CN103197055A - Method for reducing background of vector in biological reaction - Google Patents
Method for reducing background of vector in biological reaction Download PDFInfo
- Publication number
- CN103197055A CN103197055A CN2013100781567A CN201310078156A CN103197055A CN 103197055 A CN103197055 A CN 103197055A CN 2013100781567 A CN2013100781567 A CN 2013100781567A CN 201310078156 A CN201310078156 A CN 201310078156A CN 103197055 A CN103197055 A CN 103197055A
- Authority
- CN
- China
- Prior art keywords
- carrier
- dye
- biological
- deionized water
- biological respinse
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The present invention relates to a method for reducing background of a vector in a biological reaction, and belongs to the technical field of biological detection. According to the method for reducing the background of the vector in the biological reaction, the used biological reaction vector is a colored vector formed by coloring process. The biological reaction vector is a vector which can be combined with coated biological materials. The method of the present invention can significantly reduce the background during biological reaction, and improve the detection sensitivity, and has the advantages of low cost and easiness to operate.
Description
Technical field
The present invention relates to a kind of method that reduces carrier background when biological respinse, belong to technical field of biological.
Background technology
Current, bio-reaction system is widely used in each research field of life sciences such as genome research, medical research and study of pharmacy, and progressively permeates to each big field such as preferred good child-rearing of new drug development, clinical diagnosis, health forecast, disease treatment, judicial expertise, military protection, Food Hygiene Surveillance, commerce and trade inspection and quarantine, environmental monitoring control and farming, forestry, husbandary and fishing crop.Particularly aspect the disease detection diagnosis, bio-reaction system has special advantages.On a carrier, can carry out the detection of multiple disease or index simultaneously to a plurality of patients, thereby provide a large amount of medical diagnosis on disease information for the medical worker at short notice.For example to the clinical examination of common diseases such as genetic disease, tumour and communicable disease and frequently-occurring disease, all can the applying biological reaction technology.People even can have " individual laboratory " can monitor the health status of oneself whenever and wherever possible from now on.
Bio-reaction system is according to special interactional principle between biomolecule, with the biochemical analysis process integration in carrier surface, thereby realize high flux fast detecting to DNA, RNA, polypeptide, protein and other biological composition.At present prepare bio-reaction system and mainly adopt the method for surface chemistry or the method for combinatorial chemistry to handle solid-phase matrix such as polystyrene board, silicon chip, cellulose membrane or nylon membrane etc., DNA, RNA, polypeptide, protein and other biological composition are arranged on the carrier by particular requirement.But usually run into the problem of background values height, difference in signal strength in detecting.
Application number is that the Chinese invention patent of 200710132686.X discloses a kind of biological chip glass carrier, and the making of this biological chip glass carrier comprises the steps: under (1) room temperature condition, earlier with silica sand and sodium carbonate mixing; (2) add MnO
2, borax, NaNO
3, carbon dust, titanium dioxide two arsenic, K
2CO
3, stir and evenly mix; (3) above-mentioned compound is dropped into smelting furnace, heating is melt into pulpous state, and continuous stirring 1 hour; (4) be cooled to 25～40 ℃ rapidly; (5) section; (6) polishing, moulding.The background values of this invention sheet base is better than common microslide sheet base, the signal intensity aspect, and this invention microslide average signal strength is better than common microslide.But these advantages all remain further to be promoted.Because the intrinsic shortcoming of glass carrier itself makes it to have limitation, what present most biological detection reagent kit adopted is the carrier of polystyrene board carrier, filter membrane class.
Summary of the invention
The objective of the invention is for solving the problems of the technologies described above, a kind of method that reduces carrier background when biological respinse is provided.
Above-mentioned purpose of the present invention is achieved by the following technical programs:
A kind of method that reduces carrier background when biological respinse is before biological respinse, to the processing of painting of biological respinse carrier.
Preferred as technique scheme, described biological respinse carrier are the carriers that can be combined by biological substance with bag.
Described colouring handle used last toner be can significantly reduce after the carrier colouring background of carrier when biological respinse, raising detection sensitivity on toner.
Preferred as technique scheme, described carrier is the supporting body that miillpore filter or tygon, polystyrene, Polyvinylchloride, polypropylene or silastic material are made; Described colouring is treated to carrier in manufacture process, adopts coloring matter to be scattered in the raw material of making described carrier, makes described coloring matter evenly be blended in the carrier through moulding process.
Preferred as technique scheme, described carrier are any one in the supporting body made of miillpore filter or tygon, polystyrene, Polyvinylchloride, polypropylene, silastic material; It is before making biological detection reagent that described colouring is handled, and adopts to contaminate or be coated with the technology of dying, and in dye liquor or be coated with and be covered with dye liquor, makes described carrier become coloured carrier described carrier impregnation.
Preferred as technique scheme, described carrier are any one in the supporting body made of miillpore filter or tygon, polystyrene, Polyvinylchloride, polypropylene, silastic material; It is in the process of making biological detection reagent that described colouring is handled, and adopts to contaminate or be coated with the technology of dying, and in dye liquor or be coated with and be covered with dye liquor, makes described carrier become coloured carrier described carrier impregnation.
Preferred as technique scheme, described carrier is miillpore filter.
Described cellulose family miillpore filter refers to film that the potpourri of nitrocellulose filter, cellulose acetate membrane, cellulose acetate and cellulose nitrate makes, cellulose membrane, the plain film of glass fibre etc.
Described polyvinyl-fluoride miillpore filter refers to polyvinylidene fluoride film or poly tetrafluoroethylene.
Preferred as technique scheme, described to go up toner be that direct dyes, azoic dyes insoluble azo dyes, reactive dye, reducing dye, soluble vat dye, sulfur dye, sulphur vat dye, phthalocyanine dye, oxidation dye, the condensation dye, disperse dyes, acid dyes, acidic intermedium and acidity contain mordant dye, alkalescence and the dye of positive ion, coating.
Preferred as technique scheme, described painting methods is for contaminating.
Preferred as technique scheme, described colouring is handled and is: in the Tween-20 solution that described carrier is soaked in, take out after the soaking at room temperature earlier; Described carrier deionized water rinsing; Immerse and begin dyeing in the dye liquor, rising temperature and insulation; Take out carrier, in dye liquor, add HAC, put into carrier, soak; Remove dye liquor, use deionized water rinsing; Be soaked in a period of time in the deionized water; Remove deionized water, wash with cleansing solution; Remove cleansing solution, use deionized water rinsing; Take out washed carrier oven dry.
Further preferred as technique scheme, described colouring is handled and is: earlier described carrier is soaked in 0.1～0.5% the Tween-20 solution, soaking at room temperature was taken out after 3 hours; Described carrier deionized water rinsing; Immerse and to begin dyeing in the dye liquor, by 2～4 ℃/min rising temperature, rise to 90～95 ℃ of insulation 20min, during per 8～10min jiggle once; Take out carrier, in dye liquor, add HAC, put into carrier, 90～95 ℃ of insulation 15～20min; Remove dye liquor, use deionized water rinsing; Be soaked in the deionized water, room temperature is more than 12 hours; Remove deionized water, washed 3～5 hours with cleansing solution; Remove cleansing solution, use deionized water rinsing; Take out washed 35～37 ℃ of oven dry of carrier.
Preferred as technique scheme, it is when making described carrier that described colouring is handled, and 0.1～5% dyestuff is sneaked in the raw material paint.
Further preferred as technique scheme, it is when making described carrier that described colouring is handled, and 0.1～0.5% dyestuff is sneaked in the raw material paint.
Preferred as technique scheme, described colouring is handled: be that to add massfraction in confining liquid be that 0.1～5% dyestuff seals carrier surface; Described biological detection reagent is biological substance bag not of the same race by in the carrier of carrier surface, the biological detection reagent through being assembled into.
Further preferred as technique scheme, described colouring is handled: be that to add massfraction in confining liquid be that 0.1～0.5% dyestuff seals the carrier surface sheet; Described biological detection reagent is biological substance bag not of the same race by in the carrier of carrier surface, the biological detection reagent through being assembled into.
The present invention has following beneficial effect:
The present invention can significantly reduce carrier background, raising detection sensitivity, other the present invention when biological respinse and also have low, the simple operation and other advantages of cost.
Description of drawings
Fig. 1 is not colouring and the bio-reaction system experimental image of having painted;
Fig. 2 is the analysis of results table of experimental image.
Embodiment
Below in conjunction with accompanying drawing the present invention is further explained.
Among Fig. 1, I1 and I 2 holes are negative sample; III 1 and III 2 holes are positive sample.Test figure specifically as shown in Figure 2, as can be seen from Figure 1, background colour is dusky comparatively fuzzy in the III 1, background colour is totally limpid in the III 2; Data from the table of Fig. 2 also as can be seen, the membrane carrier of handling through colouring has the advantage that improves signal to noise ratio (S/N ratio) and reduce the background signal value, and with the not contrast of colouring, difference is clearly.
Embodiment one
The preparation of genetic chip
(1) NC film dyeing:
1. the NC film is soaked in the 0.2% Tween-20 solution, soaking at room temperature was taken out after 3 hours; Film after soaking is immersed in the dyestuff, and 85 ℃ were dyed 1 hour, during every 5min jiggle once; 2. remove dye liquor, use deionized water rinsing 3 times;
3. will wash good film and be soaked in deionized water, room temperature is more than 12 hours;
4. remove deionized water, and with behind twice of the deionized water rinsing, add cleansing solution (0.2M PBS-T), washed 5 hours for 50 ℃;
5. remove cleansing solution, use deionized water rinsing 3 times;
6. add deionized water, 50 ℃, soaked 1 hour, and repeat again once;
7. take out washed film, behind deionized water rinsing one time, 37 ℃ of oven dry, standby.
(2) bag quilt: with different nucleic acid probes with certain requirement bag by in the nitrocellulose filter surface.
(3) assembling: will contain the nitrocellulose filter of biological substance after bag is done, and be assembled into genetic chip.
(4) sealing: the confining liquid with every hole 100ul seals biochip.
(5) drying: biochip has sealed the back and carried out drying in drying box.
(6) packing: pack after the biochip drying finishes.
Embodiment two
The preparation of immuno-chip (fluorescence method)
(1) bag quilt: with biological substance not of the same race with certain requirement bag by in the cellulose acetate membrane surface.
(2) will contain the cellulose acetate membrane of biological substance after the assembling bag is done, be assembled into immuno-chip.
(3) colouring liquid modulation: aniline blue (aniline blue) 0.5 gram is dissolved in the distilled water of 100ml, boils the back cooling, adds orange G 1 gram again, and acid fuchsin 1.5 grams are proofreaied and correct pH to 1.9 with HCC, can use.
(4) sealing: add 1.0% colouring liquid in the confining liquid, with the confining liquid of every hole 100ul immuno-chip is sealed.
(5) drying: immuno-chip has sealed the back and carried out drying in drying box.
(6) packing: pack after the immuno-chip drying finishes.
Embodiment three
The preparation of biochip
(1) bag quilt: with biological substance not of the same race with certain requirement bag by in the nitrocellulose filter surface.
(2) will contain the nitrocellulose filter of biological substance after the assembling bag is done, be assembled into biochip.
(3) sealing: add 0.4% methyl violet in the confining liquid, with the confining liquid of every hole 100ul biochip is sealed.
(4) drying: biochip has sealed the back and carried out drying in drying box.
(5) packing: pack after the biochip drying finishes.
Embodiment four
The preparation of biochip
(1) making of carrier:
1. nylon membrane is soaked in the 0.2% Tween-20 solution, soaking at room temperature was taken out after 3 hours; 2. the nylon membrane after soaking is soaked and use deionized water rinsing 3 times;
3. immerse in the dyestuff, 40 ℃ of beginning dyeing by 3 ℃/min rising temperature, rise to 95 ℃ of insulation 20min; Every 10min jiggles once during this time;
4. take out nylon membrane, in dye liquor, add HAC, put into nylon membrane, 95 ℃ of insulation 20min;
5. remove dye liquor, use deionized water rinsing 3 times;
6. will wash good film and be soaked in deionized water, room temperature is more than 12 hours;
7. remove deionized water, and with behind twice of the deionized water rinsing, add cleansing solution (0.2M PBS-T), washed 5 hours for 40 ℃;
8. remove cleansing solution, use deionized water rinsing 3 times;
9. add deionized water, 40 ℃, soaked 1 hour, and repeat again once;
10. take out washed film, behind deionized water rinsing one time, 37 ℃ of oven dry, standby.
(2) bag quilt: will biological substance not of the same race with certain requirement bag by the nylon membrane surface after dyeing.
(3) will contain the nylon membrane of biological substance after the assembling bag is done, be assembled into biochip.
(4) sealing: the confining liquid with every hole 100ul seals biochip.
(5) drying: biochip has sealed the back and carried out drying in drying box.
(6) packing: pack after the biochip drying finishes.
Embodiment five
The preparation of biological detection reagent
(1) making of carrier: under the constant prerequisite of the original technology of polystyrene batten, 0.2% black dyes sneaked in the polystyrene raw material paint, carry out lath production by original technology.
(2) bag quilt: envelope antigen is cushioned liquid preparation coating buffer with bag, and every hole is got 100 mL and is added ELISA Plate, builds the cover plate film, and 4 ℃ of bags are by 16-20h.
(3) wash plate: bag by after to carry out twice washing, 250 microlitres/hole, after washing finishes, should be at towel arsis trepang and liquid, and in time carry out next step sealing.
(4) sealing: confining liquid should shift to an earlier date taking-up in a hour tentatively rises again, and with before shaking up, every hole is got 180 mL and added ELISA Plate, builds the cover plate film, 37 ℃ of sealing 1.5h, and the spacing between the ELISA Plate should be consistent.
(5) oven dry: after having sealed, fall down liquid in the hole, and pat dry at towel, next take 37 ℃ of mode finish-drying ELISA Plate of being inverted oven dry or drying.
(6) pack: after the plank drying, the valve bag of should in time packing into is packed into according to the amount of 1 bag of drying agent of every block of plate, and the sack outside is finished writing plank and prepared the date, puts into this warehouse for finished product cold storage environment and preserves standby.
Claims (10)
1. method that reduces carrier background when biological respinse is characterized in that: before biological respinse, to the processing of painting of biological respinse carrier.
2. a kind of method that reduces carrier background when biological respinse according to claim 1 is characterized in that: described biological respinse carrier is the carrier that can be combined by biological substance with bag.
3. a kind of method that reduces carrier background when biological respinse according to claim 1 is characterized in that: described carrier is the supporting body that miillpore filter or tygon, polystyrene, Polyvinylchloride, polypropylene, silastic material are made; Described colouring is treated to carrier in manufacture process, adopts coloring matter to be scattered in the raw material of making described carrier, makes described coloring matter evenly be blended in the carrier through moulding process.
4. a kind of method that reduces carrier background when biological respinse according to claim 1 is characterized in that: described carrier is any one in the supporting body made of miillpore filter or tygon, polystyrene, Polyvinylchloride, polypropylene, silastic material; It is before making biological detection reagent that described colouring is handled, and adopts to contaminate or be coated with the technology of dying, and in dye liquor or be coated with and be covered with dye liquor, makes described carrier become coloured carrier described carrier impregnation.
5. a kind of method that reduces carrier background when biological respinse according to claim 1 is characterized in that: described carrier is any one in the supporting body made of miillpore filter or tygon, polystyrene, Polyvinylchloride, polypropylene, silastic material; It is in the process of making biological detection reagent that described colouring is handled, and adopts to contaminate or be coated with the technology of dying, and in dye liquor or be coated with and be covered with dye liquor, makes described carrier become coloured carrier described carrier impregnation.
6. according to claim 3 or 4 or 5 described a kind of methods that reduce carrier background when the biological respinse, it is characterized in that: described dye liquor is direct dyes, azoic dyes insoluble azo dyes, reactive dye, reducing dye, soluble vat dye, sulfur dye, sulphur vat dye, phthalocyanine dye, oxidation dye, the condensation dye, disperse dyes, acid dyes, acidic intermedium and acidly contains in mordant dye, alkalescence and the dye of positive ion, the coating any one.
7. a kind of method that reduces carrier background when biological respinse according to claim 4, it is characterized in that: described carrier is miillpore filter; Described colouring is handled: in the Tween-20 solution that described carrier is soaked in, take out after the soaking at room temperature earlier; Described carrier deionized water rinsing; Immerse and begin dyeing in the dye liquor, rising temperature and insulation; Take out carrier, in dye liquor, add HAC, put into carrier, soak; Remove dye liquor, use deionized water rinsing; Be soaked in the deionized water a period of time; Remove deionized water, wash with cleansing solution; Remove cleansing solution, use deionized water rinsing; Take out washed carrier oven dry.
8. a kind of method that reduces carrier background when biological respinse according to claim 3 is characterized in that: it is when making described carrier that described colouring is handled, and 0.1～5% dyestuff is sneaked in the raw material paint.
9. a kind of method that reduces carrier background when biological respinse according to claim 5, it is characterized in that: described carrier is miillpore filter; Described colouring is handled: earlier described carrier is soaked in 0.1～0.5% the Tween-20 solution, soaking at room temperature was taken out after 3 hours; Described carrier deionized water rinsing; Immerse and to begin dyeing in the dye liquor, by 2～4 ℃/min rising temperature, rise to 90～95 ℃ of insulation 20min, during per 8～10min jiggle once; Take out carrier, in dye liquor, add HAC, put into carrier, 90～95 ℃ of insulation 15～20min; Remove dye liquor, use deionized water rinsing; Be soaked in the deionized water, room temperature is more than 12 hours; Remove deionized water, washed 3～5 hours with cleansing solution; Remove cleansing solution, use deionized water rinsing; Take out washed 35～37 ℃ of oven dry of carrier.
10. a kind of method that reduces carrier background when biological respinse according to claim 6 is characterized in that: described colouring is handled: be that to add massfraction in confining liquid be that 0.1～5% dyestuff seals carrier surface; Described biological detection reagent be with biological substance bag not of the same race by behind carrier surface, the biological detection reagent through being assembled into.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310078156.7A CN103197055B (en) | 2013-03-12 | 2013-03-12 | Method for reducing background of vector in biological reaction |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310078156.7A CN103197055B (en) | 2013-03-12 | 2013-03-12 | Method for reducing background of vector in biological reaction |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103197055A true CN103197055A (en) | 2013-07-10 |
| CN103197055B CN103197055B (en) | 2015-04-15 |
Family
ID=48719746
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310078156.7A Expired - Fee Related CN103197055B (en) | 2013-03-12 | 2013-03-12 | Method for reducing background of vector in biological reaction |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103197055B (en) |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1082070A (en) * | 1992-07-15 | 1994-02-16 | 考脱沃兹Plc | Coloured film |
| CN1514244A (en) * | 2003-04-08 | 2004-07-21 | 成都夸常科技有限公司 | Method of preoceeding qualitative and/or quantitative analysis against target substance in sample and its detecting device |
| CN1771438A (en) * | 2003-04-10 | 2006-05-10 | 库诺公司 | Improved composite microarry slides |
| CN201096776Y (en) * | 2007-10-30 | 2008-08-06 | 江苏三联生物工程有限公司 | Biologic chip |
-
2013
- 2013-03-12 CN CN201310078156.7A patent/CN103197055B/en not_active Expired - Fee Related
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1082070A (en) * | 1992-07-15 | 1994-02-16 | 考脱沃兹Plc | Coloured film |
| CN1514244A (en) * | 2003-04-08 | 2004-07-21 | 成都夸常科技有限公司 | Method of preoceeding qualitative and/or quantitative analysis against target substance in sample and its detecting device |
| CN1771438A (en) * | 2003-04-10 | 2006-05-10 | 库诺公司 | Improved composite microarry slides |
| CN201096776Y (en) * | 2007-10-30 | 2008-08-06 | 江苏三联生物工程有限公司 | Biologic chip |
Non-Patent Citations (2)
| Title |
|---|
| COURTENAY KEMPER,ET AL: "An improved, luminescent europium-based stain for detection of electroblotted proteins on nitrocellulose or polyvinylidene difluoride membranes", 《ELECTROPHORESIS》 * |
| J.G.JONES AND B.M.SIMON: "An Investigation of Errors in Direct Counts of Aquatic Bacteria by Epifluorescence Microscopy, with Reference to a New Method for Dyeing Membrane Filters", 《J.APPL.BACT.》 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103197055B (en) | 2015-04-15 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103265947B (en) | A kind of indolepyridinium salt fluorescent probe for RNA in viable cell and kernel imaging | |
| CN105115878A (en) | Circulating tumor cell detection kit, preparing method thereof and application thereof | |
| Demuth et al. | Novel probes for pH and dissolved oxygen measurements in cultivations from millilitre to benchtop scale | |
| KR20200037755A (en) | Contact-type staining-assist patch, manufacturing method thereof and staining method using the patch | |
| CN1421700A (en) | Method and apparatus for measuring concentration of analyte | |
| ES2606716T3 (en) | Method to detect wound infection | |
| CN103275699B (en) | Pyrrole pyridine salt fluorescent probe used for RNA (ribonucleic acid) and nucleolus imaging in living cell | |
| CN109387626A (en) | For multiple diagnostic and alignment biochip | |
| CN108345498A (en) | A kind of dyeing scheduling system and dispatching method based on multitask staining protocols | |
| CN116445258A (en) | PCR tube and detection method thereof applied to target nucleic acid molecules | |
| CN109679836B (en) | Slide trace detection system applied to prawn pathogen high-flux detection | |
| US8524469B2 (en) | Two-step gram staining method | |
| CN103197055B (en) | Method for reducing background of vector in biological reaction | |
| CN110988344B (en) | Fluorescent staining reagent for rapidly identifying staphylococcus aureus and preparation method thereof | |
| CN108593392A (en) | A kind of vaginal fluid dyeing liquor and preparation method thereof | |
| Toda et al. | Continuous monitoring of ammonia removal activity and observation of morphology of microbial complexes in a microdevice | |
| CN109540633B (en) | Special preparation of high-resolution chromosome dye solution for karyotype analysis | |
| CN106323723A (en) | Double-blue staining method | |
| CN105861626A (en) | Antibacterial drug screening method based on hydrogel microarray chip | |
| CN108774325A (en) | The synthetic method of metal coordinating polymer and its application in the detection of diamines substance and equipment | |
| CN103981082A (en) | Cell amyloidosis detection chip and reagent thereof | |
| CN108329316A (en) | A kind of biological fluorescent labeling and preparation method thereof of detection nickel ion | |
| CN108444996B (en) | Rapid detection kit and detection method for protein residues | |
| CN207570882U (en) | Double-face dyeing device | |
| CN101539554A (en) | Quality control substance for identifying quality of medical cleaning liquid (agent) products |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20150415 Termination date: 20200312 |