CN1077538A - Utilize sub-micron latex to carry out the method for enzyme immune reaction - Google Patents
Utilize sub-micron latex to carry out the method for enzyme immune reaction Download PDFInfo
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- CN1077538A CN1077538A CN 93104249 CN93104249A CN1077538A CN 1077538 A CN1077538 A CN 1077538A CN 93104249 CN93104249 CN 93104249 CN 93104249 A CN93104249 A CN 93104249A CN 1077538 A CN1077538 A CN 1077538A
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- latex
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- micron
- micron latex
- immune reaction
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- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Enzymes And Modification Thereof (AREA)
- Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
Abstract
A kind of method of utilizing sub-micron latex to carry out enzyme immune reaction, comprise the modification of sub-micron latex preparation, sub-micron latex and the application in enzyme immune reaction thereof, used sub-micron latex is through ferrous iron and the magnetized magnetic latex of ferric iron, this latex can separate by magnetic separator, also can be suspended in fully in the reactant liquor simultaneously, institute is so that reaction velocity is accelerated, and operating performance is simplified.
Description
The present invention relates to a kind of method of enzyme immune reaction, especially utilize sub-micron latex to carry out the method for above-mentioned reaction.
The enzyme immune reaction technology has begun to be widely used in food hygiene, biomedical Biological Detection.Method at present commonly used is for being fixed in reaction reagent such as antigen or antibody on the solid phase carrier, and then detecting material to be checked in the solution with the solid phase carrier of combination, this liquid-solid phase hybridization reaction reaches the balance required time, and to reach the balance required time than liquid-liquid phase hybridization long doubly a lot of.Enzyme linked immunoassay commonly used at present adsorbs antigen or antibody on the enzyme immune reaction plate of being everlasting, and makes the quantitative changeization of antigen or antibodies bigger like this, and the absorption required time is long, often need spend the night, and therefore operation and agents useful for same etc. is required relatively stricter.
The object of the present invention is to provide a kind of method of utilizing sub-micron latex to carry out enzyme immune reaction, because sub-micron latex has been carried out the magnetization processing, above-mentioned defective can be overcome, and the reaction time is shortened dramatically in this method.
Method of the present invention comprises the preparation of sub-micron latex, the modification of sub-micron latex and the application in enzyme immune reaction thereof, and described sub-micron latex is the magnetic sub-micron latex of handling through magnetization.
The present invention has following advantage: magnetic latex can separate by magnetic separator, can be suspended in fully in the reactant liquor again simultaneously, and institute is so that reaction velocity is accelerated simplified control; Also can realize operation automation simultaneously by combining with porous enzyme immune reaction detector; Magnetic latex is used for nucleic acid hybridization reaction, the conversion of bioactivator and preparation etc. with after nucleic acid probe, enzyme, antibody or aglucon combine.
Below in conjunction with example method of the present invention is done more detailed explanation.
One, the preparation of magnetic sub-micron latex: 100 milliliters of ferrous irons that 100 milliliters in polystyrene sub-micron latex are suspended in process micropore (0.21 μ m) membrane filtration, ferric iron (iron protochloride 10.8 grams, iron sulfate 5.6 grams, mole ratio is about 2: 1) aqueous solution in, at room temperature stir 4-6 hour (realizing the balance of latex and ferric ion) with magnetic stirring apparatus, slowly drip the NaOH of 100-200 milliliter 5% or other alkaline solution, accelerate the speed of magnetic stirring apparatus simultaneously, after stirring 1-2 hour under the room temperature, rinsing is colourless until eluate repeatedly with distilled water or damping fluid and magnetic separator, till PH is the PH of distilled water or damping fluid.Magnetic sub-micron latex is in 4 ℃ of storages.
Two, the modification of magnetic sub-micron latex: 100 milliliters of magnetic sub-micron latex are washed with 2 premium on currency and 1 liter of anhydrous dioxane, and after making its dehydration, be suspended in 200 milliliters the dioxane, add N-hydroxyl amber acid imide, the polystyrene N-hydroxy-succinamide ester of dioxane rinsing gained is used in room temperature jolting 90 minutes then.This polystyrene N-hydroxy-succinamide ester can be preserved in dioxane, after also dioxane can being drained, with antigen, after antibody or the nucleic acid probe combination, carry out the sandwich reaction of enzyme linked immunoassay or nucleic acid, also can combine, carry out the conversion of material or the separation of bioactivator, promptly can be used as the zymophore of immobilized enzyme reaction with enzyme or part.
Three, the application of magnetic sub-micron polystyrene N-hydroxy-succinamide ester in enzyme linked immunoassay:
(1) combining of magnetic sub-micron polystyrene N-hydroxy-succinamide ester and antigen that contains the terminal amino group group or antibody:
Magnetic sub-micron polystyrene N-hydroxy-succinamide ester after dioxane drained join with the isopyknic solution that contains antigen or antibody of latex in, the solution of dissolving antigen or antibody is generally used citric acid, phosphoric acid, the damping fluid of acetate or supercarbonate, PH is 5-8.5,4 ℃ of reactions, reaction time can be from 10 minutes to 6 hours, after finishing, reaction adds the 1M glycocoll, PH9.0, room temperature reaction 2 hours, after removing unreacted Acibenzolar group, after magnetic latex fully washs with damping fluid again, or freeze-drying preserves, or 4 ℃ are stored in the damping fluid that contains Sodium azide
(2) operate in below and carry out (enzyme immune reaction) on the enzyme immune reaction plate
Add the above-mentioned magnetic of 20-100 μ l latex/hole
↓
Add 37 ℃ of materials to be checked of 20-100 μ l 15-30 minute (the immune response plate can be vibrated in the horizontal direction)
↓
Washing 3 minutes X3 time (behind magnetic separator sedimentation magnetic latex, topples over and blots cleansing solution with filter paper.Cleansing solution is phosphate buffer or physiological saline)
↓
Added 37 ℃ of 50-100 μ l enzyme mark bonds 15-30 minute
↓
Wash 3 minutes X3 time
↓
The substrate solution 200 μ l that add new preparation
↓
Room temperature 30-60 minute or 37 ℃ 5-30 minute
↓
Cessation reaction, colorimetric detection reaction result or range estimation reaction result
Utilize this magnetic latex can make to react in 1.5-2 hour and finish, fast doubly a lot of than commonsense method 28-36 hour.
Claims (5)
1, a kind of method of utilizing sub-micron latex to carry out enzyme immune reaction, comprise the preparation of sub-micron latex, the modification of sub-micron latex and the application in enzyme immune reaction thereof is characterized in that described sub-micron latex is the magnetic sub-micron latex of handling through magnetization.
2, the method for claim 1 is characterized in that magnetic sub-micron latex is suspended in polystyrene latex in ferrous iron and the ferric aqueous solution to be prepared from.
3, method as claimed in claim 2 is characterized in that described ferrous iron is an iron protochloride, and ferric iron is an iron sulfate.
4, the method for claim 1, the modification that it is characterized in that magnetic sub-micron latex are by latex being suspended in the dioxane, adding N-hydroxy-succinamide to generate magnetic sub-micron polystyrene N-hydroxy-succinamide ester.
5, the method for claim 1 is characterized in that carrying out enzyme immune reaction with after the magnetic sub-micron latex of modified and antigen that contains the terminal amino group group or the antibodies.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 93104249 CN1077538A (en) | 1993-04-12 | 1993-04-12 | Utilize sub-micron latex to carry out the method for enzyme immune reaction |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 93104249 CN1077538A (en) | 1993-04-12 | 1993-04-12 | Utilize sub-micron latex to carry out the method for enzyme immune reaction |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN1077538A true CN1077538A (en) | 1993-10-20 |
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Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 93104249 Pending CN1077538A (en) | 1993-04-12 | 1993-04-12 | Utilize sub-micron latex to carry out the method for enzyme immune reaction |
Country Status (1)
| Country | Link |
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| CN (1) | CN1077538A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100389327C (en) * | 2004-06-22 | 2008-05-21 | 北京倍爱康生物技术股份有限公司 | Magnetic separation enzyme-linked immune detecting method for thyroid gland peroxidase antibody |
| CN102301238A (en) * | 2009-01-29 | 2011-12-28 | 株式会社日立高新技术 | Automatic Analyzer |
-
1993
- 1993-04-12 CN CN 93104249 patent/CN1077538A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100389327C (en) * | 2004-06-22 | 2008-05-21 | 北京倍爱康生物技术股份有限公司 | Magnetic separation enzyme-linked immune detecting method for thyroid gland peroxidase antibody |
| CN102301238A (en) * | 2009-01-29 | 2011-12-28 | 株式会社日立高新技术 | Automatic Analyzer |
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Legal Events
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| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C06 | Publication | ||
| PB01 | Publication | ||
| C01 | Deemed withdrawal of patent application (patent law 1993) | ||
| WD01 | Invention patent application deemed withdrawn after publication |