CN101846678A - Rapid immunochromatographic reagent strip test method suitable for multiple samples - Google Patents
Rapid immunochromatographic reagent strip test method suitable for multiple samples Download PDFInfo
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- CN101846678A CN101846678A CN200910056992A CN200910056992A CN101846678A CN 101846678 A CN101846678 A CN 101846678A CN 200910056992 A CN200910056992 A CN 200910056992A CN 200910056992 A CN200910056992 A CN 200910056992A CN 101846678 A CN101846678 A CN 101846678A
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Abstract
The invention discloses a rapid immunochromatographic reagent strip test method suitable for multiple samples, comprising the following steps: 1) preparing a reagent strip composed of a sticky baseplate, a porous fibre material and a water absorption material, wherein, a reaction reagent corresponding to a detection item and a quality-controlled reagent corresponding to a color-developing agent are fixed on the porous fibre material; 2) mixing a sample to be tested with treating fluid to form the mixture of the tested sample; 3) mixing the reagent corresponding to the sample to be tested with the color-developing agent prepared by a conventional method to form the mixture of the color-developing agent; 4) dipping one end of the porous fibre material of the reagent strip into the liquid level of the mixture the tested sample; and 5) taking out the reagent strip when the sample climbs to the water absorption material, dipping one end of the porous fibre material into the liquid level of the mixture of the color-developing agent once again, and judging the result within an interpretation time slot. The method of the invention has the advantages of unnecessary glass fibrous membranes and dry color-developing agent technique, low cost, simple production process, wide sample application range, distinct judgment background, short interpretation time and improved sensitivity and the like.
Description
Technical field
The present invention relates to a kind of immune chromatography method, particularly relate to a kind of rapid immunochromatographireagent reagent strip test method glass fibre membrane and solid-state developer, that be applicable to several samples that do not adopt.
Background technology
Colloidal gold immunochromatographimethod (GICA) technology is a kind of simple and easy technology of setting up on the immunity percolation technical foundation early 1990s of immunology detection fast, is used for the mensuration of human chorionic gonadotrophin (HCG) at first by Beggs etc.GICA is to be carrier with nitrocellulose filter (NC film), utilizes the capillarity of microporous barrier, dropping is slowly oozed to the other end at the liquid of film bar one end move, as chromatography.Immunity Au composite dry plate sticks in nearly NC film bar lower end (c), film bar test section (t) is surrounded by specific antibody, when the test strips lower end enters in the liquid sample sample, lower end absorbent material imbitition moves to the upper end, when flowing through the c place, immune Au composite on the dry plate is redissolved, and drive it and ooze to the film bar and move, when if sample has specific antigen, can with the antibodies of immune Au composite, the gold of formation mark antigen-antibody complex flow to the test section, is caught by insolubilized antibody, form antibody-antigen-Jin labeling antibody compound, red control line appears in the t place on film.
Traditional immunochromatography reagent bar mainly is made up of base plate, nitrocellulose membrane, thieving paper and glass fibre membrane and solid-state developer, and owing to adopt solid-state developer, therefore need drying process, and for the certain requirement of storing of developer, as being stored in the good aluminium foil bag of sealing, promptly lose effect in case open to make moist.
And existing immunochromatography reagent bar is in the different samples of test, because the mode difference of sample preparation needs to adopt different reagent strips.As when testing whole blood sample, need on sample pad, add such as anti-RBC (red blood cell) antibody or some chemical substances and play aggegation or filter erythrocytic effect, in order to avoid the red blood cell in the whole blood sample causes false sample etc. to influence the phenomenon of test result in reaction or Quality Control zone colour developing.Therefore, traditional immunochromatography reagent bar method of testing has shortcomings such as cost height, the identical product sample scope of application be not wide.
Summary of the invention
The technical problem to be solved in the present invention provides a kind of rapid immunochromatographireagent reagent strip test method that is applicable to several samples, by adopting reagent strip and the liquid developer of forming by adhesive base, porous fibrous material and absorbent material, the present invention is compared with traditional method, have characteristics such as cost is low, the expansion of the sample scope of application, do not lose the sensitivity of traditional method simultaneously again.
For solving the problems of the technologies described above, a kind of rapid immunochromatographireagent reagent strip test method that is applicable to several samples of the present invention comprises step:
1) prepares reagent strip according to a conventional method
This reagent strip is made up of adhesive base, porous fibrous material and absorbent material, be provided with test section and Quality Control district successively by the chromatography direction on the porous fibrous material, be fixed with on the test section and the corresponding reaction reagent of detection, the Quality Control district is fixed with and the corresponding quality control reagent of developer;
2) testing sample is mixed with treating fluid, form the specimen potpourri, testing sample is 1: 1～1: 100 with the mixed volume for the treatment of fluid than scope, wherein, treating fluid is to contain the damping fluid that percent by volume is 0.1%～3% surfactant, and its damping fluid comprises Tris-HCl damping fluid, phosphate buffer, and the buffer concentration scope is at 0.001mol/L～1mol/L, pH of buffer value scope is 4.0～10.5, and surfactant comprises Tween-20, polyvinyl alcohol (PVA) and polyglycol;
3) prepare developer according to conventional method, and corresponding reagent is mixed with developer, form the developer potpourri, wherein, the concentration of the corresponding reagent that contains in the developer potpourri is 0.001mg/ml～100mg/ml;
4) specimen potpourri and developer potpourri are placed two micropore containers respectively;
5) porous fibrous material one end is dipped under the liquid level of specimen potpourri, liquid level position is no more than the position, test section that is fixed with reaction reagent on the reagent strip, and sample can climb to absorbent material one end by capillary action;
6) treat that sample rises to absorbent material, take out reagent strip, porous fibrous material one end is dipped under the developer potpourri liquid level again, liquid level position is no more than the position, test section that is fixed with reaction reagent on the reagent strip, the developer potpourri can climb to absorbent material one end by capillary action equally, in the interpretation time period, the interpretation test result.
Porous fibrous material in the described step 1) comprises nitrocellulose membrane, and its pore diameter range is at 0.02～15 μ m; Absorbent material comprises thieving paper; Reaction reagent is corresponding with detection and the reagent of idiosyncrasy can take place that comprise albumen, antigen or antibody, concentration range is 0.001mg/ml～100mg/ml; Quality control reagent is and the corresponding reagent of developer, comprises albumen, antigen or antibody, and concentration is 0.001mg/ml～100mg/ml.
Developer in the described step 3) is the developer of liquid condition, comprises collaurum, latex and fluorescence; Corresponding reagent is and the testing sample reagent corresponding, comprises albumen, antibody or antigen.
Judgement time segment limit in the described step 6) is 2～15 minutes.
In sum, the present invention compares with traditional reagent strip chromatography, has the following advantages:
1) need not glass fibre membrane, thereby save cost;
2) with the solid-state developer in the liquid developer replacement classic method, avoided drying process required in the classic method, production technology is more simple, control easily, and liquid developer has convenient the storage and (need not aluminium foil bag, the drying agent sealing is preserved), the long characteristics such as (more than 2 years) of the term of validity;
3) owing to contain surfactant component in the treating fluid, when handling sample, also can be so that the race sample process of sample is more smooth, and make that the final judgement background of test can be more clear;
4) owing to adopt the blend step of sample and treating fluid, it is wide that the scope of application of sample becomes, go for multiple specimen samples, as whole blood sample, the pedotheque of thickness etc., these samples itself just need mix with treating fluid, erythrocytic cracking and thickness dilution of sample are all finished in this step, and this step itself is the steps necessary that test is carried out, it also is a unified step, can't bring extra operation to the operator, understand easily and accept easy operating, thereby make multi-form sample adopt with a kind of reagent, accomplished with the method that a kind of operation is tested;
5) the interpretation time of test shortens, and sensitivity is constant, and the sensitivity of whole blood sample test increases;
6) the relative consumption of sample reduces.
Description of drawings
The present invention is further detailed explanation below in conjunction with accompanying drawing and embodiment:
Fig. 1 is a reagent strip structural drawing of the present invention;
Fig. 2 is the operational flowchart of immunochromatography reagent bar method of testing of the present invention;
Fig. 3 is the reagent strip structural drawing in the classic method;
Fig. 4 is the experimental result picture of the embodiment of the invention 1, a-classic method, b-method of the present invention;
Fig. 5 is the experimental result picture of the embodiment of the invention 2, a-classic method, b-method of the present invention.
Embodiment
Embodiment 1 classic method and method of the present invention detect HIV (AIDS virus) antibody
In the present embodiment, adhesive base, porous fibrous material and absorbent material that reagent strip in two kinds of methods adopts are identical, wherein, porous fibrous material is a nitrocellulose membrane, its cellulose nitrate membrane aperture is 15 μ m, absorbent material adopts thieving paper, and the reagent strip in the classic method also is stained with glass fibre membrane as application of sample part (structure as shown in Figure 3) at the nitrocellulose membrane other end.All routinely the method preparations of reagent strip in two kinds of methods, developer, the reagent strip width all is 10mm.
Wherein, fixing SPA of 1mg/ml (recombination staphylococcus staphylococcus albumin A) all in the Quality Control district, consumption 1 μ l/ reagent strip fixedly is wire.The fixing HIV antigen of 0.5mg/ml in the test section, consumption 1 μ l/ reagent strip fixedly is wire.Developer potpourri: adopt the blue latex composition (10mg/ml SPA: the volume ratio of blue latex composition is 1: 100) that combines with SPA.
The sample of two kinds of method employings is HIV positive serum sample (concentration is approximately 0.0001mg/ml).
The operation steps of classic method is:
1) on glass fibre membrane, adds 10 μ l testing samples;
2) behind testing sample, add 100 μ l developer potpourris;
3) sentence read result after 10 minutes.
The operation steps of method of the present invention is:
1) in 200 μ l micropore containers, mixes testing sample and treating fluid, its mixed volume ratio is 1: 20, and total amount is 105 μ l, and promptly the primary sample consumption is 5 μ l, wherein, treating fluid is that to contain 0.5mol/L, pH that percent by volume is 0.5% Tween-20 be 6.5 PBS damping fluid (phosphate buffer);
2) nitrocellulose membrane one end of reagent strip is placed the micropore container of step 1);
3) in another 200 μ l micropore container, add 100 μ l developer potpourris;
4) treat that sample mix liquid ran the Quality Control zone after, reagent strip is put into developer potpourri micropore container, sentence read result after 10 minutes.
The result as shown in Figure 4, two kinds of methods can both detect HIV antibody, but compare with classic method, method amount of samples of the present invention still less, and the test background more clear.
In the present embodiment, adhesive base, porous fibrous material and absorbent material that reagent strip in two kinds of methods adopts are identical, wherein, porous fibrous material is a nitrocellulose membrane, its cellulose nitrate membrane aperture is 5 μ m, absorbent material adopts thieving paper, but the reagent strip in the classic method also is stained with glass fibre membrane as application of sample part (structure as shown in Figure 3) at the nitrocellulose membrane other end.All routinely the method preparations of reagent strip in two kinds of methods, developer, the reagent strip width all is 5mm.
Wherein, fixing SPA of 1mg/ml (recombination staphylococcus staphylococcus albumin A) all in the Quality Control district, consumption 0.5 μ l/ reagent strip fixedly is wire.The fixing HIV antigen of 0.5mg/ml in the test section, consumption 0.5 μ l/ reagent strip fixedly is wire.Developer potpourri: adopt the collaurum preparation (1mg/ml SPA: the volume ratio of collaurum is 1: 500) that combines with SPA.
The sample of two kinds of method employings is normal person's whole blood sample.
The operation steps of classic method is:
1) on glass fibre membrane, adds 5 μ l testing samples;
2) behind testing sample, add 100 μ l developer potpourris;
3) sentence read result after 10 minutes.
The operation steps of method of the present invention is:
1) in 500 μ l micropore containers, mixes testing sample and treating fluid, its mixed volume ratio is 1: 50, and total amount is 255 μ l, and promptly the primary sample consumption is 5 μ l, wherein, treating fluid is that to contain 1mol/L, pH that percent by volume is 1.5% polyglycol be 9.5 Tris-HCl damping fluid;
2) nitrocellulose membrane one end of reagent strip is placed the micropore container of step 1);
3) in another 500 μ l micropore container, add 100 μ l developer potpourris;
4) treat that sample mix liquid ran the Quality Control zone after, reagent strip is put into developer potpourri micropore container, sentence read result after 10 minutes.
The result as shown in Figure 5, classic method is owing to can't handle the red blood cell in the whole blood, the whole background of result presents redness, can't sentence read result; And method display result of the present invention is correct, and promptly test zone does not have the p-wire appearance, and testing background is clear relatively.
Claims (9)
1. rapid immunochromatographireagent reagent strip test method that is applicable to several samples comprises step:
1) prepares reagent strip according to a conventional method
This reagent strip is made up of adhesive base, porous fibrous material and absorbent material, is provided with test section and Quality Control district successively by the chromatography direction on the porous fibrous material, is fixed with reaction reagent on the test section, and the Quality Control district is fixed with quality control reagent;
2) testing sample is mixed with treating fluid, form the specimen potpourri;
3) prepare developer according to a conventional method, and corresponding reagent is mixed with developer, form the developer potpourri;
4) specimen potpourri and developer potpourri are placed two micropore containers respectively;
5) reagent strip porous fibrous material one end is dipped under the liquid level of specimen potpourri, liquid level position is no more than the position, test section that is fixed with reaction reagent on the reagent strip, and sample climbs to absorbent material one end by capillary action;
6) treat that sample rises to absorbent material, take out reagent strip, test strips porous fibrous material one end is dipped under the developer potpourri liquid level again, and liquid level position is no more than the position, test section that is fixed with reaction reagent on the reagent strip, interpretation test result in the interpretation time period.
2. the rapid immunochromatographireagent reagent strip test method that is applicable to several samples as claimed in claim 1 is characterized in that: the porous fibrous material in the described step 1) comprises nitrocellulose membrane, and its pore diameter range is at 0.02～15 μ m; Absorbent material comprises thieving paper.
3. the rapid immunochromatographireagent reagent strip test method that is applicable to several samples as claimed in claim 1 is characterized in that: the reaction reagent in the described step 1) is corresponding with detection and the reagent of idiosyncrasy can take place, comprises albumen, antigen or antibody.
4. the rapid immunochromatographireagent reagent strip test method that is applicable to several samples as claimed in claim 1 is characterized in that: the quality control reagent in the described step 1) is and the corresponding reagent of developer, comprises albumen, antigen or antibody.
5. the rapid immunochromatographireagent reagent strip test method that is applicable to several samples as claimed in claim 1, it is characterized in that: the treating fluid described step 2) is to contain the damping fluid that percent by volume is 0.1%～3% surfactant, and testing sample is 1: 1～1: 100 with the mixed volume for the treatment of fluid than scope.
6. the rapid immunochromatographireagent reagent strip test method that is applicable to several samples as claimed in claim 5, it is characterized in that: described damping fluid comprises Tris-HCl damping fluid, phosphate buffer, the buffer concentration scope is at 0.001mol/L～1mol/L, and pH of buffer value scope is 4.0～10.5; Surfactant comprises Tween-20, polyvinyl alcohol (PVA) and polyglycol.
7. the rapid immunochromatographireagent reagent strip test method that is applicable to several samples as claimed in claim 1 is characterized in that: the developer in the described step 3) is the developer of liquid condition, comprises collaurum, latex and fluorescence.
8. the rapid immunochromatographireagent reagent strip test method that is applicable to several samples as claimed in claim 1, it is characterized in that: the corresponding reagent in the described step 3) is and the testing sample reagent corresponding, comprise albumen, antibody or antigen, wherein, the concentration of the corresponding reagent that contains in the developer potpourri is 0.001mg/ml～100mg/ml.
9. the rapid immunochromatographireagent reagent strip test method that is applicable to several samples as claimed in claim 1 is characterized in that: the judgement time segment limit in the described step 6) is 2～15 minutes.
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| CN200910056992A CN101846678A (en) | 2009-03-26 | 2009-03-26 | Rapid immunochromatographic reagent strip test method suitable for multiple samples |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102645532A (en) * | 2012-01-04 | 2012-08-22 | 深圳市易瑞生物技术有限公司 | Device and method for detecting beta-agonists drug |
| CN103048460A (en) * | 2012-12-15 | 2013-04-17 | 武汉珈源生物医学工程有限公司 | Method for detecting by using quantum dot fluorescence immunochromatographic test strips |
| CN103336122A (en) * | 2013-06-24 | 2013-10-02 | 中国科学院苏州生物医学工程技术研究所 | Immunofluorescence chromatography test strip for detecting platelet antibody and detection method of platelet antibody |
| CN106198966A (en) * | 2016-09-06 | 2016-12-07 | 中国农业大学 | A kind of near-infrared fluorescent immune chromatography reagent kit detecting aflatoxin and application thereof |
-
2009
- 2009-03-26 CN CN200910056992A patent/CN101846678A/en active Pending
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102645532A (en) * | 2012-01-04 | 2012-08-22 | 深圳市易瑞生物技术有限公司 | Device and method for detecting beta-agonists drug |
| CN103048460A (en) * | 2012-12-15 | 2013-04-17 | 武汉珈源生物医学工程有限公司 | Method for detecting by using quantum dot fluorescence immunochromatographic test strips |
| CN103336122A (en) * | 2013-06-24 | 2013-10-02 | 中国科学院苏州生物医学工程技术研究所 | Immunofluorescence chromatography test strip for detecting platelet antibody and detection method of platelet antibody |
| CN103336122B (en) * | 2013-06-24 | 2016-01-20 | 中国科学院苏州生物医学工程技术研究所 | The immunofluorescence chromatograph test strip that a kind of platelet antibody detects and detection method |
| CN106198966A (en) * | 2016-09-06 | 2016-12-07 | 中国农业大学 | A kind of near-infrared fluorescent immune chromatography reagent kit detecting aflatoxin and application thereof |
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Application publication date: 20100929 |