CN102323427B9 - A kind of kit and method thereof that detects concentration of complement Clq in human serum - Google Patents
A kind of kit and method thereof that detects concentration of complement Clq in human serum Download PDFInfo
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- CN102323427B9 CN102323427B9 CN201110226543.1A CN201110226543A CN102323427B9 CN 102323427 B9 CN102323427 B9 CN 102323427B9 CN 201110226543 A CN201110226543 A CN 201110226543A CN 102323427 B9 CN102323427 B9 CN 102323427B9
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- 230000000295 complement Effects 0.000 title claims abstract description 27
- 210000002966 Serum Anatomy 0.000 title claims abstract description 15
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 37
- 229920004890 Triton X-100 Polymers 0.000 claims abstract description 14
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 claims abstract description 12
- 229910000397 disodium phosphate Inorganic materials 0.000 claims abstract description 12
- 235000019800 disodium phosphate Nutrition 0.000 claims abstract description 12
- GNSKLFRGEWLPPA-UHFFFAOYSA-M Monopotassium phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 claims abstract description 10
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims abstract description 10
- 235000019796 monopotassium phosphate Nutrition 0.000 claims abstract description 10
- 241000283973 Oryctolagus cuniculus Species 0.000 claims abstract description 9
- 239000000203 mixture Substances 0.000 claims abstract description 7
- 229920002584 Polyethylene Glycol 6000 Polymers 0.000 claims abstract 6
- 230000035484 reaction time Effects 0.000 claims description 3
- 239000011734 sodium Substances 0.000 claims 2
- ZGTMUACCHSMWAC-UHFFFAOYSA-L disodium;2-[2-[carboxylatomethyl(carboxymethyl)amino]ethyl-(carboxymethyl)amino]acetate Chemical compound [Na+].[Na+].OC(=O)CN(CC([O-])=O)CCN(CC(O)=O)CC([O-])=O ZGTMUACCHSMWAC-UHFFFAOYSA-L 0.000 claims 1
- 238000004879 turbidimetry Methods 0.000 abstract description 6
- 238000004458 analytical method Methods 0.000 abstract description 4
- 238000002965 ELISA Methods 0.000 abstract description 3
- 238000002835 absorbance Methods 0.000 description 8
- 230000033228 biological regulation Effects 0.000 description 8
- LYCAIKOWRPUZTN-UHFFFAOYSA-N glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 4
- 239000007788 liquid Substances 0.000 description 4
- 238000000034 method Methods 0.000 description 3
- PIICEJLVQHRZGT-UHFFFAOYSA-N 1,2-ethanediamine Chemical compound NCCN PIICEJLVQHRZGT-UHFFFAOYSA-N 0.000 description 2
- 238000005352 clarification Methods 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- QXNVGIXVLWOKEQ-UHFFFAOYSA-N disodium Chemical compound [Na][Na] QXNVGIXVLWOKEQ-UHFFFAOYSA-N 0.000 description 2
- UFHFLCQGNIYNRP-UHFFFAOYSA-N hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 2
- 230000036039 immunity Effects 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 2
- 229910052700 potassium Inorganic materials 0.000 description 2
- 239000011591 potassium Substances 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 238000010998 test method Methods 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 108010047814 Antigen-Antibody Complex Proteins 0.000 description 1
- 210000001268 Chyle Anatomy 0.000 description 1
- 102000008186 Collagen Human genes 0.000 description 1
- 108010035532 Collagen Proteins 0.000 description 1
- 239000003109 Disodium ethylene diamine tetraacetate Substances 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 206010018910 Haemolysis Diseases 0.000 description 1
- 206010018913 Haemolysis Diseases 0.000 description 1
- LLJZKKVYXXDWTB-UHFFFAOYSA-N acetic acid;sodium Chemical compound [Na].[Na].CC(O)=O LLJZKKVYXXDWTB-UHFFFAOYSA-N 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 108090001123 antibodies Proteins 0.000 description 1
- 102000004965 antibodies Human genes 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 102000038129 antigens Human genes 0.000 description 1
- 108091007172 antigens Proteins 0.000 description 1
- 238000004166 bioassay Methods 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 230000004154 complement system Effects 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 235000019301 disodium ethylene diamine tetraacetate Nutrition 0.000 description 1
- 231100000673 dose–response relationship Toxicity 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N edta Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 238000011017 operating method Methods 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 239000000376 reactant Substances 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000002834 transmittance Methods 0.000 description 1
- 238000005429 turbidity Methods 0.000 description 1
- 230000000007 visual effect Effects 0.000 description 1
Abstract
The invention belongs to bioengineering field, provide a kind of immune turbidimetry and immune scattering turbidimetry to detect the kit of concentration of complement Clq in human serum, solve available technology adopting SRID and ELISA double-antibody sandwich technology and measured C1Q concentration step complexity, accuracy is not high, the technical problem of poor repeatability, comprise two kinds of reagent, the first reagent is by sodium hydrogen phosphate, potassium dihydrogen phosphate, PEG6000, EDTA-NA2With TX-100 composition, the second reagent is by sodium hydrogen phosphate, potassium dihydrogen phosphate, PEG6000, EDTA-NA2, TX-100 and the anti-human C1Q antiserum of rabbit composition. The present invention also provides the method that adopts above-mentioned kit to detect concentration of complement Clq in human serum, and kit of the present invention and method are simple and convenient by the step of measuring C1Q concentration, and accuracy is high, reproducible, for automated analysis instrument.
Description
Technical field
The invention belongs to bioengineering field, relate in particular to a kind of detection kit, particularly a kind of immune transmissionTurbidimetry and immune scattering turbidimetry detect complement Clq concentration in human serum and in arthral fluid kit andIts method.
Background technology
Complement Clq is the first composition of complement system Cl, is a giant molecule amount glycoprotein, a ClqMolecule is become by 18 polypeptide chains, and chemical composition is collagen protein molecular weight: 410KD. Available technology adoptingSRID and ELISA double-antibody sandwich technology are measured complement Clq concentration, but its step complexity, accuratelyProperty is not high.
Summary of the invention
The object of the present invention is to provide a kind of kit and method thereof that detects concentration of complement Clq in human serum,The kit of described this detection concentration of complement Clq in human serum and method thereof will solve in prior art adoptsMeasure complement Clq concentration process complexity by SRID and ELISA double-antibody sandwich technology, accuracy is not highTechnical problem.
The invention provides a kind of kit that detects concentration of complement Clq in human serum, comprise two kinds of reagent, theA kind of reagent is by sodium hydrogen phosphate, potassium dihydrogen phosphate, Macrogol 6000 (PEG6000), ethylenediamine tetremAcid disodium (EDTA-NA2) and Triton X-100 (TX-100) composition, in described the first reagent, instituteThe mass percent concentration of the sodium hydrogen phosphate of stating is 28.6g/L, the mass percent of described potassium dihydrogen phosphateConcentration is 2.7g/L, and the mass percent concentration of described Macrogol 6000 is 50g/L, described second twoThe mass percent concentration of amine tetraacethyl disodium is 1g/L, and the volumetric concentration of described Triton X-100 is0.2ml/L, the second reagent is by sodium hydrogen phosphate, potassium dihydrogen phosphate, Macrogol 6000, ethylenediamine tetraaceticThe anti-human complement Clq of acetic acid disodium and Triton X-100 and rabbit antiserum composition, in described the second reagent,The mass percent concentration of described sodium hydrogen phosphate is 28.6g/L, the quality percentage of described potassium dihydrogen phosphateSpecific concentration is 2.7g/L, and the mass percent concentration of described Macrogol 6000 is 50g/L, describedThe mass percent concentration of disodium ethylene diamine tetraacetate is 1g/L, and the volumetric concentration of described Triton X-100 is0.2ml/L, the sero-fast volumetric concentration of the anti-human complement Clq of described rabbit is 300ml/L.
Concrete, the purity of above-mentioned Triton X-100 is 100%, anti-human complement Clq is sero-fast for above-mentioned rabbitTire 1: 64.
The present invention also provides a kind of method that detects concentration of complement Clq in human serum, adopts above-mentioned reagentBox, comprises a step that adopts Biochemical Analyzer to measure in sample, sample is adopted biochemical at described oneIn the step of analysis-e/or determining, sample is added after the first reagent R1, hatch 5min for 37 DEG C, read to survey the 1stPoint sample A1, adds the second reagent R2, hatches 5min for 37 DEG C, reads to survey the 2nd sample A2, sample A=Sample A2-sample A1; Also comprise a step that adopts Biochemical Analyzer to measure calibration object, described oneIn the individual step that calibration is adopted to Biochemical Analyzer mensuration, calibration is added after the first reagent R1, incubate for 37 DEG CEducate 5min, read to survey the 1st calibration A1, add the second reagent R2, hatch 5min for 37 DEG C, read to survey the 2ndPoint calibration A2, calibration A=calibration A2-calibration A1;
Result is calculated:
The parameter of said determination is: 37 DEG C of temperature; Dominant wavelength 340nm, commplementary wave length 700nm, sample or calibrationProduct 4 μ l; R1：240μl；R2: 60 μ l, 10 minutes reaction time.
The anti-human complement Clq of rabbit of the present invention antiserum can be bought in market, also can be by conventional immunity sideLegal system is standby.
Kit of the present invention and method are to adopt the mensuration of immune turbidimetry and immune scattering turbidimetry formerReason, utilizes complement Clq antigen and specific antibody (the anti-human complement Clq of rabbit antiserum) to combine, and formsInsolubilized immune complexes, produces reactant liquor muddy, its turbidity height, i.e. and light transmittance minimizing, absorbance increaseThe concentration that adds complement Clq in reflection human serum sample, the dosage that the concentration of complement Clq can be done by calibration object is anti-Answer curve to calculate.
Adopt the concentration of the CLq that kit of the present invention and method record to can be used as scientific research and teaching is used.
The present invention compares with prior art, and its technological progress is significant. Kit of the present invention and method are used forThe process of measuring complement Clq concentration is simple and convenient, and accuracy is high.
Brief description of the drawings
Fig. 1 is the complement Clq concentration done by calibration object and the dose-effect curve figure of absorbance.
Detailed description of the invention
Embodiment 1
1.1 kit specifications
1.2 component
R1: sodium hydrogen phosphate 28.6g/L potassium dihydrogen phosphate 2.7g/L
PEG6000 50g/L EDTA-NA2 1g/L
TX-100 0.2ml/L
R2: sodium hydrogen phosphate 28.6g/L potassium dihydrogen phosphate 2.7g/L
PEG6000 50g/L EDTA-NA2 1g/L
The anti-human complement Clq of TX-100 0.2ml/L rabbit antiserum 300ml/L
1.3 are suitable for instrument
Semi-automatic, automatic clinical chemistry analyzer.
1.4 analytical method
Immunity turbidimetry.
1.5 sample requirements
End user's serum, 2-8 DEG C of preservation, detected in 24 hours, as do not detected within 24 hours, putPreserve April in-20 DEG C, haemolysis and serious piarhemia (as chyle shape) sample can not use.
1.6 performance requirement
1.6.1 reagent outward appearance
R1: achromaticity and clarification transparency liquid;
R2: faint yellow supernatant liquid.
1.6.2 reagent blank absorbance (A)
Absorbance (A): R1+R2≤ 0.04A is (37 DEG C of temperature; Dominant wavelength 340nm (commplementary wave length 700nm)).
1.6.3 precision
1.6.3.1 withinrun precision
CV≤10％。
1.6.4 betweenrun precision
Extreme difference≤10% relatively.
1.6.5 the degree of accuracy
Inaccuracy: in ± 10% scope.
1.6.6 sensitivity for analysis
Absorbance (A) > 0.04A.
1.6.7 linear
Within the scope of 50mg/L-400mg/L, coefficient correlation (γ) >=0.9900.
1.6.8 stability
Reagent, detects to effective former and later two months in the end of term 2 DEG C of-8 DEG C of lucifuges storages, and its quality meetsThe regulation of item.
Embodiment 2 experimental techniques
2.1 testing conditions
2.1.1 Biochemical Analyzer (hereinafter to be referred as instrument)
Hitachi's 7060 automatic biochemistry analyzers.
2.1.2 operating ambient temperature
Room temperature 15-32 DEG C.
Indoor humidity 45-85%RH.
2.1.3 main location parameter and operating procedure
37 DEG C of temperature; Dominant wavelength 340nm (commplementary wave length 700nm), sample or calibration object 4 μ l; R1：240μ
l；R2: 60 μ l, 10 minutes reaction time.
Method type: 2 end-point methods.
After sample or calibration object reagent adding R1, hatch 5min for 37 DEG C, read to survey the 1st point (A1), add reagent R2,Hatch 5min for 37 DEG C, read to survey the 2nd point (A2), sample A=A2-A1.
Result is calculated:
2.1.4 calibration object
In this standard, calibration object used is the standard items that south, Yuhuan county of Zhejiang Province chemical reagent work produces.
2.2 reagent outward appearances
Visual observation agent box under light, R1: achromaticity and clarification transparency liquid; R2: faint yellow supernatant liquid.
2.3 reagent blank absorbances
Get R1240 μ l, adding distil water 4 μ l, put 37 DEG C and hatch after 5min, add R260 μ l, put 37DEG C hatch after 5min, on 7060 automatic biochemistry analyzers, use 340nm wavelength, measuring its absorbance shouldMeet the regulation of 1.6.2 item in embodiment 1.
2.4 precision
2.4.1 withinrun precision test method
Under instrument normal running conditions, use with a collection of reagent follow-on test (about 200mg/L) sample 20Inferior, calculate the mean value of its measured valueAnd standard deviation (SD), then calculate as follows the coefficient of variation (CV%)Value, its result should meet the regulation of 1.6.3.1 item in embodiment 1.
n
SD＝{∑(Xi-X)2/(N-1)}1/2
I＝1
In formula: SD-standard deviation;
The CV-coefficient of variation;
The average of-n time measured value;
XiThe measured value that-is i time;
N-measures number of times.
2.4.2 betweenrun precision test method
Under instrument normal running conditions, get three lot number reagent, each lot number is got a set of. Measure (approximately respectively200mg/L) each 3 times of sample. Then calculate the average of every batch of measurement resultSurvey with three batches of reagentDetermine the grand mean of resultObtain according to the following formula the antipode of the mensuration average of three lot number reagentPoor (%), its result should meet the regulation of 1.6.4 item in embodiment 1.
In formula:ForMiddle maximum,ForMiddle minimum of a value.
2.5 the degree of accuracy
Under instrument normal running conditions, the standard items that reagent is produced with Zhejiang Yuhuan south chemical reagent work are through calibrationAfter, the sample of mensuration concentration known, replication 10 times, the relative deviation of the average of its measurement result should accord withClose the regulation of 1.6.5 item in embodiment 1.
RE%=absolute deviation/TV × 100%
Absolute deviation=testing result average-standard items target value
In formula: the target value of TV-bioassay standard product
2.6 sensitivity for analysis
R1Reagent 240 μ l, add 50mg/L sample 4 μ l, put 37 DEG C and hatch after 5min, add R260μL, puts 37 DEG C and hatches after 5min, on 7060 automatic biochemistry analyzers, with 340nm dominant wavelength (commplementary wave length700nm), survey absorbance, its result should meet the regulation of 1.6.6 item in embodiment 1.
2.7 linear test
Sample or calibration object: select prepare by embodiment 31#、2#、3#、4#, and 5#Each test concentrations sampleBasis or calibration object.
Determination step: under instrument normal running conditions, after calibration, five concentration samples more than measuring with reagentBasis or calibration object, each concentration samples or calibration object are measured respectively 3 times, and getting average is measured value Yi.
Calculate: calculate as follows coefficient correlation γ, its result should meet the rule of 1.6.7 item in embodiment 1Fixed.
In formula: γ-coefficient correlation
Xi-i#The theoretical value of sample
Yi-i#The measured value of sample
The numbering of i-variable concentrations test sample book
5.8 stability
Be taken at and under regulation storage requirement, be saved to the reagent in two months before and after effective end of term and detect, its resultShould meet the regulation of 1.6.8 item in embodiment 1.
The preparation of embodiment 3 reagent
Getting the sample that a content is about 200mg/L, is sample 4#, by sample 4#According to the form below method physiological salineOr pure water is mixed with 5 test sample books. (this sample matching while using)
| 1# | 2# | 3# | 4# | 5# | |
| Calibration object | - | 4μl | 2μl | 4μl | 8μl |
| Physiological saline | 4μl | 12μl | - | - | - |
| Sampling amount after dilution | - | 4μl | - | - | - |
| Concentration (mg/L) | 0 | 50 | 100 | 200 | 400 |
After sample or calibration object reagent adding R1, hatch 5min for 37 DEG C, read to survey the 1st point (A1), add reagent R2,Hatch 5min for 37 DEG C, read to survey the 2nd point (A2), sample A=A2-A1.
Result instrument calculates:
By the mensuration of above-mentioned sample, the complement Clq concentration shown in Fig. 1 and the dose response of absorbance are obtainedCurve map.
(Fig. 1 is the curve doing according to the OD value of wavelength 340nm, the use only for reference of wavelength 700nm. )
By Fig. 1, measure 500 routine normal persons' serum, result is described as following table, cures political affairs according to the Ministry of Public HealthComplement Clq normal reference value in serum of introducing in department's " national clinical examination code ": 157-237mg/L,Illustrate adopt the Clq that kit of the present invention and method measure concentration still accurately and reliably.
500 routine Healthy People concentration of complement Clq in human serums (mg/L)
| Number | Numerical value | Number | Numerical value | Number | Numerical value | Number | Numerical value | Number | Numerical value | Number | Numerical value |
| 1 | 179 | 9 | 196 | 17 | 196 | 25 | 221 | 33 | 206 | 41 | 195 |
| 2 | 174 | 10 | 199 | 18 | 178 | 26 | 205 | 34 | 195 | 42 | 205 |
| 3 | 166 | 11 | 201 | 19 | 221 | 27 | 192 | 35 | 206 | 43 | 200 |
| 4 | 198 | 12 | 193 | 20 | 207 | 28 | 196 | 36 | 182 | 44 | 198 |
| 5 | 206 | 13 | 194 | 21 | 197 | 29 | 213 | 37 | 198 | 45 | 215 |
| 6 | 195 | 14 | 203 | 22 | 194 | 30 | 206 | 38 | 196 | 46 | 161 |
| 7 | 216 | 15 | 221 | 23 | 184 | 31 | 191 | 39 | 194 | 47 | 180 |
| 8 | 194 | 16 | 196 | 24 | 207 | 32 | 192 | 40 | 226 | 48 | 196 |
| 49 | 205 | 78 | 195 | 107 | 196 | 136 | 206 | 165 | 196 | 194 | 193 |
| 50 | 200 | 79 | 194 | 108 | 194 | 137 | 200 | 166 | 192 | 195 | 203 |
| 51 | 160 | 80 | 206 | 109 | 196 | 138 | 192 | 167 | 194 | 196 | 191 |
| 52 | 194 | 81 | 187 | 110 | 199 | 139 | 196 | 168 | 195 | 197 | 200 |
| 53 | 206 | 82 | 198 | 111 | 199 | 140 | 197 | 169 | 217 | 198 | 198 |
| 54 | 184 | 83 | 225 | 112 | 200 | 141 | 195 | 170 | 198 | 199 | 183 |
| 55 | 163 | 84 | 217 | 113 | 196 | 142 | 196 | 171 | 196 | 200 | 226 |
| 56 | 206 | 85 | 193 | 114 | 195 | 143 | 198 | 172 | 193 | 201 | 195 |
| 57 | 207 | 86 | 195 | 115 | 196 | 144 | 199 | 173 | 188 | 202 | 198 |
| 58 | 205 | 87 | 204 | 116 | 194 | 145 | 232 | 174 | 198 | 203 | 194 |
| 59 | 198 | 88 | 196 | 117 | 196 | 146 | 193 | 175 | 195 | 204 | 206 |
| 60 | 195 | 89 | 198 | 118 | 201 | 147 | 194 | 176 | 194 | 205 | 198 |
| 61 | 230 | 90 | 196 | 119 | 195 | 148 | 196 | 177 | 229 | 206 | 198 |
| 62 | 197 | 91 | 195 | 120 | 191 | 149 | 163 | 178 | 195 | 207 | 191 |
| 63 | 194 | 92 | 182 | 121 | 193 | 150 | 195 | 179 | 204 | 208 | 195 |
| 64 | 195 | 93 | 207 | 122 | 195 | 151 | 196 | 180 | 199 | 209 | 193 |
| 65 | 195 | 94 | 198 | 123 | 179 | 152 | 172 | 181 | 198 | 210 | 195 |
| 66 | 198 | 95 | 217 | 124 | 197 | 153 | 192 | 182 | 183 | 211 | 161 |
| 67 | 205 | 96 | 203 | 125 | 197 | 154 | 203 | 183 | 195 | 212 | 199 |
| 68 | 204 | 97 | 192 | 126 | 166 | 155 | 229 | 184 | 196 | 213 | 219 |
| 69 | 187 | 98 | 196 | 127 | 195 | 156 | 197 | 185 | 198 | 214 | 195 |
| 70 | 228 | 99 | 229 | 128 | 194 | 157 | 226 | 186 | 205 | 215 | 192 |
| 71 | 199 | 100 | 197 | 129 | 196 | 158 | 198 | 187 | 206 | 216 | 187 |
| 72 | 196 | 101 | 192 | 130 | 196 | 159 | 196 | 188 | 196 | 217 | 196 |
| 73 | 198 | 102 | 195 | 131 | 195 | 160 | 200 | 189 | 201 | 218 | 193 |
| 74 | 196 | 103 | 181 | 132 | 193 | 161 | 192 | 190 | 233 | 219 | 206 |
| 75 | 184 | 104 | 186 | 133 | 201 | 162 | 184 | 191 | 226 | 220 | 197 |
| 76 | 202 | 105 | 194 | 134 | 193 | 163 | 188 | 192 | 188 | 221 | 194 |
| 77 | 196 | 106 | 198 | 135 | 228 | 164 | 204 | 193 | 194 | 222 | 195 |
| 223 | 207 | 252 | 204 | 281 | 193 | 310 | 182 | 339 | 165 | 368 | 198 |
| 224 | 228 | 253 | 216 | 282 | 207 | 311 | 196 | 340 | 196 | 369 | 219 |
| 225 | 199 | 254 | 196 | 283 | 205 | 312 | 189 | 341 | 202 | 370 | 192 |
| 226 | 200 | 255 | 207 | 284 | 191 | 313 | 206 | 342 | 216 | 371 | 221 |
| 227 | 196 | 256 | 199 | 285 | 196 | 314 | 198 | 343 | 184 | 372 | 196 |
| 228 | 198 | 257 | 192 | 286 | 221 | 315 | 200 | 344 | 195 | 373 | 206 |
| 229 | 197 | 258 | 222 | 287 | 195 | 316 | 229 | 345 | 194 | 374 | 196 |
| 230 | 196 | 259 | 201 | 288 | 173 | 317 | 194 | 346 | 183 | 375 | 192 |
| 231 | 197 | 260 | 194 | 289 | 198 | 318 | 186 | 347 | 196 | 376 | 189 |
| 232 | 196 | 261 | 206 | 290 | 191 | 319 | 191 | 348 | 203 | 377 | 203 |
| 233 | 195 | 262 | 171 | 291 | 184 | 320 | 163 | 349 | 196 | 378 | 196 |
| 234 | 197 | 263 | 192 | 292 | 217 | 321 | 197 | 350 | 201 | 379 | 197 |
| 235 | 219 | 264 | 199 | 293 | 182 | 322 | 231 | 351 | 192 | 380 | 196 |
| 236 | 194 | 265 | 206 | 294 | 183 | 323 | 190 | 352 | 201 | 381 | 190 |
| 237 | 199 | 266 | 194 | 295 | 186 | 324 | 189 | 353 | 199 | 382 | 179 |
| 238 | 193 | 267 | 207 | 296 | 200 | 325 | 187 | 354 | 198 | 383 | 192 |
| 239 | 196 | 268 | 196 | 297 | 199 | 326 | 192 | 355 | 194 | 384 | 190 |
| 240 | 201 | 269 | 198 | 298 | 182 | 327 | 196 | 356 | 197 | 385 | 196 |
| 241 | 198 | 270 | 193 | 299 | 233 | 328 | 196 | 357 | 195 | 386 | 195 |
| 242 | 200 | 271 | 171 | 300 | 189 | 329 | 191 | 358 | 162 | 387 | 183 |
| 243 | 196 | 272 | 203 | 301 | 159 | 330 | 160 | 359 | 209 | 388 | 208 |
| 244 | 163 | 273 | 189 | 302 | 194 | 331 | 196 | 360 | 219 | 389 | 198 |
| 245 | 196 | 274 | 195 | 303 | 201 | 332 | 188 | 361 | 233 | 390 | 194 |
| 246 | 227 | 275 | 206 | 304 | 182 | 333 | 194 | 362 | 221 | 391 | 203 |
| 247 | 159 | 276 | 219 | 305 | 166 | 334 | 198 | 363 | 190 | 392 | 220 |
| 248 | 191 | 277 | 207 | 306 | 184 | 335 | 201 | 364 | 213 | 393 | 195 |
| 249 | 195 | 278 | 196 | 307 | 196 | 336 | 196 | 365 | 196 | 394 | 180 |
| 250 | 209 | 279 | 206 | 308 | 201 | 337 | 195 | 366 | 184 | 395 | 159 |
| 251 | 226 | 280 | 194 | 309 | 184 | 338 | 205 | 367 | 229 | 396 | 204 |
| 397 | 192 | 421 | 199 | 445 | 197 | 469 | 199 | 493 | 203 | 397 | 192 |
| 398 | 195 | 422 | 197 | 446 | 196 | 470 | 184 | 494 | 233 | 398 | 195 |
| 399 | 197 | 423 | 182 | 447 | 196 | 471 | 191 | 495 | 172 | 399 | 197 |
| 400 | 195 | 424 | 205 | 448 | 201 | 472 | 197 | 496 | 224 | 400 | 195 |
| 401 | 193 | 425 | 197 | 449 | 178 | 473 | 196 | 497 | 160 | ||
| 402 | 187 | 426 | 203 | 450 | 207 | 474 | 204 | 498 | 164 | ||
| 403 | 191 | 427 | 190 | 451 | 213 | 475 | 205 | 499 | 184 | ||
| 404 | 181 | 428 | 195 | 452 | 164 | 476 | 196 | 500 | 159 | ||
| 405 | 197 | 429 | 161 | 453 | 229 | 477 | 180 |
| 406 | 232 | 430 | 181 | 454 | 192 | 478 | 194 | ||||
| 407 | 210 | 431 | 198 | 455 | 172 | 479 | 196 | ||||
| 408 | 182 | 432 | 191 | 456 | 200 | 480 | 193 | ||||
| 409 | 179 | 433 | 170 | 457 | 194 | 481 | 196 | ||||
| 410 | 188 | 434 | 233 | 458 | 181 | 482 | 177 | ||||
| 411 | 209 | 435 | 193 | 459 | 178 | 483 | 196 | ||||
| 412 | 196 | 436 | 232 | 460 | 171 | 484 | 229 | ||||
| 413 | 180 | 437 | 204 | 461 | 189 | 485 | 198 | ||||
| 414 | 173 | 438 | 230 | 462 | 202 | 486 | 196 | ||||
| 415 | 194 | 439 | 159 | 463 | 166 | 487 | 163 | ||||
| 416 | 164 | 440 | 174 | 464 | 208 | 488 | 160 | ||||
| 417 | 200 | 441 | 184 | 465 | 211 | 489 | 233 | ||||
| 418 | 189 | 442 | 164 | 466 | 195 | 490 | 160 | ||||
| 419 | 173 | 443 | 189 | 467 | 196 | 491 | 191 | ||||
| 420 | 181 | 444 | 193 | 468 | 199 | 492 | 161 |
Claims (1)
1. one kind is detected the method for concentration of complement Clq in human serum, it is characterized in that comprising a step that adopts Biochemical Analyzer to measure in sample, sample is adopted in the step that Biochemical Analyzer measures at described one, sample is added after the first reagent R1, hatch 5min for 37 DEG C, read to survey the 1st sample A1, add the second reagent R2, hatch 5min for 37 DEG C, read to survey the 2nd sample A2, sample A=sample A2-sample A1; Also comprise a step that adopts Biochemical Analyzer to measure calibration object, calibration object is adopted in the step that Biochemical Analyzer measures at described one, calibration is added after the first reagent R1, hatch 5min for 37 DEG C, read to survey the 1st calibration A1, add the second reagent R2, hatch 5min for 37 DEG C, read to survey the 2nd calibration A2, calibration A=calibration A2-calibration A1;
Sample A2-sample A1
Complement Clq concentration (mg/L)= × calibration object concentration (mg/L)
Calibration A2-calibration A1
The parameter of said determination is: 37 DEG C of temperature, dominant wavelength 340nm, commplementary wave length 700nm, sample or calibration object 4 μ l, R1：240μl,R2: 60 μ l, the reaction time is 10 minutes, wherein, the first reagent R1 is by sodium hydrogen phosphate, potassium dihydrogen phosphate, PEG6000, EDTA-Na2Form with TX-100, in described the first reagent, the mass percent concentration of described sodium hydrogen phosphate is 28.6g/L, and the mass percent concentration of described potassium dihydrogen phosphate is 2.7g/L, the mass percent concentration of described PEG6000 is 50g/L, described EDTA-Na2Mass percent concentration be 1g/L, the volumetric concentration of described TX-100 is 0.2ml/L, the second reagent R2 is by sodium hydrogen phosphate, potassium dihydrogen phosphate, PEG6000, EDTA-Na2, TX-100 and the anti-human C1Q antiserum of rabbit composition, in described the second reagent, the mass percent concentration of described sodium hydrogen phosphate is 28.6g/L, the mass percent concentration of described potassium dihydrogen phosphate is 2.7g/L, the mass percent concentration of described PEG6000 is 50g/L, described EDTa-NA2Mass percent concentration be 1g/L, the volumetric concentration of described TX-100 is 0.2ml/L, the sero-fast volumetric concentration of the anti-human C1Q of described rabbit is 300ml/L, the anti-human C1Q of described rabbit is sero-fast tires as 1:64.
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| CN102323427A CN102323427A (en) | 2012-01-18 |
| CN102323427B CN102323427B (en) | 2013-05-22 |
| CN102323427B9 true CN102323427B9 (en) | 2016-05-04 |
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| EP0774117B1 (en) * | 1994-07-29 | 2003-04-02 | BECKMAN Coulter, Inc. | Detergent-facilitated immunoassay of pharmacological agents |
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|---|---|---|---|---|
| US4268171A (en) * | 1977-07-18 | 1981-05-19 | Beckman Instruments, Inc. | Method determining concentration in rate nephelometric immunochemical analysis |
| EP0774117B1 (en) * | 1994-07-29 | 2003-04-02 | BECKMAN Coulter, Inc. | Detergent-facilitated immunoassay of pharmacological agents |
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