CN105334318A - Portable C reaction protein detection kit - Google Patents
Portable C reaction protein detection kit Download PDFInfo
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- CN105334318A CN105334318A CN201510853865.7A CN201510853865A CN105334318A CN 105334318 A CN105334318 A CN 105334318A CN 201510853865 A CN201510853865 A CN 201510853865A CN 105334318 A CN105334318 A CN 105334318A
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- antibody
- reactive protein
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- cellulose membrane
- detection kit
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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
- G01N33/533—Production of labelled immunochemicals with fluorescent label
-
- 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
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/544—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being organic
Abstract
A portable C reaction protein detection kit comprises a test strip, an upper clamp slot, a lower clamp slot and a radio frequency identification chip; the test strip comprises a plastic cement plate, on which a sample pad, a first antibody bearing pad, a cellulose membrane and a water absorption pad are arranged in sequence; the first antibody bearing pad and the water absorption pad are respectively overlapped at two ends of the cellulose membrane; one end of the sample pad is overlapped on the first antibody bearing pad; the cellulose membrane is provided with a detection belt T for fixing a C reaction protein second antibody and a quality control belt for fixing a sheep anti-mouse antibody; the test strip is arranged in a cavity formed by splicing the upper clamp slot and the lower clamp slot; the upper clamp slot is provided with a sampling hole and a detection window; a sample diluent is a buffer solution containing a rheumatoid factor and heterophil antibody blocking agent; the lower clamp slot is provided with the radio frequency identification chip close to the left end. The kit has the advantages of being capable of accurately reflecting antibody activity, capable of avoiding wrong detection results, simple to use, and the like.
Description
Technical field
The invention belongs to biomedicine technical field, relate in particular to a kind of portable c reactive protein detection kit.
Background technology
In current quantitative immunological detection technique, mainly contain following several quantitative detecting method:
One is the immunoturbidimetry detection technique of supporting large-scale, highly integrated self-reacting device;
Two is enzyme linked immunological chromatography detection techniques;
Three is immunochromatography Fast Detection Technique;
Wherein there is the shortcomings such as sample needs pre-service, complicated operation, detection time long, the large-scale instrument that needs supporting complexity, maneuverability is poor, Turnaround Time is long in immunoturbidimetry detection technique and enzyme linked immunosorbent detection technology, be more suitable for the centralized detecting of sample in enormous quantities, be not suitable for the quick detection at emergency treatment and the outpatient service of each section office, community hospital, mini clinic, first aid scene etc.
And immunochromatography Fast Detection Technique is a kind of rapid immunoassay technology of growing up of developed country in recent years.Its principle is that sample liquid moves under capillary migration effect on nitrocellulose filter, on determinand wherein and film, the antibody (antigen) of certain area combines, by colored labels, within tens minutes, even macroscopic direct result can be obtained in a few minutes.Immunochromatography Fast Detection Technique does not need free antibody (antigen) to be separated with the antibody (antigen) forming compound, eliminate loaded down with trivial details washing step, thus simple operation, Turnaround Time is short, is widely used in places such as basic medical unit, emergency treatment, scene, family's self-inspections.
Be that the first generation immunochromatography technique of label has developed nearly more than 30 year with collaurum, still widespread use at present, but qualitative or semiquantitative detection can only be used for due to it, be difficult to meet the clinical requirement quantitatively detected trace, ultramicro-analysis thing.Be badly in need of a kind of New Product that can quantitatively detect fast and substitute colloid gold immune analytic product, meet clinical diagnosis demand.The fluorescence immunoassay that some diagnostic reagent enterprise developments can quantitatively detect detects reagent, but there is poor anti jamming capability, uses the shortcomings such as inconvenient.
Wherein China national intellecture property board web discloses a kind of immuno-chromatographic test paper strip of full-range C-reactive protein, its detection method is subject to the interference of rheumatoid factor and heterophil antibody, cause false positive, and use procedure needs to calibrate test strips, adds Operating Complexity.
China national intellecture property board web also discloses a kind of hs-CRP fluorescence immune chromatography method detection kit, comprise HS-CRP reaction plate, HS-CRP detects damping fluid, HS-CRPID chip, its use procedure is that HS-CRPID chip is inserted fluorescence detector reading ID chip data by user, then sample is added in HS-CRP detection damping fluid and react 1 minute, then added in HS-CRP reaction plate well by reacted HS-CRP detection damping fluid at once and react, reaction terminates rear insertion fluorescence detector and reads fluorescence signal.Its testing process need read ID chip data and clock reaction twice, and operating process is complicated, and ID chip is without encryption function, is easily replicated and distorts; And the method is subject to the interference of rheumatoid factor and heterophil antibody equally, causes false positive.
Summary of the invention
Problem to be solved by this invention is for the deficiencies in the prior art, provide a kind of simple to operate, not transreplication distort and be not vulnerable to the portable c reactive protein detection kit of the interference of rheumatoid factor and heterophil antibody,
In order to solve the problems of the technologies described above, the technical solution adopted in the present invention is: a kind of portable c reactive protein detection kit, comprises test strips, for the upper draw-in groove (upper shell) of fixing test strips and lower bayonet slot (lower house), radio-frequency (RF) identification chip; Described test strips comprises sample pad, the c reactive protein first antibody bearing pads of fluorescent tracing substance markers, cellulose membrane and adsorptive pads that plastics offset plate and plastics offset plate set gradually; Described first antibody bearing pads and adsorptive pads are overlapped in the two ends of cellulose membrane respectively; One end of described sample pad is overlapped on first antibody bearing pads; Described sample pad is through the process of anti-human erythrocyte antibody-solutions, the sample pad with the effect of absorption red blood cell; Described cellulose membrane has the detection zone T of fixation of C reactive protein second antibody and the quality control band C of fixing sheep anti-mouse antibody; Described test strips is placed in the cavity of draw-in groove and the mutual split formation of lower bayonet slot; The position described upper draw-in groove corresponding to sample pad is provided with the well (namely this well is for the target analytes calibration object after the dilution of interpolation sample diluting liquid or sample to be tested) of the target analytes calibration object after for the dilution of interpolation sample diluting liquid or sample to be tested, and cellulose membrane position is provided with detection window; Described sample diluting liquid is the damping fluid containing rheumatoid factor and heterophil antibody blocking agent; Described lower bayonet slot is provided with radio-frequency (RF) identification chip near left end.
Radio-frequency (RF) identification chip of the present invention is for storing the information such as corresponding relation accounting equation, test board date of manufacture, lot number, the term of validity, term of reference between test strips analyte concentration and fluorescence signal.
C reactive protein first antibody of the present invention be can the mouse monoclonal antibody of specific binding c reactive protein first epi-position (for the splenocyte that obtained by c reactive protein immune mouse and bone marrow cell are merged, screening obtains the hybridoma that forever can produce c reactive protein first epitope antibodies, and this Hybridoma Cell Culture separation and purification is obtained target mouse monoclonal antibody (procurement process of above-mentioned " mouse monoclonal antibody of energy specific binding c reactive protein first epi-position " is industry routine techniques).
C reactive protein second antibody of the present invention be can the mouse monoclonal antibody of specific binding c reactive protein second epi-position (for the splenocyte that obtained by c reactive protein immune mouse and bone marrow cell are merged, screening obtains the hybridoma that forever can produce c reactive protein second epitope antibodies, and this Hybridoma Cell Culture separation and purification is obtained target mouse monoclonal antibody; Be industry routine techniques).
Fluorescent tracing thing of the present invention be can under the exciting of certain wavelength excitation source the organic nano microballoon with functional group of generation fluorescence, further, preferably with the organic fluorescence microballoon of carboxy functional group.
Described sample pad is the one in filter paper, glass fibre, dacron, nonwoven fabrics.
Described cellulose membrane is nitrocellulose filter.
The c reactive protein first antibody bearing pads of described fluorescent tracing substance markers is the one in filter paper, glass fibre, dacron, nonwoven fabrics.
Described adsorptive pads is filter paper.
Blocking agent of the present invention is mouse thermal treatment IgG (mouse IgG (mouse IgG) is placed in 60-75 DEG C of water-bath heating and can obtains thermal treatment IgG in 10 minutes) or commercialization blocking agent compound (HBR etc. as MAB33, SCABTIBOD of Roche).
The preparation method of kit of the present invention is:
1) preparation of fluorescent tracing substance markers antibody (the c reactive protein first antibody of fluorescent tracing substance markers or fluorescence antibody)
With the phosphate buffer of 10mmol/L, pH=6.5 by carboxyl fluorescent microsphere (as Dongguan City Han Nuo Bioisystech Co., Ltd sells
yGCarboxylateMicrospheres and
one in MultifluorescentMicrospheres) to be diluted to concentration be 5mg/ml, be mixed with microsphere suspensions, EDAC (ethyldimethyl amine carbodiimide is added in the above-mentioned microsphere suspensions of every 10ml, CAS:25952-53-8) 1 ~ 10mg, dissolves mixing, and room temperature leaves standstill 1 hour, the centrifugal 20min of 10000rpm, remove supernatant, then precipitation is suspended in the phosphate buffer of isopyknic 10mmol/LpH=7.2, ultrasonic disperse; Then in carboxyl microsphere suspensions, c reactive protein first epi-position monoclonal antibody is added, antibody 1 ~ 5mg is added in the above-mentioned microsphere suspensions of every 10ml, mixing, room temperature reaction 2 hours, bovine serum albumin(BSA) is added again in above-mentioned microsphere suspensions, the concentration of bovine serum albumin(BSA) in microsphere suspensions is made to be 2 ~ 20mg/ml, mixing, room temperature reaction 1 hour; The centrifugal 20min of 10000rpm, removes supernatant, and precipitation is suspended in the phosphate buffer of isopyknic 10mmol/LpH=7.2, ultrasonic disperse, and 4 DEG C save backup;
2) preparation of first antibody bearing pads
The fluorescence antibody Spray dilutions dilution fluorescence antibody prepared by step (1), to 0.5 ~ 2mg/ml, fluorescence antibody is applied on fiberglass packing by three-dimensional specking platform, 37 DEG C of dryings 1 hour, for subsequent use;
3) cellulose membrane coated antibody
Nitrocellulose filter is pasted in plastics offset plate centre position, being diluted c reactive protein second epi-position monoclonal antibody and sheep anti-mouse antibody respectively to concentration with antibody bag by dilution is that 0.5 ~ 2.0mg/ml is prepared into spray film liquid, wherein, three-dimensional specking platform sprays film liquid with ribbon specking appropriate location (formed respectively and be fixed with the detection zone T of c reactive protein second epi-position monoclonal antibody and the quality control band C of fixing sheep anti-mouse antibody) on nitrocellulose filter, 37 DEG C of dryings 1 hour, for subsequent use.
4) pre-service of sample pad
The sample pad cut is placed in anti-human erythrocyte antibody-solutions to soak 5 minutes, drains after pulling out, be placed in 37 DEG C of dried overnight, for subsequent use;
5) assembling of fluorescent quantitation check-out console
Dried first antibody bearing pads, sample pad, thieving paper are cut on cutting machine the strip of appropriate size, by sample pad, first antibody bearing pads, cellulose membrane, the sticky note of adsorptive pads order on plastics offset plate, head and the tail are interconnected, be made into the large plate of semi-manufacture, then with cutting cutter, large for semi-manufacture plate is cut into the wide test strips of 4mm, loads onto, lower bayonet slot and radio-frequency (RF) identification chip (Shenzhen Zhong Kongmei core Science and Technology Ltd. production papery electronic tag).
6) foundation of calibration curve and the write of radio-frequency (RF) identification chip
Get each 100ul of target analytes calibration object of series concentration, being added drop-wise to check-out console sample adds in hand-hole, timing detects, and detects immediately after timing time terminates on fluorescence detector, reads the fluorescence signal intensity of fluorescence signal T (detection line) and C (nature controlling line); With calibration object concentration for ordinate, with T fluorescence signal and C fluorescence signal ratio for horizontal ordinate, carry out non-linear regression, can calibrating curve equation be obtained; Software (for the supporting configuration of commercially available radio-frequency (RF) identification chip) is read and write by the radio-frequency (RF) identification chip in the information write fluorescent quantitation check-out consoles such as calibrating curve equation, lot number, project name, the term of validity, term of reference, nature controlling line fluorescence signal effective value, Crypted password by radio-frequency (RF) identification chip.
Diluent ingredient described in step (2) is: the phosphate buffer of 10mmol/L, pH=7.2, the NaCl of the BSA of 2 ~ 10mg/ml, 5 ~ 37mg/ml, the Tween20 of 0 ~ 10mg/ml.
Antibody bag described in step (3) by diluent ingredient is: the phosphate buffer of 10mmol/L, pH=7.2, the Tween20 of the NaCl of 9mg/ml, 0 ~ 1mg/ml.
The anti-human erythrocyte antibody-solutions composition of step (4) is: the phosphate of 100mmol/L, pH7.6, the anti-human erythrocyte antibody of the tween20 of the BSA of 5 ~ 20mg/mL, 1 ~ 10mg/mL, 0.2 ~ 5mg/mL
A kind of fluorescent quantitation check-out console provided by the invention, the interference that blocking agent avoids heterophil antibody, rheumatoid factor is added in reagent, calibration parameter is write in radio-frequency (RF) identification chip in advance, improve ease-to-operate, the radio-frequency (RF) identification chip that the present invention uses can be encrypted, and avoids to be replicated and distorts.
Accompanying drawing explanation
Fig. 1 is reagent cartridge configuration figure of the present invention.
Fig. 2 is test strips structural drawing.
Fig. 3 is upper notch schematic diagram.
Fig. 4 is lower bayonet slot structural representation.
Fig. 5 is contrast and experiment figure.
Embodiment
With specific embodiment, the present invention is described in further details below, but the present invention is not only confined to following specific embodiment.
As shown in drawings, a kind of portable c reactive protein detection kit of the present invention, comprises test strips, for the upper draw-in groove 6 (upper shell) of fixing test strips and lower bayonet slot 7 (lower house), radio-frequency (RF) identification chip 8; Described test strips comprises sample pad 2, the c reactive protein first antibody bearing pads 3 of fluorescent tracing substance markers, cellulose membrane 4 and adsorptive pads 5 that plastics offset plate 1 and plastics offset plate set gradually; Described first antibody bearing pads 3 and adsorptive pads 5 are overlapped in the two ends of cellulose membrane 4 respectively; One end of described sample pad 2 is overlapped on first antibody carrying 3 pad; Described sample pad 2 is through the process of anti-human erythrocyte antibody-solutions, the sample pad with the effect of absorption red blood cell; Described cellulose membrane 3 has the detection zone T (T) of fixation of C reactive protein second antibody and the quality control band C (C) of fixing sheep anti-mouse antibody; Described test strips is placed in the cavity of draw-in groove and the mutual split formation of lower bayonet slot; The position described upper draw-in groove corresponding to sample pad is provided with the well 9 (namely this well is for the target analytes calibration object after the dilution of interpolation sample diluting liquid or sample to be tested) of the target analytes calibration object after for the dilution of interpolation sample diluting liquid or sample to be tested, and cellulose membrane position is provided with detection window 10; Described sample diluting liquid is the damping fluid containing rheumatoid factor and heterophil antibody blocking agent; Described lower bayonet slot is provided with radio-frequency (RF) identification chip 8 near left end.
The above-mentioned kit of the present invention comprises the sample diluting liquid for diluting target analytes calibration object or sample to be tested certainly.
The raw material that the embodiment of the present invention uses, does not do specified otherwise, is industry routine or commercially available prod.
The preparation of embodiment 1:C reactive protein detection kit
1) preparation of fluorescent tracing substance markers antibody
With the phosphate buffer of 10mmol/L, pH=6.5, carboxyl fluorescent microsphere being diluted to concentration is 5mg/ml, be mixed with microsphere suspensions, the EDAC of 5mg is added in every 10ml microsphere suspensions, mixing, room temperature leaves standstill 1 hour, and the centrifugal 20min of 10000rpm, removes supernatant (supernatant), precipitation is suspended in the phosphate buffer of isopyknic 10mmol/L, pH=7.2, ultrasonic disperse; Then c reactive protein monoclonal antibody 1 (the c reactive protein first epi-position monoclonal antibody of 3mg is added in the every 10ml carboxyl microsphere suspensions after ultrasonic disperse; For the splenocyte that obtained by c reactive protein immune mouse and bone marrow cell are merged, screening obtains the hybridoma that forever can produce c reactive protein first epitope antibodies, and this Hybridoma Cell Culture separation and purification is obtained target mouse monoclonal antibody, for industry routine techniques, as just described preparation process in detail in Baidupedia " monoclonal antibody ", therefore do not repeat them here:
Http:// baike.baidu.com/link? url=OiP74a66AYREMW0B0H-NP94LEwoLKNCV3YGV6jCwJ1WRhi-qxBI7 20GpIepbQ6MOgUlv7KP2mLCwsfk7_pxr2a), mixing, room temperature reaction 2 hours, adding bovine serum albumin(BSA) again makes the ultimate density of bovine serum albumin(BSA) be 20mg/ml, mixing, room temperature reaction 1 hour.The centrifugal 20min of 10000rpm, removes supernatant, and precipitation is suspended in the phosphate buffer of isopyknic 10mmol/L, pH=7.2, and ultrasonic disperse, 4 DEG C save backup.
2) preparation of first antibody bearing pads
With fluorescence antibody Spray dilutions dilution fluorescence antibody to debita spissitudo, fluorescence antibody is applied on fiberglass packing by three-dimensional specking platform, 37 DEG C of dry 1hr, for subsequent use.
3) cellulose membrane coated antibody
Debita spissitudo is diluted to by diluted second antibody and sheep anti-mouse antibody with antibody bag.Nitrocellulose filter is pasted in PVC plastic offset plate centre position, on three-dimensional specking platform by c reactive protein antibody 2 (c reactive protein second epi-position monoclonal antibody) and sheep anti-mouse antibody with ribbon specking appropriate location (formed respectively and be fixed with the detection zone T of c reactive protein second epi-position monoclonal antibody and the quality control band C of fixing sheep anti-mouse antibody) on nitrocellulose filter, 37 DEG C of dry 1hr, for subsequent use.
4) pre-service of sample pad
Preparation sample pad pretreatment fluid: the phosphate 100mmol/L of pH7.6, erythrocyte antibody (EA) 1g/L;
Nonwoven fabrics is cut into proper width, immerses in above-mentioned treating fluid and soak 5 minutes, then pull out and drain, 37 DEG C of dryings 2 hours, for subsequent use.
5) assembling of c reactive protein fluorescent quantitation check-out console
Dried first antibody bearing pads, sample pad, thieving paper (form absorption pad with) are cut on cutting machine the strip of appropriate size, by sample pad, first antibody bearing pads, cellulose membrane, the sticky note of adsorptive pads order on plastics offset plate, head and the tail are interconnected, be made into the large plate of semi-manufacture, then with cutting cutter, large for semi-manufacture plate is cut into the wide test strips of 4mm, outside test strips, fills upper and lower groove.
6) sample diluting liquid preparation
Phosphate 100mmol/L, the BSA10g/L of pH7.6, tween205g/L, thermal treatment mouse IgG1g/L;
7) foundation of calibration curve and the write of radio-frequency (RF) identification chip
Get the target analytes calibration object of series concentration, 100 times are diluted to sample diluting liquid, then drawing 100 μ l is added drop-wise in check-out console sample pipetting volume hole, timing detects, detect on fluorescence detector immediately after timing time terminates, read the fluorescence signal intensity of fluorescence signal T (detection line) and C (nature controlling line).With calibration object concentration for ordinate, carry out non-linear regression with T fluorescence signal and C fluorescence signal ratio for horizontal ordinate, can calibrating curve equation be obtained.
By radio-frequency (RF) identification chip read-write software by the information write radio-frequency (RF) identification chip such as calibrating curve equation, lot number, project name, the term of validity, term of reference, nature controlling line fluorescence signal effective value, Crypted password.
8) use of c reactive protein detection kit
Radio-frequency (RF) identification chip is inserted on fluorescence detector, by sample to be tested sample diluting liquid 100 times, then draw 100 μ l and be added drop-wise to check-out console sample and add in hand-hole, timing detects, detect on fluorescence detector immediately after timing time terminates, read testing result.
The Performance Evaluation of embodiment 2:C reactive protein detection kit
1) contrast experiment
Kit of the present invention and beckmanIMMAGE800 special protein instrument detect 243 parts of c reactive protein concentration ranges (whole blood/blood plasma) sample in 0.5-200mg/L scope simultaneously, as accompanying drawing 5, and the coefficient R 2=0.987 of both results.
2) interference experiment
Measure after adding the interfering material of different content respectively in a human serum sample.Add the measured value after interfering material and the difference between the measured value before adding interfering material is jamming rate divided by adding the measured value ratio before chaff interference, test findings show the human rheumatoid factor, human anti-mouse antibody concentration respectively when 1200IU/mL and below 6.5mg/L, they are to the jamming rate of measurement result all below 3% (see table 1).
Table 1
These results suggest that kit of the present invention can meet clinical requirement, by force anti-interference to the chaff interference such as heterophil antibody, rheumatoid factor, calibration parameter is write in radio-frequency (RF) identification chip in advance, improve ease-to-operate, the radio-frequency (RF) identification chip that the present invention uses can be encrypted, and avoids to be replicated and distorts.
Claims (10)
1. a portable c reactive protein detection kit, is characterized in that: comprise test strips, for the upper draw-in groove of fixing test strips and lower bayonet slot, radio-frequency (RF) identification chip; Described test strips comprises sample pad, the c reactive protein first antibody bearing pads of fluorescent tracing substance markers, cellulose membrane and adsorptive pads that plastics offset plate and plastics offset plate set gradually; Described first antibody bearing pads and adsorptive pads are overlapped in the two ends of cellulose membrane respectively; One end of described sample pad is overlapped on first antibody bearing pads; Described sample pad is through the process of anti-human erythrocyte antibody-solutions, the sample pad with the effect of absorption red blood cell; Described cellulose membrane has the detection zone T of fixation of C reactive protein second antibody and the quality control band C of fixing sheep anti-mouse antibody; Described test strips is placed in the cavity of draw-in groove and the mutual split formation of lower bayonet slot; The position described upper draw-in groove corresponding to sample pad is provided with the well of the target analytes calibration object after for the dilution of interpolation sample diluting liquid or sample to be tested, and cellulose membrane position is provided with detection window; Described sample diluting liquid is the damping fluid containing rheumatoid factor and heterophil antibody blocking agent; Described lower bayonet slot is provided with radio-frequency (RF) identification chip near left end.
2. portable c reactive protein detection kit according to claim 1, is characterized in that: described radio-frequency (RF) identification chip is for storing corresponding relation accounting equation between test strips analyte concentration and fluorescence signal, test board date of manufacture, lot number, the term of validity, term of reference information.
3. portable c reactive protein detection kit according to claim 1, is characterized in that: described c reactive protein first antibody is the mouse monoclonal antibody of energy specific binding c reactive protein first epi-position.
4. portable c reactive protein detection kit according to claim 1, is characterized in that: described c reactive protein second antibody is the mouse monoclonal antibody of energy specific binding c reactive protein second epi-position.
5. portable c reactive protein detection kit according to claim 1, is characterized in that: described fluorescent tracing thing be can under the exciting of certain wavelength excitation source the organic nano microballoon with functional group of generation fluorescence.
6. portable c reactive protein detection kit according to claim 1, is characterized in that: described sample pad is the one in filter paper, glass fibre, dacron, nonwoven fabrics; Described cellulose membrane is nitrocellulose filter; The c reactive protein first antibody bearing pads of described fluorescent tracing substance markers is the one in filter paper, glass fibre, dacron, nonwoven fabrics; Described adsorptive pads is filter paper.
7. portable c reactive protein detection kit according to claim 1, is characterized in that: described blocking agent is mouse thermal treatment IgG or commercialization blocking agent compound.
8. a preparation method for portable c reactive protein detection kit, is characterized in that: preparation process comprises:
1) preparation of the c reactive protein first antibody of fluorescent tracing substance markers
With the phosphate buffer of 10mmol/L, pH=6.5, carboxyl fluorescent microsphere being diluted to concentration is 5mg/ml, be mixed with microsphere suspensions, EDAC1 ~ 10mg is added in the above-mentioned microsphere suspensions of every 10ml, dissolve mixing, room temperature leaves standstill 1 hour, and the centrifugal 20min of 10000rpm, removes supernatant, then precipitation is suspended in the phosphate buffer of isopyknic 10mmol/LpH=7.2, ultrasonic disperse; Then in carboxyl microsphere suspensions, c reactive protein first epi-position monoclonal antibody is added, antibody 1 ~ 5mg is added in the above-mentioned microsphere suspensions of every 10ml, mixing, room temperature reaction 2 hours, bovine serum albumin(BSA) is added again in above-mentioned microsphere suspensions, the concentration of bovine serum albumin(BSA) in microsphere suspensions is made to be 2 ~ 20mg/ml, mixing, room temperature reaction 1 hour; The centrifugal 20min of 10000rpm, removes supernatant, and precipitation is suspended in the phosphate buffer of isopyknic 10mmol/LpH=7.2, ultrasonic disperse, and 4 DEG C save backup;
2) preparation of first antibody bearing pads
The c reactive protein first antibody spraying dilution fluorescence antibody of the fluorescent tracing substance markers prepared by step (1), to 0.5 ~ 2mg/ml, fluorescence antibody is applied on fiberglass packing by three-dimensional specking platform, 37 DEG C of dryings 1 hour, for subsequent use;
3) cellulose membrane coated antibody
Cellulose membrane is pasted in plastics offset plate centre position, being diluted c reactive protein second epi-position monoclonal antibody and sheep anti-mouse antibody respectively to concentration with antibody bag by dilution is that 0.5 ~ 2.0mg/ml is prepared into spray film liquid, wherein, three-dimensional specking platform sprays film liquid and forms the detection zone T of fixation of C reactive protein second epi-position monoclonal antibody and the detection zone C of fixing sheep anti-mouse antibody with ribbon specking appropriate location on nitrocellulose filter, 37 DEG C of dryings 1 hour, for subsequent use;
4) pre-service of sample pad
The sample pad cut is placed in anti-human erythrocyte antibody-solutions to soak 5 minutes, drains after pulling out, be placed in 37 DEG C of dried overnight, for subsequent use;
5) assembling of fluorescent quantitation check-out console
Dried first antibody bearing pads, sample pad, adsorptive pads are cut into strip on cutting machine, by sample pad, first antibody bearing pads, cellulose membrane, the sticky note of adsorptive pads order on plastics offset plate, head and the tail are interconnected, be made into the large plate of semi-manufacture, then with cutting cutter, large for semi-manufacture plate is cut into the wide test strips of 4mm, loads onto, lower bayonet slot and radio-frequency (RF) identification chip;
6) foundation of calibration curve and the write of radio-frequency (RF) identification chip
Get each 100ul of target analytes calibration object of series concentration, being added drop-wise to check-out console sample adds in hand-hole, timing detects, and detects immediately after timing time terminates on fluorescence detector, reads the fluorescence signal intensity of fluorescence signal T (detection line) and C (nature controlling line); With calibration object concentration for ordinate, with T fluorescence signal and C fluorescence signal ratio for horizontal ordinate, carry out non-linear regression, obtain calibrating curve equation; By radio-frequency (RF) identification chip read-write software by the radio-frequency (RF) identification chip in the information write fluorescent quantitation check-out consoles such as calibrating curve equation, lot number, project name, the term of validity, term of reference, nature controlling line fluorescence signal effective value, Crypted password.
9. the preparation method of portable c reactive protein detection kit according to claim 8, it is characterized in that: the diluent ingredient described in step (2) is: the phosphate buffer of 10mmol/L, pH=7.2, the NaCl of the BSA of 2 ~ 10mg/ml, 5 ~ 37mg/ml, the Tween20 of 0 ~ 10mg/ml.
10. the preparation method of portable c reactive protein detection kit according to claim 8, it is characterized in that: the antibody bag described in step (3) by diluent ingredient is: the phosphate buffer of 10mmol/L, pH=7.2, the Tween20 of the NaCl of 9mg/ml, 0 ~ 1mg/ml; The composition of the anti-human erythrocyte antibody-solutions described in step (4) is: the phosphate 100mmol/L of pH7.6, erythrocyte antibody (EA) 1g/L.
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| CN106442966A (en) * | 2016-08-31 | 2017-02-22 | 中山市创艺生化工程有限公司 | C-reactive protein immunofluorescence quantitative test strip and preparation method thereof |
| CN106645681A (en) * | 2016-10-31 | 2017-05-10 | 广州科方生物技术股份有限公司 | Blocking agent kit for immunochromatography determination and using method thereof |
| CN107621539A (en) * | 2016-07-13 | 2018-01-23 | 艾博生物医药(杭州)有限公司 | A kind of method of analyte in detection means and detection liquid sample |
| CN108548926A (en) * | 2018-03-22 | 2018-09-18 | 北京九强生物技术股份有限公司 | A kind of creatine kinase isozyme detection kit |
| CN108918883A (en) * | 2018-05-18 | 2018-11-30 | 瑞莱生物科技江苏有限公司 | A kind of immunofluorescent reagent box of rapid quantitative detection cardiac muscle troponin I, creatine kinase isozyme and myoglobins |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101620226A (en) * | 2009-08-07 | 2010-01-06 | 四川省迈克科技有限责任公司 | Quick test kit of schistosomiasis and preparation method thereof |
| WO2010026758A1 (en) * | 2008-09-05 | 2010-03-11 | 積水メディカル株式会社 | Monoclonal antibody, and immunoassay using same |
| WO2012094625A2 (en) * | 2011-01-08 | 2012-07-12 | Access Medical Systems, Ltd. | Systems for immunoassay tests |
| CN103630683A (en) * | 2013-11-13 | 2014-03-12 | 成都领御生物技术有限公司 | Test strip card |
| CN105021595A (en) * | 2015-07-23 | 2015-11-04 | 杭州金溪生物技术有限公司 | Mobile surveillance car-based single particle volatile organic compound online mass spectrum detection system and method |
| CN105092861A (en) * | 2015-09-14 | 2015-11-25 | 广州市微米生物科技有限公司 | Immunofluorescence chromatography test paper for CRP (C-reaction protein)/SAA (Serum amyloid A protein) quantitative combined detection and preparation method of immunofluorescence chromatography test paper |
-
2015
- 2015-11-28 CN CN201510853865.7A patent/CN105334318A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2010026758A1 (en) * | 2008-09-05 | 2010-03-11 | 積水メディカル株式会社 | Monoclonal antibody, and immunoassay using same |
| CN101620226A (en) * | 2009-08-07 | 2010-01-06 | 四川省迈克科技有限责任公司 | Quick test kit of schistosomiasis and preparation method thereof |
| WO2012094625A2 (en) * | 2011-01-08 | 2012-07-12 | Access Medical Systems, Ltd. | Systems for immunoassay tests |
| CN103630683A (en) * | 2013-11-13 | 2014-03-12 | 成都领御生物技术有限公司 | Test strip card |
| CN105021595A (en) * | 2015-07-23 | 2015-11-04 | 杭州金溪生物技术有限公司 | Mobile surveillance car-based single particle volatile organic compound online mass spectrum detection system and method |
| CN105092861A (en) * | 2015-09-14 | 2015-11-25 | 广州市微米生物科技有限公司 | Immunofluorescence chromatography test paper for CRP (C-reaction protein)/SAA (Serum amyloid A protein) quantitative combined detection and preparation method of immunofluorescence chromatography test paper |
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| CN106442966A (en) * | 2016-08-31 | 2017-02-22 | 中山市创艺生化工程有限公司 | C-reactive protein immunofluorescence quantitative test strip and preparation method thereof |
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| CN106645681A (en) * | 2016-10-31 | 2017-05-10 | 广州科方生物技术股份有限公司 | Blocking agent kit for immunochromatography determination and using method thereof |
| CN106645681B (en) * | 2016-10-31 | 2018-12-18 | 广州科方生物技术股份有限公司 | A kind of blocking agent kit and its application method for immunochromatographic measurement |
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