CN102628868A - Latex enhanced immunoturbidimetry kit for detection of asymmetric dimethylarginine content - Google Patents

Abstract

The invention relates to a latex enhanced immunoturbidimetry kit for the detection of asymmetric dimethylarginine content. Specifically, the invention relates to a latex enhanced immunoturbidimetry kit for the detection of asymmetric dimethylarginine (ADMA) content in human blood samples. The kit comprises a mouse anti-human monoclonal antibody solution, an ADMA-human Serum albumin complex crosslinked latex particle expansion and an ADMA calibrator. According to the method, by the utilization of competitive binding of free ADMA in blood and ADMA crosslinked on the surface of latex particles to ADMA monoclonal antibody, turbidity formed by the reaction between the ADMA-crosslinked latex particles and ADMA monoclonal antibody is reduced. Therefore, the content of ADMA is detected through the reduction degree of turbidity. The kit provided by the invention can be applied in a biochemical analyzer commonly-used in clinic, is convenient and rapid to operate and has strong specificity. Both sensitivity and detection range of the kit can satisfy clinic application needs.

Description

Detect the latex enhancing immune turbidimetry kit of Ng,Ng-Dimethylarginine content

Technical field

The invention provides a kind of kit that utilizes the immunoturbidimetry method to measure Ng,Ng-Dimethylarginine content in human serum, the plasma sample; Be specifically related to utilize Ng,Ng-Dimethylarginine and the crosslinked latex suspension competition combination mouse-anti Ng,Ng-Dimethylarginine monoclonal antibody that Ng,Ng-Dimethylarginine-human serum albumins complex is arranged in human serum, the plasma sample; Through detecting the decline of immune turbidity; Reach the higher sensitivity and the sensing range of broad, belong to medical immunology in-vitro diagnosis field.

Background technology

(asymmetric diethylarginine ADMA) is a kind of natural amino acid to Ng,Ng-Dimethylarginine, extensively is present in tissue and the cell.It can suppress nitric oxide synthetase, and (nitric oxide synthase, biological effect synthetic and that produce therefrom NOS) stops the synthetic of endothelial cell nitric oxide (NO), causes function of vascular endothelium to reduce.The order of severity of vascular lesionses such as the variation of plasma ADM A concentration and function of vascular endothelium infringement, atherosclerotic, cerebral infarction, hypertension, high lipoprotein abnormalities, diabetes, renal failure is closely related, is one of emphasis problem of present clinical research.We can say that the concentration of plasma ADM A is the risk factor of the complete and angiocardiopathy of endothelial function.

ADMA is generated by arginine methyltransferase in vivo, and arginine methyltransferase has two types: the I type methylate histone and nuclear RNA-combination albumen generation NG-monomethyl-L-arginine (NG-monomethyl-L-arginine, L-NMMA) and ADMA; The II type only methylate the myelin major protein produce L-NMMA and SDMA (symmetricdimethylarginine, SDMA).L-NMMA and ADMA all can reduce the activity of NOS, but the former PC approximately is 1/20 of ADMA, and therefore, great majority research all is directed against the detection of ADMA.The concentration of SDMA in blood plasma is similar with ADMA, but NOS is not had effect.

The method of measuring ADMA in the blood plasma comprises high performance liquid chromatography, enzyme linked immunological and immunofluorescence etc.But the mensuration of ADMA is more loaded down with trivial details in the biological sample, because exist arginine and SDMA and ADMA in detection, to show cross reactivity.ADMA and arginic different two methyl that only are on the guanidine radicals nitrogen.In addition, arginic concentration approximately is 100 times of ADMA concentration in the normal human serum.SDMA more approaches ADMA than arginine, and difference only is the position of a methyl.In normal human serum, the concentration of SDMA is suitable with the concentration of ADMA, is approximately 1uM.

Moreover, all there are the characteristics that sample preparation is loaded down with trivial details, hand-manipulated, time-consuming is long in the method for above-mentioned detection ADMA.Therefore, invent ADMA in a kind of specific detection human plasma, the kit that can be applied to simultaneously on the self-reacting device becomes very need.

Summary of the invention

Therefore, technical purpose of the present invention is to provide a kind of Ng,Ng-Dimethylarginine detection kit that can be applied on the full-automatic biochemical analyzer, the Ng,Ng-Dimethylarginine content in can accurate detection human plasma sample.

Therefore, first aspect of the present invention relates to a kind of latex enhancing immune turbidimetry kit of measuring Ng,Ng-Dimethylarginine content, it is characterized in that, comprising:

1) Ng,Ng-Dimethylarginine R1 reagent;

2) Ng,Ng-Dimethylarginine R2 reagent;

3) Ng,Ng-Dimethylarginine calibration object;

Said Ng,Ng-Dimethylarginine R1 reagent comprises Ng,Ng-Dimethylarginine monoclonal antibody specific, damping fluid, stabilizing agent, set accelerator and antiseptic;

Said Ng,Ng-Dimethylarginine R2 reagent comprises latex particle, damping fluid, stabilizing agent and the antiseptic that Ng,Ng-Dimethylarginine-the inert protein complex encapsulates;

Said Ng,Ng-Dimethylarginine calibration object is made up of Ng,Ng-Dimethylarginine, damping fluid, stabilizing agent and antiseptic.

Preferably, said Ng,Ng-Dimethylarginine-inert protein complex is to obtain after with Ng,Ng-Dimethylarginine and inert protein coupling through coupling agent.Preferably, said coupling agent is selected from a kind of in carbodiimides (EDAC), glutaraldehyde, two isocyanic acid compounds and the dihalide dinitro benzene.

Preferably, described inert protein is selected from a kind of in human serum albumins, bovine serum albumin(BSA), ox transferrins, the hemocyanin.

Preferably, said latex particle is a kind of nucleocapsid structure, and breast nuclear is poly styrene polymer, and newborn shell is made up of styrene, n-butyl acrylate and methacrylic acid copolymer.

Preferably, said latex particle according to its surface with the chemical group difference can be selected from a kind of that carboxyl, amino, hydroxyl, hydrazide group, chloromethyl modify.

Preferably, said latex particle diameter range is 50-250nm, and working concentration is selected from 0.05-0.5%.

Preferably; Said Ng,Ng-Dimethylarginine-inert protein complex can be combined in the latex particle surface through chemical bonded refractory through the chemical group that carries with latex particle itself; The common chemical group comprises carboxyl, amino, hydroxyl, hydrazide group, chloromethyl etc.; With on the compound specific carboxyl that is combined in the latex particle surface of Ng,Ng-Dimethylarginine-inert protein, the Chemical Crosslinking Methods of said optimization may be summarized to be " two pH of a step " method through the Chemical Crosslinking Methods optimized in the present invention.

Preferably; Said damping fluid is selected from 3-[N; N-two (hydroxyethyl) amino]-in 2-hydroxy-propanesulfonic acid-sodium hydrate buffer solution, 4-(2-hydroxyethyl)-1-piperazine propane sulfonic acid-sodium hydrate buffer solution, trishydroxymethylaminomethane-HCl damping fluid, phosphate buffer, glycine buffer and the barbitol buffer solution one or more, working concentration is selected from 10-200mM.

Preferably, said stabilizing agent be selected from EDTA, the 0.1-1% concentration range of sodium chloride, the 2-50mM of bovine serum albumin(BSA), the 0.5-1% concentration range of 0.1-5% concentration range Tween-20,1-10% concentration range glycerine one or more.

Preferably, said antiseptic is selected from one or more in Sodium azide, thimerosal, phenol and the ethyl mercury sodium thiosulfate, and working concentration is selected from 0.02%-0.1%.

Preferably, said set accelerator is selected from a kind of in PEG-4000, PEG-6000, PEG-8000, the sodium dextran sulfate, and the working concentration scope is selected from 1-6%.

Most preferably, the latex enhancing immune turbidimetry kit of described mensuration Ng,Ng-Dimethylarginine content is made up of following component:

R1:0.2mg/mL ADMA monoclonal antibody, 0.9% sodium chloride, 5%PEG-6000,0.2%BSA, 20mM EDTA, 0.1% sodium azide, 50mM Tris-HCl pH of buffer 7.2;

R2: the polystyrene latex particle 0.25% that Ng,Ng-Dimethylarginine-human serum albumins complex encapsulates, 50mM glycine buffer pH8.0,0.9% sodium chloride, 0.8%BSA, 20mM EDTA, 0.1% polysorbas20,0.1% sodium azide, wherein said polystyrene latex particle surface have carboxyl, diameter is 150nm;

The Ng,Ng-Dimethylarginine calibration object: concentration is respectively 0,0.5,2,5,10, the Ng,Ng-Dimethylarginine of 20umol/L, 0.9% sodium chloride, 1%BSA, 50mM EDTA, 0.1% sodium azide, 50mM Tris-HCl pH of buffer 7.2.

In other words; The present invention is based on a specificity to the monoclonal antibody of ADMA and the latex particle suspending liquid of the crosslinked ADMA-of having human serum albumins; Antigen-antibody reaction taking place in specific reaction system form certain turbidity, can be detected based on spectrophotometric Biochemical Analyzer.And if have the ADMA of free state to exist in the human plasma, can combine monoclonal antibody with the ADMA competition that is fixed on the latex particle surface, make turbidity decline, come ADMA is carried out quantitative test through the reduction degree of system turbidity behind the assaying reaction.

The present invention has solved specificity and the low problem of automaticity that ADMA detects simultaneously, can wide popularization and application.

Particularly, to achieve these goals, the technical scheme that the present invention takes is: through encapsulating latex particle and be suspended in making R2 reagent in the specific damping fluid with the ADMA that is coupled on the inert protein.Be 100-200nm as the latex particle diameter range that encapsulates wherein, concentration is 1-5mg/mL.Because the ADMA molecular weight is very little, be difficult to combine with latex particle.Therefore be used as the ADMA that encapsulates among the present invention and at first be coupled on the inertia high molecular weight protein through chemical bond, the inert protein that molecular weight is bigger can better combine with latex particle, and it is 0.1-1mg/mL that coupling has the inert protein concentration of ADMA.Coupling has the inert protein of ADMA further to be coated on the latex particle surface through the Chemical Crosslinking Methods of optimizing; Used Chemical Crosslinking Methods is " two pH of a step " method; Be that latex particle at first is activated the agent activation under low pH condition; Again with high pH solution in ADMA-inert protein complex mix, mixed pH value of solution is neutral, this method can so that coupling have the inert protein of ADMA efficient, special with the latex particle surface combination.The buffer suspension liquid that is used as is the glycocoll-NaOH damping fluid that contains stabilizing agent and antiseptic, and concentration is 20-100mM, and the pH scope is 7-9.The stabilizing agent that is used as is that bovine serum albumin(BSA) 0.1-1%, sodium chloride concentration are that 0.7-0.9%, ethylenediamine tetraacetic acid concentration are that 5-50mM, polysorbas20 concentration are 0.1-1%.Used antiseptic is Sodium azide 0.05-0.1%.

Make R1 reagent through in specific damping fluid, adding specificity to the monoclonal antibody of ADMA, will add inorganic salts, set accelerator, protein and antiseptic in the specific damping fluid.Wherein selected monoclonal antibody is the antibody that specificity is directed against ADMA, can also can buy commercial product, for example Abcam, Mlilipore etc. through art-recognized hybridoma method preparation.Selected ADMA monoclonal antibody at first will detect through indirect ELISA method, confirms can not produce cross reaction with SDMA, L-NMMA and arginine.The surge capability that wherein requires damping fluid is for regulating the pH scope between 7-9; Can select various damping fluids; Preferred HEPES, glycocoll, Tris etc. compare with other damping fluids, have that surge capability is strong, solubleness is high, good biological fitness and a reactionlessness.Wherein Tris-HCl can the long period the constant pH scope of control, therefore more preferably Tirs-HCl, its concentration range is 10-100mM, the pH scope is 7-8.Set accelerator is selected PEG-6000, can accelerate immune response speed, shortens detection time, and its final concentration is 2-6%.Protein is selected bovine serum albumin(BSA), and final concentration is 0.1-1%.Sodium chloride concentration is 0.7-0.9%, and Sodium azide concentration is 0.05-0.1%.

Be respectively 0,0.5,2,5,10 through in specific damping fluid, adding ADMA concentration, 20umol/L makes the multiple spot calibration object; The surge capability that wherein requires damping fluid is for regulating the pH scope between 7-9; Can select various damping fluids; Preferred HEPES, glycocoll, Tris etc. compare with other damping fluids, have that surge capability is strong, solubleness is high, good biological fitness and a reactionlessness.Wherein Tris-HCl can the long period the constant pH scope of control, therefore more preferably Tirs-HCl, its concentration range is 10-200mM, the pH scope is 7-8.Wherein contain specific protein stabiliser such as bovine serum albumin(BSA) concentration is 0.1-1%, contain specific antiseptic such as Sodium azide concentration is 0.05-0.1%, containing specific anti-oxidant such as ethylenediamine tetraacetic acid concentration is 5-50mM.

The present invention is provided for measuring the Ng,Ng-Dimethylarginine immunoturbidimetry assay method in human plasma, blood serum sample; It is characterized in that; Through human plasma, the serum sample that will contain Ng,Ng-Dimethylarginine; Compete the monoclonal antibody of binding specificity with the Ng,Ng-Dimethylarginine that latex particle encapsulates to ADMA; Cause turbidity to descend, the degree of decline is directly proportional with Ng,Ng-Dimethylarginine content in human plasma, the serum, and employing automatic clinical chemistry analyzer device can calculate the content of Ng,Ng-Dimethylarginine in human plasma, the serum sample.

The present invention compared with prior art has following characteristics:

1) can be applied on the automated chemical analyser, detection speed is fast;

2) detection specificity is strong, does not produce cross reaction with L-NMMA, SDMA and arginine, has very high correlativity with the HPLC method;

3) detection sensitivity and sensing range reach other just at the kit peer-level of clinical practice, can satisfy the clinical practice needs.

Description of drawings

Fig. 1 is the linear graph of the prepared kit of the present invention.

Fig. 2 detects the correlativity of clinical sample comparison for prepared kit of the present invention and HPLC method kit.

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.

Following experimental technique is conventional method if no special instructions, and employed experiment material all can easily be obtained from commercial company if no special instructions.

Embodiment

Embodiment 1: the human serum albumins of preparation coupling Ng,Ng-Dimethylarginine

With the 10mg human serum albumins with after 1mL 0.1M phosphate buffer (pH7.8) dilution; Add Ng,Ng-Dimethylarginine; Add 50mg EDAC again; 4 ℃ of reactions 24 hours, use 0.1M phosphate buffer (pH7.8) under 4 ℃ of conditions, to dialyse 24 hours again, be made into the human serum albumins (Ng,Ng-Dimethylarginine-human serum albumins complex) of coupling ADMA.

Embodiment 2: preparation Ng,Ng-Dimethylarginine R2 reagent

The surface had polystyrene latex solution (concentration 10%) (available from the Merck) 0.5mL of carboxyl, diameter 150nm, adds the 0.05M MES damping fluid (pH6.0) of 4.5mL, adds 5mg EDAC then, 37 ℃ of reactions 1 hour.After having the human serum albumins of ADMA to dilute the 0.5mg coupling again, join immediately in the above-mentioned latex solution, 37 ℃ of reactions 6 hours with 5mL 0.05M borate buffer solution (pH9.2).The glycine buffer (pH8.5) that adds 1mL 0.1M at last stirs 1h and comes cessation reaction; The centrifugal supernatant that removes; Glycine buffer (pH8.0) with 20mL 50mM washs 3 times; Contain 0.9% sodium chloride, 20mM EDTA, 0.8%BSA, 0.1% polysorbas20,0.1% sodium azide in the glycine buffer of 50mM; Last be dispersed into the milky latex suspension with the glycine buffer of same 20mL again, be R2 reagent, the polystyrene latex granule density that Ng,Ng-Dimethylarginine-human serum albumins complex encapsulates in the final R2 reagent is 0.25%.

Embodiment 3: preparation Ng,Ng-Dimethylarginine R1 reagent

In the Tris-HCl of 50mM pH7.2 damping fluid, add the ADMA monoclonal antibody (available from Abcam) of 0.2mg/mL, add sodium chloride 0.9%, PEG-60005%, BSA 0.2%, EDTA20mM, sodium azide 0.1% simultaneously, stirring is R1 reagent.

Embodiment 4: preparation Ng,Ng-Dimethylarginine calibration object

In the Tris-HCl of 50mM pH7.2 damping fluid; Add that concentration is respectively 0,0.5,2,5,10, the Ng,Ng-Dimethylarginine of 20umol/L; Add sodium chloride 0.9%, BSA1%, EDTA 50mM, sodium azide 0.1% in addition, stirring is Ng,Ng-Dimethylarginine multiple spot calibration object.

Embodiment 5: the Performance Detection of Ng,Ng-Dimethylarginine agent box

1. the usage of detection kit:

With Hitachi's 7180 Biochemical Analyzers is example: measure wavelength 570nm, at first add R1 reagent 150uL, 37 ℃ of reactions add ADMA calibration object 10uL after 30 seconds; React again and add R2 reagent 50uL after 60 seconds, the 10th second, 70 seconds absorbance of assaying reaction (A1, A2), calculating absorbance difference Δ A=A2-A1; Every pipe replication 2 times, the absorbance difference Δ A that each calibration QC is recorded for 2 times is an ordinate, corresponding concentration is horizontal ordinate; Make " concentration-absorbance difference " calibration curve, get sample to be tested, measure the absorbance difference of sample with method; The substitution calibration curve can calculate the content of ADMA in the sample to be tested.

2. repeatability detects:

With physiological saline dilution human serum sample; Compound concentration is respectively 0.1,0.5,1, the sample of 2umol/L; With the kit of above-mentioned Ng,Ng-Dimethylarginine R1, Ng,Ng-Dimethylarginine R2 and Ng,Ng-Dimethylarginine calibration object composition, with the heavier renaturation ability of ELISA reagent h box of A company.The reagent fundamental analysis principle of A company is; Use acylting agent with the ADMA acylated in the sample earlier; Add ELISA Plate then, be combined in solid phase on the competitive ADMA polyclonal antibody that combines of ADMA, adopt the standard items of varying level to compete and combine with enzymic-labelled antibody; Absorbance, production standard curve are read in colour developing then.After the same procedure of sample and standard items, on typical curve, obtain corresponding A MA concentration.Kit of the present invention is measured 10 times according to 1 described method, calculates the coefficient of variation of different ADMA content pattern detection, with A company commercial ELISA kit comparative result shown in following table 1 and 2:

The detection repeatability of table 1 kit of the present invention:

	0.1uM	0.5uM	1uM	2uM
					1	0.14	0.52	1.01	2.12
2	0.13	0.47	1.02	2.18
					3	0.18	0.48	1.11	2.07
4	0.14	0.49	1.07	2.09
					5	0.15	0.45	1.08	2.08
6	0.12	0.46	1.04	2.1
					7	0.17	0.49	0.98	2.12
8	0.19	0.55	1.01	2.04
					9	0.12	0.58	1.02	2.08
10	0.17	0.47	1.07	2.09
					Mean value	0.151	0.496	1.041	2.097
SD	0.025144	0.0416867	0.0401248	0.0374314
					CV	16.7％	8.4％	3.9％	1.8％

The detection repeatability of the ELISA kit of table 2, A company:

	0.1uM	0.5uM	1uM	2uM
						1	0.08	0.50	1.08	2.09
2	0.07	0.54	1.05	2.18
					3	0.19	0.45	1.11	2.07
4	0.14	0.44	1.07	2.11
					5	0.11	0.51	0.97	2.08
6	0.12	0.43	1.04	2.02
					7	0.17	0.48	0.98	2.12
8	0.14	0.52	1.04	2.02
					9	0.11	0.55	1.07	2.08
10	0.15	0.42	1.03	2.09
					Mean value	0.128	0.484	1.044	2.086
SD	0.0376534	0.0469515	0.0432563	0.0467143
					CV	29.42％	9.70％	4.14％	2.24％

Can find out that from last table 1 and 2 repeatability of kit of the present invention is better than the ELISA kit of A company, more helps the application of clinical diagnosis.

3. linearity test:

Screening Ng,Ng-Dimethylarginine concentration is about the sample of 20umol/L; Do the dilution of 10 different proportions with physiological saline, detect each concentration, every concentration replication 3 times according to 1 described method; Mean value and the theoretical concentration of measuring concentration are carried out recovery analysis; Result error proves that all less than 10% linearity can reach 20umol/L, and the result sees table 3 and Fig. 1.

The linearity of table 3 kit of the present invention

Dilution ratio	Theoretical concentration [um/L]	Measure concentration [um/L]	The recovery [%]
				0/10	0.0	0.0
1/10	2.00	2.07	101.0％
				2/10	4.00	4.12	104.0％
3/10	6.00	6.04	103.3％
				4/10	8.00	8.08	101.3％
5/10	10.00	10.13	104.2％
				6/10	12.00	12.07	101.7％
7/10	14.00	14.11	100.6％
				8/10	16.00	16.08	99.8％
9/10	18.00	18.13	100.2％
				10/10	20.00	20.02	104.0％

4. the correlativity that kit of the present invention and HPLC detect clinical sample compares

With kit of the present invention and HPLC 34 routine patient's sample are detected simultaneously, testing result is seen Fig. 2, and the sample ADMA result who detects with the HPLC method is a horizontal ordinate; The result who detects with kit of the present invention is that ordinate is done regretional analysis; Dependent equation is Y=0.9965X+0.018, and correlation coefficient r=0.999 shows through the statistical procedures result; It is good that the inventive method detects clinical sample measured value correlativity with the HPLC method, shows that the detection specificity of kit of the present invention is strong.

Claims (11)

1. a latex enhancing immune turbidimetry kit of measuring Ng,Ng-Dimethylarginine content is characterized in that, comprising:

1) Ng,Ng-Dimethylarginine R1 reagent;

2) Ng,Ng-Dimethylarginine R2 reagent;

3) Ng,Ng-Dimethylarginine calibration object;

Said Ng,Ng-Dimethylarginine R1 reagent comprises Ng,Ng-Dimethylarginine monoclonal antibody specific, damping fluid, stabilizing agent, set accelerator and antiseptic;

Said Ng,Ng-Dimethylarginine R2 reagent comprises latex particle, damping fluid, stabilizing agent and the antiseptic that Ng,Ng-Dimethylarginine-the inert protein complex encapsulates;

Said Ng,Ng-Dimethylarginine calibration object is made up of Ng,Ng-Dimethylarginine, damping fluid, stabilizing agent and antiseptic.

2. the latex enhancing immune turbidimetry kit of mensuration Ng,Ng-Dimethylarginine content according to claim 1; It is characterized in that said Ng,Ng-Dimethylarginine-inert protein complex is to obtain after with Ng,Ng-Dimethylarginine and inert protein coupling through coupling agent.

3. the latex enhancing immune turbidimetry kit of mensuration Ng,Ng-Dimethylarginine content according to claim 1 and 2; It is characterized in that described inert protein is selected a kind of in white man's seralbumin, bovine serum albumin(BSA), ox transferrins and the hemocyanin.

4. according to the latex enhancing immune turbidimetry kit of each described mensuration Ng,Ng-Dimethylarginine content of claim 1 to 3; It is characterized in that; Said latex particle is a kind of nucleocapsid structure; Breast nuclear is poly styrene polymer, and newborn shell is made up of styrene, n-butyl acrylate and methacrylic acid copolymer.

5. according to the latex enhancing immune turbidimetry kit of each described mensuration Ng,Ng-Dimethylarginine content of claim 1 to 4, it is characterized in that said latex particle diameter range is 50-250nm, working concentration is selected from 0.05-0.5%.

6. according to the latex enhancing immune turbidimetry kit of each described mensuration Ng,Ng-Dimethylarginine content of claim 1 to 5; It is characterized in that; Said Ng,Ng-Dimethylarginine-inert protein complex can be combined in the latex particle surface through chemical bonded refractory through the chemical group that carries with latex particle itself; The common chemical group comprises carboxyl, amino, hydroxyl, hydrazide group, chloromethyl etc.; With on the compound specific carboxyl that is combined in the latex particle surface of Ng,Ng-Dimethylarginine-inert protein, the Chemical Crosslinking Methods of said optimization is summarised as " two pH of a step " method through the Chemical Crosslinking Methods optimized in the present invention.

7. according to the latex enhancing immune turbidimetry kit of each described mensuration Ng,Ng-Dimethylarginine content of claim 1 to 6; It is characterized in that; Said damping fluid is selected from 3-[N; N-two (hydroxyethyl) amino]-in 2-hydroxy-propanesulfonic acid-sodium hydrate buffer solution, 4-(2-hydroxyethyl)-1-piperazine propane sulfonic acid-sodium hydrate buffer solution, trishydroxymethylaminomethane-HCl damping fluid, phosphate buffer, glycine buffer and the barbitol buffer solution one or more, working concentration is selected from 10-200mM.

8. according to the latex enhancing immune turbidimetry kit of each described mensuration Ng,Ng-Dimethylarginine content of claim 1 to 7; It is characterized in that, said stabilizing agent be selected from sodium chloride, the 2-50mM of bovine serum albumin(BSA), the 0.5-1% concentration range of 0.1-5% concentration range EDTA, 0.1-1% concentration range Tween-20 and 1-10% concentration range glycerine one or more.

9. according to the latex enhancing immune turbidimetry kit of each described mensuration Ng,Ng-Dimethylarginine content of claim 1 to 8; It is characterized in that; Said antiseptic is selected from one or more in Sodium azide, thimerosal, phenol and the ethyl mercury sodium thiosulfate, and working concentration is selected from 0.02%-0.1%.

10. according to the latex enhancing immune turbidimetry kit of each described mensuration Ng,Ng-Dimethylarginine content of claim 1 to 9; It is characterized in that; Said set accelerator is selected from a kind of in PEG-4000, PEG-6000, PEG-g000 and the sodium dextran sulfate, and the working concentration scope is selected from 1-6%.

11. the latex enhancing immune turbidimetry kit according to each described mensuration Ng,Ng-Dimethylarginine content of claim 1 to 10 is characterized in that it and is made up of following component:

R1:0.2mg/mL ADMA monoclonal antibody, 0.9% sodium chloride, 5%PEG-6000,0.2%BSA, 20mM EDTA, 0.1% sodium azide, 50mM Tris-HCl pH of buffer 7.2;

R2: the polystyrene latex particle 0.25% that Ng,Ng-Dimethylarginine-the human serum albumins complex encapsulates, the glycine buffer pH8.0 of 50mM, 0.9% sodium chloride, 0.8%BSA, 20mM EDTA, 0.1% polysorbas20,0.1% sodium azide, wherein said polystyrene latex particle surface have carboxyl, diameter is 150nm;

The Ng,Ng-Dimethylarginine calibration object: concentration is respectively 0,0.5,2,5,10, the Ng,Ng-Dimethylarginine of 20umol/L, 0.9% sodium chloride, 1%BSA, 50mM EDTA, 0.1% sodium azide, 50mM Tris-HCl pH of buffer 7.2.

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