain pipe L.Incubate the system of washing and comprise solution tube and rotating shaft two parts.Pvdf membrane is fixed in the rotating shaft by adhesive tape.The solution tube is the container of an outside, in holding rotating shaft.
The invention has the beneficial effects as follows: confining liquid, protein hybridization liquid, eluent etc. are successively continuously through sampling system input solution tube, emit from going out the sample system successively again, the eluent that wash-out is stylish constantly enters the solution tube by feed tube, unnecessary eluent is constantly discharged by drain pipe, thereby realized continuous wash-out, save the step of constantly changing eluent in the conventional method, sealed automatically in the experiment, hybridization and wash-out, saved time and manpower.Because the interstitial cavities between rotating shaft and the solution tube can be very little, therefore can under the less situation of antibody amount, experimentize, saved antibody.Same rotating shaft can many pvdf membranes of parallel attached subsides, thereby can realize scale operations.Hatch with the process realization of wash-out and control automatically, help keeping the consistance of experiment condition, result's good reproducibility.The method that rotation is hatched can increase the chance of antigen-antibody contact, therefore can improve hybridization efficiency.
Four, description of drawings
Fig. 1 protein hybridization elution instrument structural representation.
Fig. 2 protein hybridization elution instrument is incubated battle array and is educated the state sketch.
Fig. 3 protein hybridization elution instrument wash-out state sketch.
Fig. 4 evaluation experimental is figure as a result.
Fig. 5 automatic protein hybridization elution instrument is incubated and is washed the unit architecture diagrammatic side views.
Fig. 6 pvdf membrane stationary installation sketch.
1. feed tubes among the figure, 2. constant flow pump, 3. reagent container, 4. motor, 5. speed-regulating switch, 6. power supply, 7. drain pipe H, 8. switch, 9. drain pipe L, 10. waste collection device, 11. solution tubes, 12. rotating shafts, 13.PVDF film, 14a. first feed tube, 14b, second feed tube, 14c the 3rd feed tube, 14d the 4th feed tube, 16. magnetic film folder 17. advances the film mouth, 18. film outlet ports, 19. diaphragm holders.
Five, embodiment
Embodiment 1:
A kind of automatic protein hybridization elution instrument by incubating the system of washing, sampling system, going out the sample system and power system is formed, is wherein incubated the system of washing and is comprised solution tube 11 and rotating shaft 12, and rotating shaft 12 is located in the solution tube 11; Sampling system comprises feed tube 1, constant flow pump 2 and reagent container 3, and feed tube 1 one ends link to each other with solution tube 11 tops, and the other end links to each other with reagent container 3 by constant flow pump 2; Go out the sample system and comprise drain pipe H 7, drain pipe L 9, switch 8 and waste collection device 10, the end of drain pipe H 7 and drain pipe L 9 links to each other with solution tube 11 bottoms respectively, and the other end links to each other with waste collection device 10, and switch 8 is located on the drain pipe; Power system comprises motor 4, speed-regulating switch 5 and power supply 6, and motor 6 is controlled by speed-regulating switch 5, and links to each other with rotating shaft 12.Incubating the system of washing can be connected with some sampling systems.Be arranged with into film mouth 17 and film outlet port 18 on the solution tube.On the solution tube step respectively with the first feed tube 14a, the second feed tube 14b, the 3rd feed tube 14c 14d that links to each other with the 4th feed tube.Rotating shaft is provided with the film stationary installation, and the film stationary installation is provided with magnetic film folder 16 and diaphragm holder 19.
Embodiment 2:
This protein hybridization elution instrument comprises four systems, promptly sampling system, power system, go out the sample system and incubate the system of washing.Sampling system comprises feed tube 1, constant flow pump 2 and reagent container 3 three parts.Reagent container 3 is containers of splendid attire confining liquid, antibody-solutions and eluent, an a kind of solution of reagent bottle splendid attire.Under the promotion of constant flow pump 2, solution enters into interstitial cavities between solution tube 11 and the rotating shaft 12 via feed tube 1.Power system comprises motor 4, speed-regulating switch 5 and power supply 6 three parts.Speed-regulating switch 5 can be regulated the rotating speed of motor 4, and motor 4 rotating speeds are lower when hatching, and motor 4 rotating speeds are higher during wash-out.Motor 4 links to each other with the rotating shaft 12 of incubating the system of washing, and rotates thereby drive rotating shaft 12.Go out the sample system and comprise drain pipe H 7, switch 8, drain pipe L 9 and waste liquid bottle 10 4 parts.Liquid level of solution is lower than the top of drain pipe H 7 when hatching, and the switch 8 of drain pipe L 9 is in closed condition; Liquid level of solution is equal with the top of drain pipe H 7 during wash-out, and the switch 8 of drain pipe L9 is in closed condition, and the feed liquor system is in running order simultaneously; Open the switch 8 of drain pipe L 9 during the discharging waste liquid, waste liquid is discharged in the waste liquid bottle 10 by drain pipe L 9.Incubate the system of washing and comprise solution tube 11 and rotating shaft 12 two parts.Pvdf membrane 13 is fixed in the rotating shaft 12 by adhesive tape.Solution tube 11 is containers of an outside, in holding rotating shaft 12.Being illustrated in figure 5 as automatic protein hybridization elution instrument incubates and washes the unit architecture diagrammatic side views.Automatic protein hybridization elution instrument is provided with a plurality of feed tubes, can advance confining liquid respectively, eluent and antibody-solutions etc.The diaphragm that pvdf membrane 13 will be housed when advancing film props up in advancing film mouth 17, is adsorbed in the rotating shaft 12 by magnetic film folder 16.Incubating Solution is entered to incubate by corresponding feed tube and washes the chamber when hatching, and drain pipe is in closed condition, rotating shaft 12 low speed rotation; Eluent is continued to enter to incubate by corresponding feed tube and washes the chamber during wash-out, and fluid speed is identical with feed liquor speed, incubates the eluent of washing constant basis in the chamber thereby keep, rotating shaft 12 high speed rotating.After experiment was finished, pvdf membrane shoveled out automatic protein hybridization elution instrument by the valves of film outlet port 18.Be illustrated in figure 6 as pvdf membrane stationary installation sketch.Pvdf membrane 13 is fixed in the diaphragm holder 19 by the magnetic film folder 16 at two ends, adsorbs thereon according to the magnetic force of 16 pairs of irony rotating shafts of magnetic film folder then.
Embodiment 3:
As shown in Figure 2, pvdf membrane is fixed in the rotating shaft by adhesive tape, and confining liquid or antibody-solutions enter in the interstitial cavities between solution tube and the rotating shaft by sampling system, and its liquid level is lower than the top of drain pipe H, and the switch of drain pipe L is in closed condition.Close constant flow pump behind the sample introduction.Regulate speed-regulating switch, make rotating shaft, represent with hollow arrow among the figure with 2-10 rev/min low speed rotation.With the switch opens of drain pipe L, Incubating Solution is discharged in the waste liquid bottle by drain pipe L when hatching end.
Embodiment 4:
Be illustrated in figure 3 as protein hybridization elution instrument wash-out state embodiment.The switch of drain pipe L is in closed condition, and eluent enters in the interstitial cavities between solution tube and the rotating shaft by sampling system, and when liquid level arrived the top of drain pipe H, unnecessary eluent was discharged into waste liquid bottle by drain pipe H.Regulate speed-regulating switch, make rotating shaft, represent with two hollow arrow among the figure with 100-200 rev/min high speed rotating.Constant flow pump is lasting in running order in whole elution process, and new like this washing lotion constantly enters, and unnecessary washing lotion is constantly discharged, thereby has realized continuous wash-out.Filled arrows is depicted as the washing lotion flow direction among the figure.When finishing, wash-out, eluent is discharged in the waste liquid bottle by drain pipe L with the switch opens of drain pipe L.
Embodiment 5:
Be illustrated in figure 4 as evaluation experimental figure as a result.Sample is reorganization HLA-A*0181 heavy chain molecule, and one is anti-for HC-10, is diluted in PBST at 1: 10000.Two anti-are the sheep anti-mouse igg antibody of HRP mark, are diluted in PBST at 1: 10000.The concrete operations of conventional (RM) method and protein hybridization elution (IW) method contrast in following table to be listed.1,2 and 3 sample sizes that swimming lane adds are respectively 1 μ L, 2 μ L and 3 μ L among the figure.Used substrate is the Pico level among the figure A, and the time shutter is 118 seconds.Used substrate is that 96% Pico level adds 4% Femto level among the figure B, and the time shutter is 18 seconds.The evaluation experimental result shows that compare with conventional method, protein hybridization elution instrument is incubated at the bottom of the exposed background of washing method, and hybridization signal is strong, and the hybridization efficiency height is described.
| Conventional method | The IW method | |
| The sealing one anti-wash-out two anti-wash-outs of hatching of hatching | RT，1h RT，1h 60mL(10mL X 6) 1h(10min X 6) RT，1h 60mL(10mL X 6) 1h(10minX 6) | RT，1h RT，1h 30mL 20min RT，1h 30mL 20min |
Claims (6)
1. automatic protein hybridization elution instrument is characterized in that wherein incubating the system of washing and comprising solution tube (11) and rotating shaft (12) by incubating the system of washing, sampling system, going out the sample system and power system is formed, and rotating shaft (12) is located in the solution tube (11); Sampling system comprises feed tube (1), constant flow pump (2) and reagent container (3), and feed tube (1) one end links to each other with solution tube (11) top, and the other end links to each other with reagent container (3) by constant flow pump (2); Go out the sample system and comprise drain pipe H (7), drain pipe L (9), switch (8) and waste collection device (10), the end of drain pipe H (7) and drain pipe L (9) links to each other with solution tube (11) bottom respectively, the other end links to each other with waste collection device (10), and switch (8) is located on the drain pipe; Power system comprises motor (4), speed-regulating switch (5) and power supply (6), and motor (6) is controlled by speed-regulating switch (5), and links to each other with rotating shaft (12).
2. automatic protein hybridization elution instrument according to claim 1 is characterized in that described rotating shaft (12) is the magnetic metal material.
3. automatic protein hybridization elution instrument according to claim 1 is characterized in that the described system of washing of incubating can be connected with some sampling systems.
4. automatic protein hybridization elution instrument according to claim 1 is characterized in that being arranged with on the described solution tube into film mouth (17) and film outlet port (18).
5. automatic protein hybridization elution instrument according to claim 1, it is characterized in that described solution tube top respectively with first feed tube (14a), second feed tube (14b), the 3rd feed tube (14c) link to each other with the 4th feed tube (14d).
6. automatic protein hybridization elution instrument according to claim 1 is characterized in that described rotating shaft is provided with the film stationary installation, and the film stationary installation is provided with magnetic film folder (16) and diaphragm holder (19), and magnetic film folder (16) is located at the two ends of film stationary installation.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA200710131403XA CN101153874A (en) | 2007-08-28 | 2007-08-28 | Automatic protein hybridization elution instrument |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA200710131403XA CN101153874A (en) | 2007-08-28 | 2007-08-28 | Automatic protein hybridization elution instrument |
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| CN101153874A true CN101153874A (en) | 2008-04-02 |
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| Application Number | Title | Priority Date | Filing Date |
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| CNA200710131403XA Pending CN101153874A (en) | 2007-08-28 | 2007-08-28 | Automatic protein hybridization elution instrument |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101923096B (en) * | 2009-06-12 | 2013-01-02 | 上海迅达医疗仪器有限公司 | Automatic westem blot analyzer |
| CN103308671A (en) * | 2013-05-22 | 2013-09-18 | 北京万泰生物药业股份有限公司 | Detection film and detection system |
| CN109752532A (en) * | 2017-11-03 | 2019-05-14 | 中国人民解放军军事医学科学院放射与辐射医学研究所 | A kind of incubation sampling device based on the detection of multichannel immunochromatography |
-
2007
- 2007-08-28 CN CNA200710131403XA patent/CN101153874A/en active Pending
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101923096B (en) * | 2009-06-12 | 2013-01-02 | 上海迅达医疗仪器有限公司 | Automatic westem blot analyzer |
| CN103308671A (en) * | 2013-05-22 | 2013-09-18 | 北京万泰生物药业股份有限公司 | Detection film and detection system |
| CN109752532A (en) * | 2017-11-03 | 2019-05-14 | 中国人民解放军军事医学科学院放射与辐射医学研究所 | A kind of incubation sampling device based on the detection of multichannel immunochromatography |
| CN109752532B (en) * | 2017-11-03 | 2022-04-15 | 中国人民解放军军事医学科学院放射与辐射医学研究所 | Incubation sample loading device based on multi-channel immunochromatography detection |
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Open date: 20080402 |