CN106645108A - Microporous plate chemiluminescence detection reagent and detection method - Google Patents
Microporous plate chemiluminescence detection reagent and detection method Download PDFInfo
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- CN106645108A CN106645108A CN201610953251.0A CN201610953251A CN106645108A CN 106645108 A CN106645108 A CN 106645108A CN 201610953251 A CN201610953251 A CN 201610953251A CN 106645108 A CN106645108 A CN 106645108A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/76—Chemiluminescence; Bioluminescence
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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
- G01N33/531—Production of immunochemical test materials
- G01N33/532—Production of labelled immunochemicals
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Abstract
Provided are a microporous plate chemiluminescence detection reagent and a detection method thereof. The microporous plate chemiluminescence detection reagent comprises a luminous substrate liquid A, luminous substrate liquid B and an enzyme conjugate diluent. The microporous plate chemiluminescence detection reagent further comprises 20-fold concentrated lotion, a negative control and a positive control. The detection method comprises the steps of using distilled water or purified water to dilute the 20-fold concentrated lotion by 20 times, and preparing working concentration lotion for standby application; taking a coating batten, forming two critical value control holes, one negative control hole, one positive control hole and a plurality of blood sample holes waiting for inspection in the coating batten; respectively adding 50 microliters of critical value control samples, 50 microliters of negative control samples, 50 microliters of positive control samples and 50 microliters of to-be-detected blood samples into the corresponding holes; respectively adding 50 microliters of nzyme conjugate diluent ino each hole, and then using a plate sealing film to seal the coating batten. The goal is to provide the microporous plate chemiluminescence detection reagent and the detection method thereof which are high in specificity and sensitivity, short in detection result obtaining time, simple and convenient in operation mode and accurate and reliable in detection result.
Description
Technical field
The present invention relates to a kind of kit and its method of testing for determining serum, sends out more particularly, to a kind of microwell plate chemistry
Light detection reagent and detection method.
Background technology
The sensitivity of existing microwell plate chemiluminescence detection reagent and detection method needs further to improve, and
The time that it obtains testing result is longer, and mode of operation is complex.
The content of the invention
It is an object of the invention to provide a kind of high specificity, sensitivity is high, and the time for obtaining testing result is short, mode of operation
Simplicity, testing result accurately and reliably microwell plate chemiluminescence detection reagent and its detection method.
The microwell plate chemiluminescence detection reagent of the present invention, it includes that luminous substrate liquid A, luminous substrate liquid B and enzyme are combined
Thing dilution；
The luminous substrate liquid A is made using following methods：
Prepare 5.8 grams of raw material disodium hydrogen phosphate, 0.59 gram of sodium dihydrogen phosphate dihydrate, 8.5 grams of sodium chloride, luminol 0.1
Gram, to 3 grams of iodophenol, 0.5 milliliter of Proclin300 diagnostic reagents preservative, 1 liter of purified water；
Each component is well mixed by more than, that is, obtain luminous substrate liquid A；
The luminous substrate liquid B is made using following methods：
Prepare 5.8 grams of raw material disodium hydrogen phosphate, 0.59 gram of sodium dihydrogen phosphate dihydrate, 8.5 grams of sodium chloride, luminol 0.1
Gram, 0.1 gram of carbamide peroxide, 0.5 milliliter of Proclin300 diagnostic reagents preservative, 1 liter of purified water；
Each component is well mixed by more than, that is, obtain luminous substrate liquid B；
The enzyme combination diluent is made using following methods：
Prepare 6.057 grams of raw material trishydroxymethylaminomethane, concentration is 37.5% 3.35 grams of concentrated hydrochloric acid, 10 grams of casein-sodium,
2.0 milliliters of Proclin300 diagnostic reagents preservative, 0.1 gram of amaranth, 100 milliliters of NBCS,；
Each component is well mixed with purified water and is settled to 1 liter by more than, that is, obtain enzyme combination diluent；
Also include 20 times of concentration washing lotions, negative control and positive controls,
20 times of concentrations washing lotion is containing polysorbas20（TWEEN-20）Phosphate buffer；
The negative control is the phosphate buffer containing human serum；
The positive control is the phosphate buffer of the positive samples of HBsAg containing hepatitis B surface antigen.
The detection method of the microwell plate chemiluminescence detection reagent of the present invention, it comprises the steps：
1）, with distilled water or purified water described 20 times concentration washing lotions are diluted into 20 times, be configured to working concentration washing lotion, it is stand-by；
2）, take coated slab, critical value control product 2 hole, negative control and each 1 hole of positive control, blood sample to be checked are laid thereon
Sample wells is multiple；
3）, respectively plus critical value control product, negative control, positive control, each 50 μ l of blood sample to be checked enter in corresponding hole；
4）, in each hole be separately added into the μ l of the enzyme combination diluent 50, then seal coated slab with shrouding film, then at
42 DEG C of incubated under agitation 15 minutes, or 37 DEG C of incubated under agitation 30 minutes；
5）, board-washing 5 times, the working concentration washing lotion then prepared in each Kong Zhongjia step 1 is no less than 350 μ l, then stands not
Less than 30 seconds, then pat dry；
6）, in each Kong Zhongjia luminous substrate liquid A50 μ l and luminous substrate liquid B50 μ l, vibration is mixed even, room temperature avoid light place
5-10 minutes, with RLU value of the chemical illumination immunity analysis instrument measurement per hole, that is, obtain result.
The detection method of the microwell plate chemiluminescence detection reagent of the present invention, wherein the luminous substrate liquid A, luminous substrate
Liquid B, enzyme combination diluent, 20 times of concentration washing lotions, negative control and positive controls using forward horizontal stand temperature to room temperature.
The detection method of the microwell plate chemiluminescence detection reagent of the present invention, wherein the temperature of the room temperature is 20 DEG C -24
℃ 。
The detection method of the microwell plate chemiluminescence detection reagent of the present invention, wherein the chemical illumination immunity analysis instrument is
BK-L96C type chemical illumination immunity analysis instruments.
The microwell plate chemiluminescence detection reagent and its detection method of the present invention, shows, it has spy by many experiments
Different in nature strong, sensitivity is high, and the time for obtaining testing result is short, and mode of operation is easy, the characteristics of testing result is extremely accurate reliable,
Accuracy rate in experiment is up to 100%, therefore, the microwell plate chemiluminescence detection reagent and its detection method of the present invention have prominent
The substantive distinguishing features for going out and significant progress.
The microwell plate chemiluminescence detection reagent and its detection method of the present invention are described in further detail below.
Specific embodiment
The microwell plate chemiluminescence detection reagent of the present invention, it includes that luminous substrate liquid A, luminous substrate liquid B and enzyme are combined
Thing dilution；
The luminous substrate liquid A is made using following methods：
Prepare 5.8 grams of raw material disodium hydrogen phosphate, 0.59 gram of sodium dihydrogen phosphate dihydrate, 8.5 grams of sodium chloride, luminol 0.1
Gram, to 3 grams of iodophenol, 0.5 milliliter of Proclin300 diagnostic reagents preservative, 1 liter of purified water；
Each component is well mixed by more than, that is, obtain luminous substrate liquid A；
The luminous substrate liquid B is made using following methods：
Prepare 5.8 grams of raw material disodium hydrogen phosphate, 0.59 gram of sodium dihydrogen phosphate dihydrate, 8.5 grams of sodium chloride, luminol 0.1
Gram, 0.1 gram of carbamide peroxide, 0.5 milliliter of Proclin300 diagnostic reagents preservative, 1 liter of purified water；
Each component is well mixed by more than, that is, obtain luminous substrate liquid B；
The enzyme combination diluent is made using following methods：
Prepare 6.057 grams of raw material trishydroxymethylaminomethane, concentration is 37.5% 3.35 grams of concentrated hydrochloric acid, 10 grams of casein-sodium,
2.0 milliliters of Proclin300 diagnostic reagents preservative, 0.1 gram of amaranth, 100 milliliters of NBCS,；
Each component is well mixed with purified water and is settled to 1 liter by more than, that is, obtain enzyme combination diluent；
Also include 20 times of concentration washing lotions, negative control and positive controls,
20 times of concentrations washing lotion is containing polysorbas20（TWEEN-20）Phosphate buffer；
The negative control is the phosphate buffer containing human serum；
The positive control is the phosphate buffer of the positive samples of HBsAg containing hepatitis B surface antigen.
2. the detection method of microwell plate chemiluminescence detection reagent as above, it comprises the steps：
1）, with distilled water or purified water described 20 times concentration washing lotions are diluted into 20 times, be configured to working concentration washing lotion, it is stand-by；
2）, take coated slab, critical value control product 2 hole, negative control and each 1 hole of positive control, blood sample to be checked are laid thereon
Sample wells is multiple；
3）, respectively plus critical value control product, negative control, positive control, each 50 μ l of blood sample to be checked enter in corresponding hole；
4）, in each hole be separately added into the μ l of the enzyme combination diluent 50, then seal coated slab with shrouding film, then at
42 DEG C of incubated under agitation 15 minutes, or 37 DEG C of incubated under agitation 30 minutes；
5）, board-washing 5 times, the working concentration washing lotion then prepared in each Kong Zhongjia step 1 is no less than 350 μ l, then stands not
Less than 30 seconds, then pat dry；
6）, in each Kong Zhongjia luminous substrate liquid A50 μ l and luminous substrate liquid B50 μ l, vibration is mixed even, room temperature avoid light place
5-10 minutes, with RLU value of the chemical illumination immunity analysis instrument measurement per hole, that is, obtain result.
As a further improvement on the present invention, above-mentioned luminous substrate liquid A, luminous substrate liquid B, enzyme combination diluent, 20
Times concentration washing lotion, negative control and positive control using forward horizontal stand temperature to room temperature.
The temperature of above-mentioned room temperature is 20 DEG C -24 DEG C.Above-mentioned chemical illumination immunity analysis instrument is BK-L96C type chemiluminescences
Immunity analysis instrument.
Claims (5)
1. microwell plate chemiluminescence detection reagent, it is characterised in that：It includes that luminous substrate liquid A, luminous substrate liquid B and enzyme are combined
Thing dilution；
The luminous substrate liquid A is made using following methods：
Prepare 5.8 grams of raw material disodium hydrogen phosphate, 0.59 gram of sodium dihydrogen phosphate dihydrate, 8.5 grams of sodium chloride, luminol 0.1
Gram, to 3 grams of iodophenol, 0.5 milliliter of Proclin300 diagnostic reagents preservative, 1 liter of purified water；
Each component is well mixed by more than, that is, obtain luminous substrate liquid A；
The luminous substrate liquid B is made using following methods：
Prepare 5.8 grams of raw material disodium hydrogen phosphate, 0.59 gram of sodium dihydrogen phosphate dihydrate, 8.5 grams of sodium chloride, luminol 0.1
Gram, 0.1 gram of carbamide peroxide, 0.5 milliliter of Proclin300 diagnostic reagents preservative, 1 liter of purified water；
Each component is well mixed by more than, that is, obtain luminous substrate liquid B；
The enzyme combination diluent is made using following methods：
Prepare 6.057 grams of raw material trishydroxymethylaminomethane, concentration is 37.5% 3.35 grams of concentrated hydrochloric acid, 10 grams of casein-sodium,
2.0 milliliters of Proclin300 diagnostic reagents preservative, 0.1 gram of amaranth, 100 milliliters of NBCS,；
Each component is well mixed with purified water and is settled to 1 liter by more than, that is, obtain enzyme combination diluent；
Also include 20 times of concentration washing lotions, negative control and positive controls,
20 times of concentrations washing lotion is containing polysorbas20（TWEEN-20）Phosphate buffer；
The negative control is the phosphate buffer containing human serum；
The positive control is the phosphate buffer of the positive samples of HBsAg containing hepatitis B surface antigen.
2. the detection method of microwell plate chemiluminescence detection reagent as claimed in claim 1, it is characterised in that：Including following step
Suddenly：
1）, with distilled water or purified water described 20 times concentration washing lotions are diluted into 20 times, be configured to working concentration washing lotion, it is stand-by；
2）, take coated slab, critical value control product 2 hole, negative control and each 1 hole of positive control, blood sample to be checked are laid thereon
Sample wells is multiple；
3）, respectively plus critical value control product, negative control, positive control, each 50 μ l of blood sample to be checked enter in corresponding hole；
4）, in each hole be separately added into the μ l of the enzyme combination diluent 50, then seal coated slab with shrouding film, then at
42 DEG C of incubated under agitation 15 minutes, or 37 DEG C of incubated under agitation 30 minutes；
5）, board-washing 5 times, the working concentration washing lotion then prepared in each Kong Zhongjia step 1 is no less than 350 μ l, then stands not
Less than 30 seconds, then pat dry；
6）, in each Kong Zhongjia luminous substrate liquid A50 μ l and luminous substrate liquid B50 μ l, vibration is mixed even, room temperature avoid light place
5-10 minutes, with RLU value of the chemical illumination immunity analysis instrument measurement per hole, that is, obtain result.
3. according to the detection method of the microwell plate chemiluminescence detection reagent described in claim 2, it is characterised in that：It is described luminous
Substrate solution A, luminous substrate liquid B, enzyme combination diluent, 20 times of concentration washing lotions, negative control and positive controls are using front flat
Weighing apparatus temperature is to room temperature.
4. according to the detection method of the microwell plate chemiluminescence detection reagent described in claim 3, it is characterised in that：The room temperature
Temperature be 20 DEG C -24 DEG C.
5. according to the detection method of the microwell plate chemiluminescence detection reagent described in claim 4, it is characterised in that：The chemistry
Luminescence immunoassay instrument is BK-L96C type chemical illumination immunity analysis instruments.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
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| CN201610953251.0A CN106645108A (en) | 2016-11-03 | 2016-11-03 | Microporous plate chemiluminescence detection reagent and detection method |
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| CN201610953251.0A CN106645108A (en) | 2016-11-03 | 2016-11-03 | Microporous plate chemiluminescence detection reagent and detection method |
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| CN201610953251.0A Pending CN106645108A (en) | 2016-11-03 | 2016-11-03 | Microporous plate chemiluminescence detection reagent and detection method |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108445230A (en) * | 2018-03-21 | 2018-08-24 | 北京科卫临床诊断试剂有限公司 | Procalcitonin chemiluminescence detection reagent based on nano antibody and detection method |
| CN110455785A (en) * | 2019-08-08 | 2019-11-15 | 郑州安图生物工程股份有限公司 | A kind of chemiluminescent substrate method of inspection |
| CN116223817A (en) * | 2023-05-04 | 2023-06-06 | 北京科卫临床诊断试剂有限公司 | Liver type fatty acid binding protein measurement system and method based on neural network |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108445230A (en) * | 2018-03-21 | 2018-08-24 | 北京科卫临床诊断试剂有限公司 | Procalcitonin chemiluminescence detection reagent based on nano antibody and detection method |
| CN110455785A (en) * | 2019-08-08 | 2019-11-15 | 郑州安图生物工程股份有限公司 | A kind of chemiluminescent substrate method of inspection |
| CN116223817A (en) * | 2023-05-04 | 2023-06-06 | 北京科卫临床诊断试剂有限公司 | Liver type fatty acid binding protein measurement system and method based on neural network |
| CN116223817B (en) * | 2023-05-04 | 2023-07-14 | 北京科卫临床诊断试剂有限公司 | Liver type fatty acid binding protein measurement system and method based on neural network |
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