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Immune syntony scattering spectrometry for detecting B factor

A technology of resonance scattering and factor B, applied in the field of immune resonance scattering spectroscopy for detecting factor B, can solve the problems of long operation process, high cost, complicated operation, etc., and achieves less sample consumption, short measurement time and high sensitivity. Effect

Inactive Publication Date: 2008-04-09
GUANGXI NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, the classic immunoprecipitation method (one-way diffusion, rocket electrophoresis, etc.) can detect the content of factor B, but the one-way diffusion method has low sensitivity, poor accuracy and reproducibility, and the process is too long; the rocket electrophoresis method is complicated to operate The hemolysis method is often used to determine the activity of factor B; the double-antibody sandwich ELISA method avoids the labeling of specific antibodies, but increases the operation steps, and needs to use the enzyme-labeled antibody against the second animal, the cost of the reagent is high, and the used The enzyme substrates are often aniline and phenol reducing dyes, which are less stable and easily oxidized when exposed to light or air. What’s more, aniline substrates have carcinogenic effects, and the determination process includes coating, incubation, etc. , Washing, immune reaction, color development and other steps take a long time, at least 24 hours, and the operation is cumbersome
[0006] In recent years, some people have used immunoturbidimetry to measure factor B. Although it is simple and fast, the sensitivity is not high.
In addition, some researchers use the surface plasmon resonance B factor sensor to measure the B factor, although the detection limit is low, but the operation is complicated; there is also a piezoelectric immunosensor to measure the B factor, which has high sensitivity, selectivity and reproducibility. Both are good, but the operation process is long and complicated
The immunoresonance scattering spectrometry method of the present invention detects B factor and has not been reported yet

Method used

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  • Immune syntony scattering spectrometry for detecting B factor
  • Immune syntony scattering spectrometry for detecting B factor
  • Immune syntony scattering spectrometry for detecting B factor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] 1. Prepare a test system with known factor B concentration, add Na 2 HPO 4 -NaH 2 PO 4 Buffer solution (PBS) 0.40 mL, goat anti-human factor B solution (dilution ratio 1:20, Shanghai Beika Reagent Co., Ltd.) 200 μL, 12 μg / mL factor B antigen (Shanghai Beika Reagent Co., Ltd.) 20, 140, 260 , 380, 500, 620, 740μL, shake well, then add 0.40mL of 15% PEG6000 solution, add double distilled water to make up to 3.0mL, shake well, put the stoppered test tube in 37℃ water bath for 5 minutes, take out the running water to cool ;

[0030] 2. According to the method in step 1, but without adding factor B antigen, prepare a reagent blank control system;

[0031] 3. the above-mentioned system gained solution is poured in the quartz cuvette respectively, is placed on the Cary Eclipse fluorescence spectrophotometer (U.S. Varian company) at λ ex -λ em = 0 conditions, synchronous scanning to obtain the resonance scattering spectrum of the system (as shown in Figure 1), measure the ...

Embodiment 2

[0037] 1. Add pH 7.2 Na 2 HPO 4 -NaH 2 PO 4 0.40 mL of buffer solution, 200 μL of goat anti-human factor B solution (dilution ratio 1:20, Shanghai Beika Reagent Co., Ltd.), 12 μg / mL factor B antigen (Shanghai Beika Reagent Co., Ltd.) 50, 100, 200, 400, 800 μL, shake well, then add 0.50 mL of 15% PEG6000 solution, add twice distilled water to make up to 3.0 mL;

[0038] 2. Place the stoppered test tube in step 1 in a 37°C water bath, and after reacting for 5 minutes, take out the running water to cool;

[0039] 3. the solution gained in step 2 is poured in the quartz cuvette, placed on the Cary Eclipse fluorescence spectrophotometer (U.S. Varian company) at λ ex -λ em =0 conditions, synchronously scan to obtain the resonance scattering spectrum of the system, and simultaneously measure the resonance scattering intensity I at the wavelength of 400nm;

[0040] 4. Take no B factor antigen as the reagent blank, and the measured resonance scattering intensity is the blank valu...

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Abstract

The invention discloses a resonance scattering spectrometric for detecting B factor, which utilizes the resonant scattering effect of the immune complex to detect. When the B factor in the blood serum is compounded with the specificity goat to human B factor antibody, an insoluble complex is formed, the insoluble complex has the resonant scattering effect, the insoluble complex congregates with the effect of the PEG, and the scattering intensity is strengthened obviously. The content of the B factor can be worked out by the B factor concentration of the calibration serum--resonance scattering correct value curve. Compared with the existing method, the invention combines the immune-reaction with the resonance scattering spectrometric technique; the invention has the advantages that the sensitivity is high and the detection limit is low, and a Mu g / ml level can be achieved, the dosage of the sample used for testing low concentration B factor is little, the operation is simple and convenient, the time of test is short, the reagent used for the test is innocuous and safe, and the cost is low.

Description

Technical field: [0001] The invention relates to a detection method for B factor, in particular to an immunoresonance scattering spectrometry for detecting B factor. Background technique: [0002] B Factor (BF), also known as C 3 Activator precursor (C 3 PA), which is an important component in the alternative complement pathway activation pathway. Beta 2 Globulin, with a molecular weight of 100,000 D, has only one polypeptide chain, is rich in glycine, and is unstable to heat. It will be inactivated within 30 minutes at 50°C. Under the action of D, factor B is cleaved into two fragments a and b with different antigenicity. B factor must be with C 3b Etc and Mg 2+ Combines into C in the presence of 3b·B After that, B is released by the action of D a , forming C 3b ·B b , the complex has C 3 Invertase activity, acting on C 3 generate more C 3b , forming under the condition of P factor (C 3b )nB b P, which is the C in the alternative pathway 5 Invertase. [000...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/541G01N21/63
Inventor 李纪顺蒋治良
Owner GUANGXI NORMAL UNIV
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