CN102590524A - Assay kit for neutrophil gelatinase-associated lipocalin - Google Patents
Assay kit for neutrophil gelatinase-associated lipocalin Download PDFInfo
- Publication number
- CN102590524A CN102590524A CN2011104549917A CN201110454991A CN102590524A CN 102590524 A CN102590524 A CN 102590524A CN 2011104549917 A CN2011104549917 A CN 2011104549917A CN 201110454991 A CN201110454991 A CN 201110454991A CN 102590524 A CN102590524 A CN 102590524A
- Authority
- CN
- China
- Prior art keywords
- ngal
- antibody
- reagent
- hepes
- concentration
- 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
Classifications
-
- 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
- G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
- G01N33/54346—Nanoparticles
Landscapes
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Hematology (AREA)
- Urology & Nephrology (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Microbiology (AREA)
- Analytical Chemistry (AREA)
- Biotechnology (AREA)
- Nanotechnology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Cell Biology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention relates to an assay kit for neutrophil gelatinase-associated lipocalin (NGAL), in particular to an assay kit for detecting the concentration of the NGAL in human body fluids and a preparation method for the assay kit. The kit comprises a reagent R1, a reagent R2 and a working calibrator, wherein the reagent R1 is a buffer solution which is suitable for antigen-antibody reaction; and the reagent R2 is a solution simultaneously containing nanospheres which have two particle sizes and are combined with an anti-human NGAL antibody, and ensures that the nanospheres are coagulated to cause the change of the turbidity through antigen-antibody specific reaction, and the NGAL can be quantificationally determined by contrasting calibration curves. The kit has relatively high sensitivity when determining low NGAL concentration and has a relatively wide linear range when determining high NGAL concentration, so a prozone phenomenon is avoided. The kit has stable physical and chemical properties, is convenient to use, and has a broad application prospect in clinical laboratory science.
Description
Technical field
The application relates to a kind of neutrophil gelatinase-associated lipocalin and measures kit; More specifically, the application relates to a kind of a kind of detectable that is used for human body liquid neutrophil gelatinase-associated lipocalin (NGAL) concentration and preparation method thereof, and this reagent comprises reagent R1, reagent R2 and work calibration object; Reagent R1 is for being fit to the buffer solution of antigen-antibody reaction; R2 is the solution that contains the Nano microsphere that is combined with anti-people NGAL antibody of two kinds of particle diameters of size simultaneously, and this reagent condenses Nano microsphere through the specific reaction of antigen-antibody; Produce the variation of turbidity, the comparison calibration curve can quantitative measurement NGAL.
Background technology
Neutrophil gelatinase-associated lipocalin (neutrophil gelatinase-associated lipocalin; NGAL); Being also referred to as lipocalin protein-2 (Lipocalin-2), is a newcomer of lipocalin protein family, and it is the early stage biomarker of renal dysfunction.NGAL not only is present in the neutrophil cell; Also can appear in the specific epithelial cell; For example take place in the process of ischemic and toxicity injury of kidney, the NGAL in the renal cells will significantly increase, in two hours of beginning; The NGAL level will significantly increase in urine and the blood, so NGAL is the responsive mark of early stage acute injury of kidney.
Behind the clinical operation, acute renal failure to a certain degree can appear in the patient more than 50%, and the later stage of falling ill usually occurs making that mortality ratio increases greatly.The prognosis of acute renal failure never improves, and in from now on 40 years, will cause remarkable concern.General diagnostic method all can only detect after renal function is degenerated like the method for measuring serum creatinine or cysteine proteinase inhibitor C (Cystatin C), promptly can only damage one day after very the person detect in several days.And using the NGAL detection kit, the doctor can make diagnosis and do proper process acute renal dysfunction in several hours in damage.
Research shows that NGAL content detection distribution of results is between 0.7-9.6ng/ml in healthy volunteer's urine, and mean value is 5.3ng/ml, and the content detection result in the blood plasma is 37-106ng/ml, and mean value is 63ng/ml.Behind injury of kidney, the NGAL level will increase rapidly suddenly.With the machine testing severe case, the concentration of urine NGAL is that 110ng/ml does not wait to 40000ng/ml, and their EDTA anticoagulate plasma testing result is that 25ng/ml is to 3491ng/ml.NGAL content in urine surpasses 350ng/ml, and perhaps to reach the positive predictive value of 400ng/ml generation renal failure when above be 90% to the NGAL content in the blood plasma.
The assay method of NGAL mainly contains ELISA, radioimmunology, Western-blotting and chemoluminescence method at present.The ELISA automaticity is not high, and it is bigger influenced by human factor; Radioimmunology exists problem of environmental pollution; The Western-blotting complicated operation, it is low to measure precision; Though and chemoluminescence method sensitivity is high, the mensuration range of linearity is less and the detection cost is higher, needs particular chemical luminous detection appearance, and these reasons cause its range of application less.
The latex enhancing immune turbidimetry also is a common method of measuring NGAL concentration in human plasma or the urine, and this method is less demanding to instrument and equipment, does not have problems such as environmental protection and operating personnel's self-protection.With other assay methods relatively, this method is easy fast, sensitive reliable, common automatically or semi-automatic biochemical analyzer just can, range of application is greatly arranged, bigger practical value.
NGAL concentration is lower than 10ng/ml in normal person's urine, in severe nephrotic urine concentration can be up to 7000ng/ml more than.So broad concentration range has all proposed higher requirement to the detection kit sensitivity and the range of linearity.Existing in the market latex enhancing immune turbidimetry is measured the kit of NGAL concentration in human plasma or the urine; But these kits can't satisfy the wide range of linearity and highly sensitive requirement simultaneously; Therefore need diluted sample could measure high concentration NGAL, this has brought inconvenience in the use.
Therefore, be necessary to develop a kind of turbid detection kit of NGAL latex enhancing immune transmittance that has the high sensitivity and the wide range of linearity simultaneously.
Summary of the invention
Therefore, technical purpose of the present invention is to develop a kind of NGAL latex enhancing immune projection that has the high sensitivity and the wide range of linearity simultaneously than turbid detection kit.
Therefore, first aspect of the present invention relates to a kind of NGAL detection kit, comprises reagent R1, reagent R2 and work calibration object, and wherein reagent R1 is NGAL and anti-people NGAL antibody response damping fluid; Reagent R2 is the Nano microsphere solution that is combined with anti-people NGAL antibody; The work calibration object is made up of damping fluid, stabilizing agent, antiseptic and NGAL normative reference article; It is characterized in that, comprise the Nano microsphere that is combined with anti-people NGAL antibody of two kinds of particle diameters of size among the said reagent R2, big particle diameter is 150-500nm; Small particle diameter is 50-100nm; Preferably, big particle diameter is 200-300nm, and small particle diameter is 70-90nm.
Preferably, reagent R1 damping fluid is selected from one or more in phosphate buffer, glycine buffer, HEPES damping fluid, borate buffer, acetate buffer or the ammonium chloride buffer, and its concentration is 10-500mM; The used damping fluid of reagent R2 is one or more in phosphate buffer, glycine buffer, HEPES damping fluid, borate buffer, acetate buffer or the ammonium chloride buffer, and its concentration is 10-500mM.
Preferably, described anti-people NGAL antibody is polyclonal antibody or monoclonal antibody, and more preferably, described polyclonal antibody is a kind of of the anti-people NGAL of rabbit antibody, goat-anti people NGAL antibody or the anti-people NGAL of chicken antibody; Monoclonal antibody is a mouse-anti people NGAL antibody.
Preferably, the Nano microsphere among the reagent R2 is one or more latex that are polymerized by polystyrene, acrylic acid, acrylic acid related esters class, more preferably, comprises the latex particle of two kinds of particle diameters of size among the reagent R2, and big particle diameter diameter is 150-500nm; Latex small particle diameter diameter is 50-100nm.
Preferably, the Nano microsphere mass percent is 0.1-1% among the reagent R2, and wherein big particle diameter mass percent is 0.01-0.1%; The small particle diameter mass percent is 0.1-0.8%.
Preferably, the combination of anti-people NGAL antibody and Nano microsphere is chemical crosslinking or physisorption among the reagent R2.
Preferably, described NGAL normative reference article comprise the NGAL of five variable concentrations, are respectively: 150ng/ml, 600ng/ml, 1500ng/ml, 3000ng/ml, 5000ng/ml, or five variable concentrations of similar ratio; Wherein damping fluid is a HEPES pH of buffer 7.4, and stabilizing agent is one or more mixing of BSA, sucrose, sodium chloride, polysorbas20.
Most preferably, described NGAL protein detection kit is selected from following combination:
Kit 1: reagent R1 consists of: pH 7.3, and Main Ingredients and Appearance is:
Glycocoll 50mM,
NaCl?0.15M，
NaN3?0.1％，
EDTA?10mM，
BSA?0.1％，
Rabbit igg 1%;
R2 consists of:
PH 7.0, and principal ingredient is:
HEPES?20mM，
NaN3?0.1％，
Small grain size latex content is: 2%,
Large grain size latex content is: 0.5%,
The work calibration object is: buffering matrix, and it is by HEPES 50mM, NaCl 0.15M, NaN3 0.1%, EDTA 10mM, BSA 0.5%, sucrose 5%, all the other are the deionized water composition, are dissolved in the NGAL normative reference article of the 5000ng/ml concentration of said matrix;
Kit 2: reagent R1 consists of: pH 7.4, and Main Ingredients and Appearance is:
Glycocoll 50mM,
NaCl?0.25M，
NaN3?0.1％，
EDTA?10mM，
BSA?0.1％，
R2 consists of: pH 7.0
HEPES?20mM，
NaN3?0.1％，
Small grain size latex content is: 2%,
Large grain size latex content is: 0.5%,
The work calibration object is: buffering matrix, and it is by HEPES 50mM, NaCl 0.15M, NaN 30.1%, EDTA 10mM, BSA 0.5%, polysorbas20 0.1%, all the other are the deionized water composition, are dissolved in the NGAL normative reference article of the 5000ng/ml concentration of said matrix;
Kit 3: reagent R1 consists of: pH 7.4, and Main Ingredients and Appearance is:
Glycocoll 50mM,
NaCl?0.15M，
NaN3?0.1％，
EDTA?10mM，
BSA?0.1％，
Mouse IgG 1%,
R2 consists of: pH 7.0
HEPES?20mM，
NaN3?0.1％，
Small grain size latex content is: 2%,
Large grain size latex content is: 0.5%,
The work calibration object is: buffering matrix, and it is by HEPES 50mM, NaCl 0.15M, NaN3 0.1%, EDTA 10mM, BSA 0.5%, sucrose 5%, all the other are the deionized water composition, are dissolved in the NGAL normative reference article of the 5000ng/ml concentration of said matrix;
Second aspect of the present invention relates to the preparation method of aforesaid NGAL detection kit; Comprise step: big grain diameter nano microballoon and anti-people NGAL antibody-solutions are blended in the pH7.0 damping fluid; Stirring at room 3 hours; Antibody combines fully with Nano microsphere after, the centrifugal supernatant that goes, ultrasonic being uniformly dispersed after the adding damping fluid; Small particle diameter Nano microsphere and anti-people NGAL antibody-solutions are blended in pH 7.0 damping fluids, stirring at room 3 hours, antibody combines the fully centrifugal supernatant that goes in back, ultrasonic being uniformly dispersed after the adding damping fluid with Nano microsphere; The size particles solution in certain proportion is mixed.
In other words; The application's problem to be solved is to overcome the defective that above-mentioned available reagent box exists; Provide the compound latex enhancing immune of a kind of size particles than turbid kit, detect the content of NGAL in blood plasma or the urine, to reach the purpose of the high sensitivity and the wide range of linearity simultaneously.
For reaching the purpose that a kind of kit possesses the high sensitivity and the wide range of linearity simultaneously, the application mainly starts with from immune response speed, promptly under low NGAL concentration, higher immune agglutination speed arranged, and guarantees high sensitivity; Under high NGAL concentration, lower immune agglutination speed arranged simultaneously, guarantee the wide range of linearity.
Particularly, the application's inventive concept is: behind (1) large grain size latex pan coating antibody, itself and antigen compatibility are higher, antigen-antibody reaction speed when greatly having improved low concentration NGAL, thereby the sensitivity that has improved reagent; (2) behind the small particle diameter pan coating antibody, its compatibility is relatively low, under high antigen concentration, still can form antigen antibody complex, thereby has improved the range of linearity of reagent; (3) after big small particle diameter is blended in same buffer solution system, high sensitivity and wide range of linearity purpose have been reached simultaneously.
The application's technical scheme is:
A kind of NGAL detection kit is characterized in that, comprises reagent R1, reagent R2, and described reagent R1 is for being fit to the damping fluid of NGAL and anti-people NGAL antibody response; Described reagent R2 is the Nano microsphere that is combined with anti-people NGAL antibody, places suitable damping fluid.
Described R1 damping fluid is one or more in phosphate buffer, glycine buffer, HEPES damping fluid, borate buffer, acetate buffer or the ammonium chloride buffer.Its concentration is 10-500mM; Reagent R2 damping fluid is one or more in phosphate buffer, glycine buffer, HEPES damping fluid, borate buffer, acetate buffer or the ammonium chloride buffer.Its concentration is 10-500mM.
Described anti-people NGAL antibody is polyclonal antibody or monoclonal antibody.
Described polyclonal antibody is the anti-people NGAL of rabbit antibody, goat-anti people NGAL antibody or the anti-people NGAL of chicken antibody.
Described monoclonal antibody is a mouse-anti people NGAL antibody.
Nano microsphere among the described reagent R2 is one or more latex that are polymerized by polystyrene latex, acrylic acid, acrylic acid related esters class.
Latex among the described reagent R2 comprises the latex of two kinds of particle diameters of size.The bulky grain diameter is 150-500nm; The latex small particle diameters is 50-100nm.
Latex quality percentage is 0.1-1% among the described reagent R2, and wherein the bulky grain mass percent is 0.01-0.1%; Latex granule mass percent is 0.1-0.8%.
Anti-people NGAL antibody and latex particle combination are physisorption or chemical crosslinking among the described reagent R2.
The application's major advantage is: (1) bulky grain latex surface hydrophobic is higher, and behind the coated antibody, itself and antigen compatibility are higher, antigen-antibody reaction speed when greatly having improved low concentration NGAL, thereby the sensitivity that has improved reagent.(2) behind the granule pan coating antibody, its compatibility is relatively low, under high antigen concentration, still can form antigen antibody complex, thereby has improved the range of linearity of reagent; (3) after size particles is blended in same buffer solution system; Because bulky grain and NGAL and antigen adhesion are strong, therefore at first combine, like this under low NGAL concentration with antigen; Reagent also has higher sensitivity; When NGAL concentration was higher, antigen can combine with granule to form immune complex, and reagent has the wide range of linearity like this.Though prior art also has the Nano microsphere that utilizes two kinds of particle diameters of size simultaneously to realize improving detectability and the report that enlarges the range of linearity that detects; But the prior art size particles adopts conventional bag by mode, and is higher for the antibody performance requirement, two kinds of antibody of the needs that have; A kind of high-affinity antibody of the needs that have; And the application adopts different coated antibody modes owing to size particles, and antagonist does not have special demands, only needs a kind of antibody just.
Description of drawings
Fig. 1: NGAL kit typical curve.
Fig. 2: the present invention and enzyme-linked immunosorbent assay blood plasma measured value correlativity.
Fig. 3: the present invention and enzyme-linked immunosorbent assay urine measured value correlativity.
Embodiment
To further specify the present invention through following non-limiting example below, as well known to those skilled in the art, under the situation that does not deviate from spirit of the present invention, can make many modifications to the present invention, such modification also falls into scope of the present invention.For example, as well known to those skilled in the art, can use the NGAL antibody in different plant species source, the particle size range of the Nano microsphere of said two kinds of particle diameters also can be wideer, and such variation also all falls within protection scope of the present invention.
Following experimental technique is conventional method if no special instructions, and employed experiment material such as antibody and Nano microsphere all can easily obtain from commercial company if no special instructions.
Embodiment
The preparation of embodiment 1:NGAL detection kit
The application has exemplarily prepared three kinds of NGAL detection kit, and its preparation raw material, preparation steps, performance are following:
(1) kit 1:
R1：pH?7.3，
Main Ingredients and Appearance is:
Glycocoll 50mM,
NaCl?0.15M，
NaN3?0.1％，
EDTA?10mM，
BSA?0.1％，
Rabbit igg 1%,
It is following that R2 prepares process:
The small grain size latex reagent preparation:
1, at room temperature dilute the anti-people NGAL of 4ml rabbit polyclonal antibody (10mg/ml) with 20mM HEPES solution (pH 7.0), to the antibody final concentration be 2mg/ml;
2, at room temperature dilute 4ml 74nm latex solution (10%wt) with 20mM HEPES solution (pH 7.0), making latex concentration is 4%;
3, will dilute good antibody and latex mixing, the room temperature concussion adds the EDAC solution 10ml with 20mM HEPES solution (pH 7.0) preparation 5mg/ml after 30 minutes, and room temperature reaction 3h adds 5ml confining liquid (1%BSA, 1% polysorbas20 solution) again, sealing 2h;
4, centrifugal, abandon supernatant, with 20mM HEPES solution (pH 7.0) diluted latex to 2.5%, and add 0.1% antiseptic, for use after the ultrasonic dispersion.
The large grain size latex reagent preparation:
1, at room temperature dilute the anti-people NGAL of 4ml rabbit polyclonal antibody (10mg/ml) with 20mM HEPES solution (pH 7.0), to the antibody final concentration be 2mg/ml;
2, at room temperature dilute 1ml 220nm latex solution (10%wt) with 20mM HEPES solution (pH 7.0), making latex concentration is 4%;
3, will dilute good antibody and latex mixing; The room temperature concussion adds the EDAC solution 10ml with 20mM HEPES solution (pH 7.0) preparation 5mg/ml after 30 minutes, room temperature reaction 3h adds the 5ml confining liquid again and (contains 1%BSA; 1% polysorbas20 solution), sealing 2h;
4, centrifugal, abandon supernatant, with 20mM HEPES solution (pH 7.0) diluted latex to 2.5%, and add 0.1% antiseptic, for use behind the ultrasonic dispersion latex particle.
5, large and small particle reagents is mixed with final R2 reagent: after size particles is mixed, stir, the R2 preparation finishes:
R2：pH?7.0，
Principal ingredient is:
HEPES?20mM，
NaN3?0.1％，
Small grain size latex content is: 2%,
Large grain size latex content is: 0.5%.
It is following that normative reference is tasted with discrimination the damping fluid matrix components:
HEPES?50mM，
NaCl?0.15M，
NaN3?0.1％，
EDTA?10mM，
BSA?0.5％，
Sucrose 5%,
All the other are deionized water,
Add in the above-mentioned damping fluid matrix by the required concentration of the normative reference article pure article of NGAL of will recombinating; Make the NGAL normative reference article of 5000ng/ml concentration, the normative reference article of all the other concentration (150ng/ml, 600ng/ml, 1500ng/ml, 3000ng/ml) can be obtained with the dilution of damping fluid matrix by the 5000ng/ml standard items.
(2) kit 2:
R1：pH?7.4，
Main Ingredients and Appearance is:
Glycocoll 50mM
NaCl?0.25M
NaN3?0.1％
EDTA 10mM
BSA?0.1％
It is following that R2 prepares process:
The small grain size latex reagent preparation:
1, at room temperature dilute the anti-people NGAL of 5ml chicken polyclonal antibody (3mg/ml) with 20mM HEPES solution (pH 7.0), to the antibody final concentration be 2mg/ml;
2, at room temperature dilute 4ml 74nm latex solution (10%wt) with 20mM HEPES solution (pH 7.0), making latex concentration is 4%;
3, will dilute good antibody and latex mixing, the room temperature concussion adds the EDAC solution 10ml with 20mM HEPES solution (pH 7.0) preparation 5mg/ml after 30 minutes, and room temperature reaction 3h adds 5ml confining liquid (1%BSA, 1% polysorbas20 solution) again, sealing 2h;
4, centrifugal, abandon supernatant, with 20mM HEPES solution (pH 7.0) diluted latex to 2.5%, and add 0.1% antiseptic, for use after the ultrasonic dispersion.
The large grain size latex reagent preparation:
1, at room temperature dilute the anti-people NGAL of 4ml chicken polyclonal antibody (5mg/ml) with 20mM HEPES solution (pH 7.0), to the antibody final concentration be 2mg/ml;
2, at room temperature dilute 1ml 220nm latex solution (10%wt) with 20mM HEPES solution (pH 7.0), making latex concentration is 4%;
3, will dilute good antibody and latex mixing; The room temperature concussion adds the EDAC solution 10ml with 20mM HEPES solution (pH 7.0) preparation 5mg/ml after 30 minutes, room temperature reaction 3h adds the 5ml confining liquid again and (contains 1%BSA; 1% polysorbas20 solution), sealing 2h;
4, centrifugal, abandon supernatant, with 20mM HEPES solution (pH 7.0) diluted latex to 2.5%, and add 0.1% antiseptic, for use behind the ultrasonic dispersion latex particle.
5, large and small particle reagents is mixed with final R2 reagent: after size particles is mixed, stir, the R2 preparation finishes.
R2：pH?7.0，
Principal ingredient is:
HEPES?20mM，
NaN3?0.1％，
Small grain size latex content is: 2%,
Large grain size latex content is: 0.5%.
It is following that normative reference is tasted with discrimination the damping fluid matrix components:
HEPES?50mM，
NaCl?0.15M，
NaN3?0.1％，
EDTA?10mM，
BSA?0.5％，
Polysorbas20 0.1%,
All the other are deionized water,
Add in the above-mentioned damping fluid matrix by the required concentration of the normative reference article pure article of NGAL of will recombinating; Make the NGAL normative reference article of 5000ng/ml concentration, the normative reference article of all the other concentration (150ng/ml, 600ng/ml, 1500ng/ml, 3000ng/ml) can be obtained with the dilution of damping fluid matrix by the 5000ng/ml standard items.
(3) kit 3:
R1：pH?7.4，
Main Ingredients and Appearance is:
Glycocoll 50mM,
NaCl?0.15M，
NaN3?0.1％，
EDTA?10mM，
BSA?0.1％，
Mouse IgG 1%,
It is following that R2 prepares process:
The small grain size latex reagent preparation:
1, at room temperature dilute 5ml mouse-anti people NGAL polyclonal antibody (5mg/ml) with 20mM HEPES solution (pH 7.0), to the antibody final concentration be 2mg/ml;
2, at room temperature dilute 4ml 74nm latex solution (10%wt) with 20mM HEPES solution (pH 7.0), making latex concentration is 4%;
3, will dilute good antibody and latex mixing, the room temperature concussion adds the EDAC solution 10ml with 20mM HEPES solution (pH 7.0) preparation 5mg/ml after 30 minutes, and room temperature reaction 3h adds 5ml confining liquid (1%BSA, 1% polysorbas20 solution) again, sealing 2h;
4, centrifugal, abandon supernatant, with 20mM HEPES solution (pH 7.0) diluted latex to 2.5%, and add 0.1% antiseptic, for use after the ultrasonic dispersion.
The large grain size latex reagent preparation:
1, at room temperature dilute 4ml mouse-anti people NGAL polyclonal antibody (5mg/ml) with 20mM HEPES solution (pH 7.0), to the antibody final concentration be 2mg/ml;
2, at room temperature dilute 1ml 220nm latex solution (10%wt) with 20mM HEPES solution (pH 7.0), making latex concentration is 4%;
3, will dilute good antibody and latex mixing; The room temperature concussion adds the EDAC solution 10ml with 20mM HEPES solution (pH 7.0) preparation 5mg/ml after 30 minutes, room temperature reaction 3h adds the 5ml confining liquid again and (contains 1%BSA; 1% polysorbas20 solution), sealing 2h;
4, centrifugal, abandon supernatant, with 20mM HEPES solution (pH 7.0) diluted latex to 2.5%, and add 0.1% antiseptic, for use behind the ultrasonic dispersion latex particle.
5, large and small particle diameter reagent mix prepares final R2 reagent: after size particles is mixed, stir, the R2 preparation finishes.
R2：pH?7.0，
Principal ingredient is:
HEPES?20mM，
NaN3?0.1％，
Small grain size latex content is: 2%,
Large grain size latex content is: 0.5%.
It is following that normative reference is tasted with discrimination the damping fluid matrix components:
HEPES?50mM
NaCl?0.15M
NaN3?0.1％
EDTA 10mM
BSA?0.5％
Polysorbas20 0.1%
Sucrose 5%
All the other are deionized water
Add in the above-mentioned damping fluid matrix by the required concentration of the normative reference article pure article of NGAL of will recombinating; Make the NGAL normative reference article of 5000ng/ml concentration, the normative reference article of all the other concentration (0ng/ml, 150ng/ml, 600ng/ml, 1500ng/ml, 3000ng/ml) can be obtained with the dilution of damping fluid matrix by the 5000ng/ml standard items.
Embodiment 2:NGAL detection kit determination step, the drafting of calibration curve compares research with the product of certain famous brand name on the market; It also is the latex enhancing immune turbidimetry that this producer's reagent detects principle, the main composition: R1,10mmol Tris; 0.1%NAN3,0.5%BSA; R2,0.15% is coated with the latex suspension of anti-people NGAL monoclonal antibody, and the contrast situation is following:
The application's determination step is as shown in table 1.
Table 1: the determination step of the application's kit:
Kit 2 with embodiment 1-(2) is an example, and standard items concentration is transverse axis, corresponding Δ OD
570Be the longitudinal axis, adopt nonlinear fitting, draw out typical curve like spline, the result is as shown in Figure 1.
The linearity and the LDL of embodiment 3:NGAL kit
Linear experiment: adopt method known to those skilled in the art; A certain high NGAL concentration samples (urine specimen during the acute injury of kidney patient 2h of operation back) is done doubling dilution; Kit 2 with shown in the embodiment 1-(2) is a detection kit, measures the concentration after diluting, and measures three times and averages; Compare with theoretical concentration, calculate its deviation from linearity.Simultaneously with above-mentioned market on certain famous brand name compare, measure the result shown in table 2, table 3, table 4 and table 5.
The high value scope of table 2 the application kit 2 is linear: the ng/ml of unit
Table 3: the high value scope of contrast agents is linear: the ng/ml of unit
| Extension rate | Measure 1 | Measure 1 | Measure 1 | Measure average | Theoretical value | Deviation from linearity % |
| 1 | 4102.3 | 4056.5 | 4085.6 | 4081.467 | 6839.523 | 40.33 |
| 0.9 | 4110.5 | 4125.4 | 4108.7 | 4114.867 | 6162.513 | 33.23 |
| 0.8 | 4178.4 | 4165.2 | 4182.6 | 4175.4 | 5485.503 | 23.88 |
| 0.7 | 4034.5 | 3955.1 | 3927.5 | 3972.367 | 4808.493 | 17.39 |
| 0.6 | 3893.6 | 3851.8 | 3906.6 | 3884 | 4131.483 | 5.99 |
| 0.5 | 3426 | 3466.5 | 3439.3 | 3443.933 | 3454.473 | 0.305 |
| 0.4 | 2793.6 | 2781.5 | 2815.5 | 2796.867 | 2777.463 | -0.7 |
| 0.3 | 2071.2 | 2039.1 | 2063.4 | 2057.9 | 2100.453 | 2.026 |
| 0.2 | 1486.1 | 1499.5 | 1491.9 | 1492.5 | 1423.443 | -4.85 |
| 0.1 | 712.9 | 714.6 | 705.7 | 711.0667 | 746.433 | 4.738 |
Table 4: the low value scope of the application's kit 2 is linear: the ng/ml of unit
| Extension rate | Measure 1 | Measure 1 | Measure 1 | Measure average | Theoretical value | Deviation from linearity % |
| 1 | 720.3 | 720.5 | 720.6 | 720.47 | 711.07 | 1.321 |
| 0.9 | 630.5 | 632.4 | 638.7 | 633.87 | 639.12 | -0.82 |
| 0.8 | 568.4 | 565.2 | 562.6 | 565.4 | 567.18 | -0.31 |
| 0.7 | 504.5 | 495.1 | 492.5 | 497.37 | 495.23 | 0.43 |
| 0.6 | 419.6 | 415.8 | 420.6 | 418.67 | 423.29 | -1.09 |
| 0.5 | 346 | 346.5 | 349.3 | 347.27 | 351.35 | -1.16 |
| 0.4 | 278.6 | 271.5 | 280.5 | 276.87 | 279.40 | -0.91 |
| 0.3 | 206.2 | 209.1 | 203.4 | 206.23 | 207.46 | -0.59 |
| 0.2 | 141.1 | 140.5 | 141.9 | 141.17 | 135.51 | 4.17 |
| 0.1 | 65.7 | 68.6 | 69.7 | 68 | 63.57 | 6.97 |
| 0.05 | 25.5 | 25.6 | 25.4 | 25.5 | 27.60 | -7.60 |
| 0 | 0.02 | -0.01 | 0.01 | 0.01 | 0 | - |
Table 5: the low value scope of contrast agents is linear: the ng/ml of unit
| Extension rate | Measure 1 | Measure 1 | Measure 1 | Measure average | Theoretical value | Deviation from linearity % |
| 1 | 740.3 | 740.5 | 740.6 | 740.4667 | 731.066 | -1.29 |
| 0.9 | 650.5 | 652.4 | 658.7 | 653.8667 | 659.122 | 0.797 |
| 0.8 | 588.4 | 585.2 | 582.6 | 585.4 | 587.178 | 0.303 |
| 0.7 | 524.5 | 515.1 | 512.5 | 517.3667 | 515.234 | -0.41 |
| 0.6 | 439.6 | 435.8 | 440.6 | 438.6667 | 443.29 | 1.043 |
| 0.5 | 366 | 366.5 | 369.3 | 367.2667 | 371.346 | 1.099 |
| 0.4 | 298.6 | 291.5 | 300.5 | 296.8667 | 299.402 | 0.847 |
| 0.3 | 226.2 | 229.1 | 223.4 | 226.2333 | 227.458 | 0.538 |
| 0.2 | 161.1 | 160.5 | 161.9 | 161.1667 | 155.514 | -3.63 |
| 0.1 | 85.7 | 88.6 | 89.7 | 88 | 83.57 | -5.3 |
| 0.05 | 45.5 | 45.6 | 45.4 | 45.5 | 47.598 | 4.408 |
| 0 | 740.3 | 740.5 | 740.6 | 740.4667 | 731.066 | -1.29 |
Can find out that from last table the range of linearity of the application's kit 2 is 0-7000ng/ml, simultaneously, the result of the test result of two other kit of the present invention and kit 2 is similar.The range of linearity of contrast agents box is merely 0-3800ng/ml.
LDL: adopt method known to those skilled in the art; With blank solution with a several low concentration samples that dilute with physiological saline; With the kit 2 shown in the embodiment 1-(2) is that detection kit is measured, and replication 15 times reads the absorbance variable quantity.Calculate the absorbance of each sample of deduction behind the blank absorbency then, computation of mean values, standard deviation.The possibility of employing 99.7% is calculated LDL.The average of each sample deducted 3 times standard deviation separately; 3 times standard deviation with blank solution compares then; If the former is higher than the latter, just assert has minimum absorbance that 99.7% possibility occurs necessarily greater than blank absorbency, can quantitative reporting the result.Measure the result shown in table 6-9.
Table 6: the LDL of the application's kit 2:
Table 7: the absorbance that the said determination result is corresponding
| (A-A0)*10000 | 1 | 2 | 3 | Water is blank | |
| 1 | 41 | 24 | 15 | 8 | 1 |
| 2 | 38 | 22 | 15 | 5 | 6 |
| 3 | 35 | 22 | 12 | 9 | 4 |
| 4 | 29 | 23 | 10 | 5 | 0 |
| 5 | 34 | 19 | 13 | 6 | 2 |
| 6 | 42 | 19 | 14 | 3 | 3 |
| 7 | 36 | 22 | 14 | 4 | 1 |
| 8 | 39 | 22 | 15 | 1 | 1 |
| 9 | 40 | 23 | 14 | 3 | 3 |
| 10 | 34 | 24 | 16 | 7 | -1 |
| 11 | 42 | 22 | 14 | 4 | 2 |
| 12 | 37 | 21 | 17 | 5 | 6 |
| 13 | 39 | 22 | 11 | 8 | 0 |
| 14 | 44 | 21 | 12 | 4 | -1 |
| 15 | 41 | 23 | 16 | 4 | 5 |
| Average | 38.06667 | 21.93333 | 13.86667 | 5.066667 | 2.133333 |
| SD | 3.954503 | 1.486447 | 1.959106 | 2.186539 | 2.325838 |
| CV％ | 10.38836 | 6.777113 | 14.12817 | 43.15537 | 109.0237 |
| 3SD | 11.86351 | 4.45934 | 5.877317 | 6.559617 | 6.977515 |
| A-3SD | 26.20316 | 17.47399 | 7.989349 | -1.49295 | 9.110848 |
| A-3SD&3S0 | 17.09231 | 8.363145 | -1.1215 | -10.6038 |
Table 8: the LDL of contrast agents
Table 9: the absorbance that the said determination result is corresponding
Learnt that by above table the lowest detection of the application's kit is limited to 10.39ng/ml, the result of the test result of two other kit of the present invention and kit 2 is similar simultaneously.Contrast agents is 14.67ng/ml.
Embodiment 4: the correlativity of the application NGAL kit and enzyme-linked immunosorbent assay (Elisa) measured value
Carrying out two kinds of sample correlations investigates: EDTA anticoagulate plasma, urine specimen.
The EDTA anticoagulate plasma NGAL measured value that obtains with embodiment 1-(2) compares with the measured value through the enzyme-linked immunosorbent assay gained, and as shown in Figure 2, correlativity R=0.9982 (y=0.9646x+4.7736) is known in the line retrace analysis of going forward side by side; Demonstrate present embodiment and enzyme-linked immunosorbent assay and have good correlativity aspect the EDTA anticoagulate plasma NGAL mensuration.
The urine NGAL measured value that obtains with the said kit of embodiment 1-(2) compares with the measured value through the enzyme-linked immunosorbent assay gained; As shown in Figure 3; The line retrace analysis of going forward side by side; Know correlativity R=0.9998 (y=0.9794x-9.0372), this kit 2 that demonstrates this present invention also has good correlativity with enzyme-linked immunosorbent assay aspect the urine NGAL mensuration.Simultaneously, the result of the test result of two other kit of the present invention and kit 2 is similar.
Claims (11)
1. a NGAL detection kit comprises reagent R1, reagent R2 and work calibration object, and wherein reagent R1 is NGAL and anti-people NGAL antibody response damping fluid; Reagent R2 is the Nano microsphere solution that is combined with anti-people NGAL antibody; The work calibration object is made up of damping fluid, stabilizing agent, antiseptic and NGAL normative reference article; It is characterized in that, comprise the Nano microsphere that is combined with anti-people NGAL antibody of two kinds of particle diameters of size among the said reagent R2, big particle diameter is 150-500nm; Small particle diameter is 50-100nm; Preferably, big particle diameter is 200-300nm, and small particle diameter is 70-90nm.
2. NGAL detection kit according to claim 1; It is characterized in that; Reagent R1 damping fluid is selected from one or more in phosphate buffer, glycine buffer, HEPES damping fluid, borate buffer, acetate buffer or the ammonium chloride buffer, and its concentration is 10-500mM; The used damping fluid of reagent R2 is one or more in phosphate buffer, glycine buffer, HEPES damping fluid, borate buffer, acetate buffer or the ammonium chloride buffer, and its concentration is 10-500mM.
3. NGAL protein detection kit according to claim 1 and 2 is characterized in that, described anti-people NGAL antibody is polyclonal antibody or monoclonal antibody.
4. NGAL detection kit according to claim 3 is characterized in that, described polyclonal antibody is a kind of of the anti-people NGAL of rabbit antibody, goat-anti people NGAL antibody or the anti-people NGAL of chicken antibody; Monoclonal antibody is a mouse-anti people NGAL antibody.
5. according to each described NGAL detection kit of claim 1 to 4, it is characterized in that the Nano microsphere among the reagent R2 is one or more latex that are polymerized by polystyrene, acrylic acid, acrylic acid related esters class.
6. NGAL detection kit according to claim 5 is characterized in that, comprises the latex particle of two kinds of particle diameters of size among the reagent R2, and big particle diameter diameter is 150-500nm; Latex small particle diameter diameter is 50-100nm.
7. according to each described NGAL detection kit of claim 1 to 6, it is characterized in that the Nano microsphere mass percent is 0.1-1% among the reagent R2, wherein big particle diameter mass percent is 0.01-0.1%; The small particle diameter mass percent is 0.1-0.8%.
8. according to each described NGAL detection kit of claim 1 to 7, it is characterized in that the combination of anti-people NGAL antibody and Nano microsphere is chemical crosslinking or physisorption among the reagent R2.
9. according to each described NGAL protein detection kit of claim 1 to 8, it is characterized in that described NGAL normative reference article comprise the NGAL of five variable concentrations; Be respectively: 150ng/ml; 600ng/ml, 1500ng/ml, 3000ng/ml; 5000ng/ml, or five variable concentrations of similar ratio; Wherein damping fluid is a HEPES pH of buffer 7.4, and stabilizing agent is one or more mixing of BSA, sucrose, sodium chloride and polysorbas20.
10. according to each described NGAL protein detection kit of claim 1 to 9, it is characterized in that its composition is selected from following combination:
Kit 1: reagent R1 consists of: pH 7.3, and Main Ingredients and Appearance is:
Glycocoll 50mM,
NaCl?0.15M，
NaN3?0.1％，
EDTA?10mM，
BSA?0.1％，
Rabbit igg 1%;
R2 consists of:
PH 7.0, and principal ingredient is:
HEPES?20mM，
NaN3?0.1％，
Small grain size latex content is: 2%,
Large grain size latex content is: 0.5%,
The work calibration object is: buffering matrix, and it is by HEPES 50mM, NaCl 0.15M, NaN3 0.1%, EDTA 10mM, BSA 0.5%, sucrose 5%, all the other are the deionized water composition, are dissolved in the NGAL normative reference article of the 5000ng/ml concentration of said matrix;
Kit 2: reagent R1 consists of: pH 7.4, and Main Ingredients and Appearance is:
Glycocoll 50mM,
NaCl?0.25M，
NaN3?0.1％，
EDTA?10mM，
BSA?0.1％，
R2 consists of: pH 7.0
HEPES?20mM，
NaN3?0.1％，
Small grain size latex content is: 2%,
Large grain size latex content is: 0.5%,
The work calibration object is: buffering matrix, and it is by HEPES 50mM, NaCl 0.15M, NaN3 0.1%, EDTA 10mM, BSA 0.5%, polysorbas20 0.1%, all the other are the deionized water composition, are dissolved in the NGAL normative reference article of the 5000ng/ml concentration of said matrix;
Kit 3: reagent R1 consists of: pH 7.4, and Main Ingredients and Appearance is:
Glycocoll 50mM,
NaCl?0.15M，
NaN3?0.1％，
EDTA?10mM，
BSA?0.1％，
Mouse IgG 1%,
R2 consists of: pH 7.0
HEPES?20mM，
NaN3?0.1％，
Small grain size latex content is: 2%,
Large grain size latex content is: 0.5%,
The work calibration object is: buffering matrix, and it is by HEPES 50mM, NaCl 0.15M, NaN3 0.1%, EDTA 10mM, BSA 0.5%, sucrose 5%, all the other are the deionized water composition, are dissolved in the NGAL normative reference article of the 5000ng/ml concentration of said matrix.
11. preparation method according to each described NGAL detection kit of claim 1 to 9; Comprise step: big grain diameter nano microballoon and anti-people NGAL antibody-solutions are blended in the pH7.0 damping fluid; Stirring at room 3 hours; Antibody combines fully with Nano microsphere after, the centrifugal supernatant that goes, ultrasonic being uniformly dispersed after the adding damping fluid; Small particle diameter Nano microsphere and anti-people NGAL antibody-solutions are blended in pH 7.0 damping fluids, stirring at room 3 hours, antibody combines the fully centrifugal supernatant that goes in back, ultrasonic being uniformly dispersed after the adding damping fluid with Nano microsphere; The size particles solution in certain proportion is mixed.
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201110454991.7A CN102590524B (en) | 2011-12-30 | 2011-12-30 | Neutrophil gelatinase-associated lipocalin detection kit |
| PCT/CN2012/086508 WO2013097606A1 (en) | 2011-12-30 | 2012-12-13 | Assay kit for neutrophil gelatinase-associated lipocalin |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201110454991.7A CN102590524B (en) | 2011-12-30 | 2011-12-30 | Neutrophil gelatinase-associated lipocalin detection kit |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102590524A true CN102590524A (en) | 2012-07-18 |
| CN102590524B CN102590524B (en) | 2016-01-06 |
Family
ID=46479453
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201110454991.7A Active CN102590524B (en) | 2011-12-30 | 2011-12-30 | Neutrophil gelatinase-associated lipocalin detection kit |
Country Status (2)
| Country | Link |
|---|---|
| CN (1) | CN102590524B (en) |
| WO (1) | WO2013097606A1 (en) |
Cited By (33)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102967714A (en) * | 2012-12-10 | 2013-03-13 | 天津市协和医药科技集团有限公司 | Neutrophil gelatinase associated lipocalin (NGAL) chemiluminescence detection kit |
| CN103048464A (en) * | 2012-10-17 | 2013-04-17 | 武汉生之源生物科技有限公司 | Neutrophile granulocyte gelatinase related lipid transport protein detection kit and preparation method thereof |
| WO2013097606A1 (en) * | 2011-12-30 | 2013-07-04 | 北京九强生物技术股份有限公司 | Assay kit for neutrophil gelatinase-associated lipocalin |
| CN103235130A (en) * | 2013-04-18 | 2013-08-07 | 宁波瑞源生物科技有限公司 | High-sensitivity high-linearity D-D dimer detection kit and application method thereof |
| CN103323596A (en) * | 2012-10-31 | 2013-09-25 | 武汉生之源生物科技有限公司 | Detection kit for myeloperoxidase content and preparation method thereof |
| CN103777023A (en) * | 2013-12-16 | 2014-05-07 | 深圳市安之酶生物技术有限公司 | Kit for measuring adiponectin by nano latex enhanced immune turbidimetry |
| CN103995128A (en) * | 2014-05-08 | 2014-08-20 | 北京玖佳宜科技有限公司 | Neutrophil gelatinase-associated lipocalin detection kit and preparation |
| CN104155458A (en) * | 2013-11-27 | 2014-11-19 | 新昌县美迪生物科技有限公司 | Quantitative detection kit for human chorionic gonadotropin and application of quantitative detection kit |
| CN104198732A (en) * | 2014-08-28 | 2014-12-10 | 宁波瑞源生物科技有限公司 | Kit for detecting content of neutrophil gelatinase-associated lipocalin (NGAL) |
| CN104215769A (en) * | 2014-08-14 | 2014-12-17 | 上海睿康生物科技有限公司 | Latex enhanced immunoturbidimetry NGAL detection kit |
| CN104407154A (en) * | 2014-12-05 | 2015-03-11 | 重庆中元生物技术有限公司 | High-sensitivity latex enhanced immunoturbidimetry kit |
| CN104865386A (en) * | 2015-05-08 | 2015-08-26 | 浙江蓝森生物科技有限公司 | Method, reagent and kit for quantitative determination of NGAL content in human serum |
| CN105158476A (en) * | 2015-06-03 | 2015-12-16 | 南京闻智生物科技有限公司 | Full-scale range C-reactive protein latex-enhanced immunoturbidimetry detection kit |
| CN105738617A (en) * | 2016-04-01 | 2016-07-06 | 武汉生之源生物科技股份有限公司 | Cystatin C latex-particle-enhanced turbidimetry detection reagent kit and application thereof |
| CN106483297A (en) * | 2016-09-29 | 2017-03-08 | 浙江达美生物技术有限公司 | Neutrophil gelatinase-associated lipocalin determines reagent and preparation method |
| CN106501526A (en) * | 2016-10-03 | 2017-03-15 | 王贤俊 | A kind of combination directing agent optimizes latex coupled antibody detection prealbumin（PA）Test kit |
| CN106526198A (en) * | 2016-10-03 | 2017-03-22 | 王贤俊 | Combined orientation agents optimized latex coupled antibody detection method of prealbumin (PA) |
| CN106568944A (en) * | 2016-10-03 | 2017-04-19 | 王贤俊 | Latex immunoturbidimetric kit for detecting AGEs |
| CN107167586A (en) * | 2017-07-06 | 2017-09-15 | 四川新健康成生物股份有限公司 | A kind of antistreptolysin O (ASO) detection kit and preparation method thereof |
| CN107340395A (en) * | 2017-07-05 | 2017-11-10 | 深圳开立生物医疗科技股份有限公司 | A kind of Immunoturbidimetric kit for detecting Procalcitonin |
| CN107543931A (en) * | 2017-09-12 | 2018-01-05 | 四川新健康成生物股份有限公司 | Kit based on latex enhancing immune turbidimetry detection TnI and preparation method thereof |
| CN107894509A (en) * | 2017-10-20 | 2018-04-10 | 柏荣诊断产品（上海）有限公司 | A kind of method for improving latex immunoturbidimetry antigen excess and the range of linearity |
| CN107991496A (en) * | 2017-11-29 | 2018-05-04 | 山东诺安爱迪尔生物工程有限公司 | A kind of neutrophil gelatinase-associated lipocalin detection kit |
| WO2018119626A1 (en) * | 2016-12-27 | 2018-07-05 | 菲鹏生物股份有限公司 | Assay kit for neutrophil gelatinase-associated lipocalin |
| CN108362895A (en) * | 2018-02-12 | 2018-08-03 | 北京九强生物技术股份有限公司 | Folic acid detection kit and preparation method thereof |
| CN108414775A (en) * | 2018-03-02 | 2018-08-17 | 江西英大生物技术有限公司 | Neutrophil gelatinase-associated lipocalin detection kit |
| CN108896773A (en) * | 2018-08-23 | 2018-11-27 | 潍坊市康华生物技术有限公司 | A kind of gastrin 17 detection kit and its preparation and application |
| CN110133247A (en) * | 2019-05-24 | 2019-08-16 | 深圳上泰生物工程有限公司 | A kind of immune latex turbidimetry method assay kit for testing goal antigen |
| CN110568182A (en) * | 2019-09-12 | 2019-12-13 | 苏州普瑞斯生物科技有限公司 | Adiponectin-latex enhanced immunoturbidimetry kit and preparation method thereof |
| CN110824176A (en) * | 2019-11-28 | 2020-02-21 | 江西英大生物技术有限公司 | Neutrophil gelatinase-associated lipocalin detection kit |
| CN111721937A (en) * | 2019-03-29 | 2020-09-29 | 北京九强生物技术股份有限公司 | Human epididymis protein 4 immunoturbidimetry kit |
| CN113125699A (en) * | 2019-12-31 | 2021-07-16 | 科美诊断技术股份有限公司 | beta-hCG homogeneous phase chemiluminescence detection kit and application thereof |
| CN116413256A (en) * | 2023-02-13 | 2023-07-11 | 上海索昕生物科技有限公司 | Photosensitive microsphere for photoexcitation chemiluminescence detection |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104020283B (en) * | 2014-06-12 | 2016-01-20 | 长春理工大学 | A kind of monofilm dry plate for renal function detection and preparation method |
| CN111912990B (en) * | 2020-08-12 | 2023-12-29 | 山东博科生物产业有限公司 | Neutrophil gelatinase-associated lipocalin assay kit |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0898169A2 (en) * | 1997-08-11 | 1999-02-24 | F. Hoffmann-La Roche Ag | Microparticle enhanced light scattering assay and microparticle reagents therefor |
| CN1769894A (en) * | 2004-11-01 | 2006-05-10 | 上海申索佑福医学诊断用品有限公司 | Kit for determination of high-sensitive C-reactive protein |
| CN101566633A (en) * | 2009-04-17 | 2009-10-28 | 武汉华美生物工程有限公司 | Method for diagnosing, evaluating or testing cancer and foreseeing cancer severity |
| CN101680891A (en) * | 2007-11-21 | 2010-03-24 | 爱科来株式会社 | Measurement reagent, immune nephelometry using the same, and analyte analysis tool |
| CN102175871A (en) * | 2010-12-30 | 2011-09-07 | 北京九强生物技术股份有限公司 | Liquid double-reagent kit for measuring free light chains in serum or urine by double-latex method |
| CN102253217A (en) * | 2011-04-07 | 2011-11-23 | 武汉生之源生物科技有限公司 | Detection kit of latex particle enhanced neutrophil gelatinase-associated lipid transfer protein |
Family Cites Families (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US9182391B2 (en) * | 2008-05-09 | 2015-11-10 | Arkray, Inc. | Method of producing insoluble carrier particles, insoluble carrier particles, measurement reagent, specimen analyzing tool, and immunoturbidimetric assay |
| US7977110B2 (en) * | 2008-06-02 | 2011-07-12 | Children's Hospital Medical Center | Method for distinguishing between kidney dysfunctions |
| CN102072960A (en) * | 2010-03-29 | 2011-05-25 | 武汉生之源生物科技有限公司 | Method for detecting neutrophil gelatinase-associated lipocalin (NGAL) in sample |
| CN102243241A (en) * | 2011-04-07 | 2011-11-16 | 武汉生之源生物科技有限公司 | Homogeneous phase aerosol particle-type neutrophile granulocyte gelatinase-related lipid carrier protein determination kit and preparation method thereof |
| CN102680698B (en) * | 2011-07-29 | 2016-03-30 | 南京诺尔曼生物技术有限公司 | Neutrophil gelatinase-associated lipocalin (NGAL) measures kit (latex enhancing immune turbidimetry) |
| CN102590524B (en) * | 2011-12-30 | 2016-01-06 | 北京九强生物技术股份有限公司 | Neutrophil gelatinase-associated lipocalin detection kit |
| CN102662059B (en) * | 2012-05-11 | 2014-08-13 | 北京美康生物技术研究中心 | Latex-enhanced immunoturbidimetry kit for measuring Helicobacter pylori antibody as well as preparation method and application thereof |
-
2011
- 2011-12-30 CN CN201110454991.7A patent/CN102590524B/en active Active
-
2012
- 2012-12-13 WO PCT/CN2012/086508 patent/WO2013097606A1/en active Application Filing
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0898169A2 (en) * | 1997-08-11 | 1999-02-24 | F. Hoffmann-La Roche Ag | Microparticle enhanced light scattering assay and microparticle reagents therefor |
| CN1769894A (en) * | 2004-11-01 | 2006-05-10 | 上海申索佑福医学诊断用品有限公司 | Kit for determination of high-sensitive C-reactive protein |
| CN101680891A (en) * | 2007-11-21 | 2010-03-24 | 爱科来株式会社 | Measurement reagent, immune nephelometry using the same, and analyte analysis tool |
| CN101566633A (en) * | 2009-04-17 | 2009-10-28 | 武汉华美生物工程有限公司 | Method for diagnosing, evaluating or testing cancer and foreseeing cancer severity |
| CN102175871A (en) * | 2010-12-30 | 2011-09-07 | 北京九强生物技术股份有限公司 | Liquid double-reagent kit for measuring free light chains in serum or urine by double-latex method |
| CN102253217A (en) * | 2011-04-07 | 2011-11-23 | 武汉生之源生物科技有限公司 | Detection kit of latex particle enhanced neutrophil gelatinase-associated lipid transfer protein |
Cited By (42)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2013097606A1 (en) * | 2011-12-30 | 2013-07-04 | 北京九强生物技术股份有限公司 | Assay kit for neutrophil gelatinase-associated lipocalin |
| CN103048464A (en) * | 2012-10-17 | 2013-04-17 | 武汉生之源生物科技有限公司 | Neutrophile granulocyte gelatinase related lipid transport protein detection kit and preparation method thereof |
| CN103048464B (en) * | 2012-10-17 | 2015-04-15 | 武汉生之源生物科技有限公司 | Neutrophile granulocyte gelatinase related lipid transport protein detection kit and preparation method thereof |
| CN103323596A (en) * | 2012-10-31 | 2013-09-25 | 武汉生之源生物科技有限公司 | Detection kit for myeloperoxidase content and preparation method thereof |
| CN103323596B (en) * | 2012-10-31 | 2015-06-03 | 武汉生之源生物科技有限公司 | Detection kit for myeloperoxidase content and preparation method thereof |
| CN102967714A (en) * | 2012-12-10 | 2013-03-13 | 天津市协和医药科技集团有限公司 | Neutrophil gelatinase associated lipocalin (NGAL) chemiluminescence detection kit |
| CN103235130A (en) * | 2013-04-18 | 2013-08-07 | 宁波瑞源生物科技有限公司 | High-sensitivity high-linearity D-D dimer detection kit and application method thereof |
| CN104155458A (en) * | 2013-11-27 | 2014-11-19 | 新昌县美迪生物科技有限公司 | Quantitative detection kit for human chorionic gonadotropin and application of quantitative detection kit |
| CN103777023A (en) * | 2013-12-16 | 2014-05-07 | 深圳市安之酶生物技术有限公司 | Kit for measuring adiponectin by nano latex enhanced immune turbidimetry |
| CN103777023B (en) * | 2013-12-16 | 2015-04-08 | 深圳市安之酶生物技术有限公司 | Kit for measuring adiponectin by nano latex enhanced immune turbidimetry |
| CN103995128A (en) * | 2014-05-08 | 2014-08-20 | 北京玖佳宜科技有限公司 | Neutrophil gelatinase-associated lipocalin detection kit and preparation |
| CN104215769A (en) * | 2014-08-14 | 2014-12-17 | 上海睿康生物科技有限公司 | Latex enhanced immunoturbidimetry NGAL detection kit |
| CN104198732A (en) * | 2014-08-28 | 2014-12-10 | 宁波瑞源生物科技有限公司 | Kit for detecting content of neutrophil gelatinase-associated lipocalin (NGAL) |
| CN104198732B (en) * | 2014-08-28 | 2015-08-19 | 宁波瑞源生物科技有限公司 | A kind of neutrophil gelatinase-associated lipocalin reagent box for detecting content |
| CN104407154A (en) * | 2014-12-05 | 2015-03-11 | 重庆中元生物技术有限公司 | High-sensitivity latex enhanced immunoturbidimetry kit |
| CN104407154B (en) * | 2014-12-05 | 2016-03-30 | 重庆中元生物技术有限公司 | A kind of high sensitivity latex enhancing immune turbidimetry kit |
| CN104865386A (en) * | 2015-05-08 | 2015-08-26 | 浙江蓝森生物科技有限公司 | Method, reagent and kit for quantitative determination of NGAL content in human serum |
| CN105158476A (en) * | 2015-06-03 | 2015-12-16 | 南京闻智生物科技有限公司 | Full-scale range C-reactive protein latex-enhanced immunoturbidimetry detection kit |
| CN105738617A (en) * | 2016-04-01 | 2016-07-06 | 武汉生之源生物科技股份有限公司 | Cystatin C latex-particle-enhanced turbidimetry detection reagent kit and application thereof |
| CN106483297A (en) * | 2016-09-29 | 2017-03-08 | 浙江达美生物技术有限公司 | Neutrophil gelatinase-associated lipocalin determines reagent and preparation method |
| CN106501526A (en) * | 2016-10-03 | 2017-03-15 | 王贤俊 | A kind of combination directing agent optimizes latex coupled antibody detection prealbumin（PA）Test kit |
| CN106568944A (en) * | 2016-10-03 | 2017-04-19 | 王贤俊 | Latex immunoturbidimetric kit for detecting AGEs |
| CN106526198A (en) * | 2016-10-03 | 2017-03-22 | 王贤俊 | Combined orientation agents optimized latex coupled antibody detection method of prealbumin (PA) |
| WO2018119626A1 (en) * | 2016-12-27 | 2018-07-05 | 菲鹏生物股份有限公司 | Assay kit for neutrophil gelatinase-associated lipocalin |
| CN107340395A (en) * | 2017-07-05 | 2017-11-10 | 深圳开立生物医疗科技股份有限公司 | A kind of Immunoturbidimetric kit for detecting Procalcitonin |
| CN107167586A (en) * | 2017-07-06 | 2017-09-15 | 四川新健康成生物股份有限公司 | A kind of antistreptolysin O (ASO) detection kit and preparation method thereof |
| CN107543931B (en) * | 2017-09-12 | 2020-02-21 | 四川新健康成生物股份有限公司 | Kit for detecting TnI based on latex enhanced immunoturbidimetry and preparation method thereof |
| CN107543931A (en) * | 2017-09-12 | 2018-01-05 | 四川新健康成生物股份有限公司 | Kit based on latex enhancing immune turbidimetry detection TnI and preparation method thereof |
| CN107894509A (en) * | 2017-10-20 | 2018-04-10 | 柏荣诊断产品（上海）有限公司 | A kind of method for improving latex immunoturbidimetry antigen excess and the range of linearity |
| CN107991496A (en) * | 2017-11-29 | 2018-05-04 | 山东诺安爱迪尔生物工程有限公司 | A kind of neutrophil gelatinase-associated lipocalin detection kit |
| CN108362895B (en) * | 2018-02-12 | 2020-05-08 | 北京九强生物技术股份有限公司 | Folic acid detection kit and preparation method thereof |
| CN108362895A (en) * | 2018-02-12 | 2018-08-03 | 北京九强生物技术股份有限公司 | Folic acid detection kit and preparation method thereof |
| CN108414775A (en) * | 2018-03-02 | 2018-08-17 | 江西英大生物技术有限公司 | Neutrophil gelatinase-associated lipocalin detection kit |
| CN108896773A (en) * | 2018-08-23 | 2018-11-27 | 潍坊市康华生物技术有限公司 | A kind of gastrin 17 detection kit and its preparation and application |
| CN111721937A (en) * | 2019-03-29 | 2020-09-29 | 北京九强生物技术股份有限公司 | Human epididymis protein 4 immunoturbidimetry kit |
| CN110133247A (en) * | 2019-05-24 | 2019-08-16 | 深圳上泰生物工程有限公司 | A kind of immune latex turbidimetry method assay kit for testing goal antigen |
| CN110568182A (en) * | 2019-09-12 | 2019-12-13 | 苏州普瑞斯生物科技有限公司 | Adiponectin-latex enhanced immunoturbidimetry kit and preparation method thereof |
| CN110824176A (en) * | 2019-11-28 | 2020-02-21 | 江西英大生物技术有限公司 | Neutrophil gelatinase-associated lipocalin detection kit |
| CN113125699A (en) * | 2019-12-31 | 2021-07-16 | 科美诊断技术股份有限公司 | beta-hCG homogeneous phase chemiluminescence detection kit and application thereof |
| CN113125699B (en) * | 2019-12-31 | 2024-03-26 | 科美博阳诊断技术(上海)有限公司 | beta-hCG homogeneous phase chemiluminescence detection kit and application thereof |
| CN116413256A (en) * | 2023-02-13 | 2023-07-11 | 上海索昕生物科技有限公司 | Photosensitive microsphere for photoexcitation chemiluminescence detection |
| CN116413256B (en) * | 2023-02-13 | 2024-05-28 | 上海索昕生物科技有限公司 | Screening method of photosensitive microspheres for photo-excitation chemiluminescence detection |
Also Published As
| Publication number | Publication date |
|---|---|
| CN102590524B (en) | 2016-01-06 |
| WO2013097606A1 (en) | 2013-07-04 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102590524B (en) | Neutrophil gelatinase-associated lipocalin detection kit | |
| CN102628867B (en) | Double antibody latex intensified Retinal-binding protein detection kit | |
| CN104198732B (en) | A kind of neutrophil gelatinase-associated lipocalin reagent box for detecting content | |
| CN102590497B (en) | Cysteine protease inhibitor C test kit | |
| CN101699287B (en) | Homogeneous phase sol particle type cystatin C measuring kit and preparation method thereof | |
| CN106353507A (en) | Kit for detecting serum amyloid protein and application thereof | |
| CN103134934A (en) | Kit for simultaneously detecting retinol-binding protein (RBP) in urine sample and serum sample | |
| CN105510604A (en) | Method for improving sensitivity and linearity of latex reagent | |
| CN103389381B (en) | Human epididymal secretory protein E4 chemiluminescence detection kit and preparation method thereof | |
| CN101310186B (en) | Method of assaying antigen and kit to be used therein | |
| CN107402308A (en) | Immue quantitative detection reagent boxes of IL 6 and preparation method thereof | |
| CN102507918B (en) | Kit for determining anti-cyclic citrullinated peptide (Anti-CCP) antibody | |
| CN102944679A (en) | Kit for performing retinol binding protein detection by using latex turbidimetry | |
| CN103823070A (en) | Cystatin C determination kit with high sensitivity | |
| CN103018464A (en) | Reagent for determining procalcitonin and preparation method of reagent | |
| CN103308681B (en) | Trypsinogen-2 detection kit and preparation method | |
| CN107860929A (en) | The immunoturbidimetry detection reagent and method of a kind of serum amyloid A protein | |
| CN104614528A (en) | Wider linear range retinol binding protein determination kit | |
| CN104535770A (en) | Myoglobin determination kit of compound antibody | |
| CN104849473A (en) | Microalbuminuria detection kit and preparation thereof | |
| CN103645323A (en) | Cystatin C detection kit and preparation method therefor | |
| CN105765386A (en) | Competitive ligand binding assay for detecting neutralizing antibodies | |
| CN108051439A (en) | A kind of single reagent heparin-binding protein detection kit and preparation method thereof | |
| CN104360056A (en) | Method for improving sensitivity of latex enhanced turbidimetric immunoassay | |
| CN104820102A (en) | Kit of detecting the content of Lp-PLA2 and CRP on the basis of chemiluminescence, and method and application thereof |
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 |