PLGF magnetic particle chemiluminiscence kit and detection method thereof

A chemiluminescence reagent and chemiluminescence technology, applied in chemiluminescence/bioluminescence, analysis by making materials undergo chemical reactions, measurement devices, etc., can solve problems, only applicable to the detection and identification of trace substances, and the cost of excitation solution Low, low background luminescence and other issues, to achieve the effect of short response time, high sensitivity, and simple operation

Pending Publication Date: 2019-09-20
NINGBO AUCHEER BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] So far, the methods used to detect placental growth factor residues in human serum mainly include: enzyme-linked immunoassay (ELISA). However, although the detection price of ELISA is low and fast, the sensitivity is not enough, and it is only suitable for the detection of trace substances. Detection and identification; the method adopted in the present invention is the direct chemiluminescence method, and the use of acridinium ester as a chemiluminescent marker has obvious advantages, mainly in that the reaction does not require a catalyst, as long as the alkaline environment can be carried out, the reaction is rapid, and can be completely captured Photons generated by reaction, low background luminescence, high signal-to-noise ratio, less interference factors, good reagent stability, simple system, low cost of excitation solution, acridinium ester is easy to bind with protein, and the photon yield does not decrease after the association

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] A PLGF magnetic particle chemiluminescence kit, comprising

[0024] Biotin-labeled placental growth factor antibody;

[0025] Streptavidin magnetic particle suspension;

[0026] Acridinium ester-labeled placental growth factor antibody;

[0027] calibrator;

[0028] Cleaning fluid;

[0029] Diluent;

[0030] Chemiluminescence pre-excitation solution A;

[0031] Chemiluminescence pre-excitation solution B;

[0032] The streptavidin magnetic particle suspension is a magnetic particle suspension formed by suspending ferric oxide particles coated with streptavidin on the surface in a magnetic particle coating buffer, and the magnetic particle coating The buffer is 20mM-200mM Tris (trishydroxymethylaminomethane) buffer.

[0033] The biotin-labeled placental growth factor antibody is a monoclonal antibody;

[0034] The cleaning solution is a Tris-HCl buffer solution with a concentration of 0.025 mol / L, which contains NaCl and 0.05% Tween-20 with a concentration of 0.1...

Embodiment 2

[0044]Included in the present invention is the preparation method of streptavidin magnetic particle solution, and concrete steps are:

[0045] Dissolve 20g of magnetic fluid and 2g of polyethylene glycol in 73.68g of absolute ethanol, and move into a 2 In the three-necked bottle at the entrance, N 2 In the atmosphere, under the condition of 65°C, stir at 300r / min for 30min, then raise the temperature to 70°C, add 19.78g styrene, 2g maleic anhydride, 1.37g acrylic acid, 0.25g divinylbenzene, 6g benzoyl peroxide, After 10min, the reactant became emulsion, keeping N 2 Atmosphere, 70°C, stirring speed 300r / min, reaction time 12h, after the reaction is completed, polystyrene magnetic particles are collected for future use.

[0046] The PBS buffer solution was autoclaved, 1 mg of streptavidin was dissolved in 1 ml of PBS buffer solution, and 200 ul was put into an EP tube for use.

[0047] Soak 50 mg of polystyrene magnetic particles in ultrapure water, wash with PBS buffer solut...

Embodiment 3

[0050] Included in the present invention is the preparation method of streptavidin magnetic particle solution, and concrete steps are:

[0051] 25g of magnetic fluid (ferric oxide) and 3g of polyethylene glycol were dissolved in 72g of absolute ethanol, and moved into 2 In the three-necked bottle at the entrance, N 2 In the atmosphere, under the condition of 65°C, stir at 300r / min for 30min, then raise the temperature to 70°C, add 20g styrene, 2g maleic anhydride, 2g acrylic acid, 0.25g divinylbenzene, 4.75g benzoyl peroxide, 10min Afterwards, the reactant becomes an emulsion, keeping N 2 Atmosphere, 70°C, stirring speed 300r / min, reaction time 12h, after the reaction is completed, polystyrene magnetic particles are collected for later use;

[0052] Autoclave the PBS buffer, dissolve 1mg of streptavidin in 1ml of PBS buffer, take 200ul and put it in an EP tube for use;

[0053] Soak 50mg of polystyrene magnetic particles in ultrapure water, wash with PBS buffer solution for...

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Abstract

The present invention discloses a PLGF (Placental Growth Factor) magnetic particle chemiluminiscence kit and a detection method thereof. The kit comprises a placental growth factor antibody with biotin labeling, streptavidin magnetic particle suspension liquid, a placental growth factor antibody with acridinium ester labeling, a calibration product, cleanout fluid, a diluent, chemiluminiscence pre-excitation liquid A, and chemiluminiscence pre-excitation liquid B. The streptavidin and the biotin have a high-specificity combining capacity, the high-purity antibody of the streptavidin and the biotin labeling is subjected to specific binding through a non covalent bond and has a cascade amplification effect. Besides, an acridinium ester chemiluminescent system is adopted, and the acridinium ester does not need a catalyst and can emit light when being put in a H2O2 solution with no need for a catalytic process and an enhancer. The sensitivity is higher, the reaction time is short, the operation is simple, and the anti-interference performance is high.

Description

technical field [0001] The invention relates to the field of in vitro diagnosis, in particular to a PLGF magnetic particle chemiluminescence kit and a detection method thereof. Background technique [0002] Placental growth factor (PLGF) was first isolated and purified from human placental cDNA library by Maglione et al. in 1991. PLGF is mainly synthesized by syncytiotrophoblast cells and can bind to tyrosinase receptors located in trophoblast cells and vascular endothelial cells. It is a protein that has autocrine effects on trophoblast cell functions and paracrine effects on angiogenesis. PLGF has a unique regulatory effect on the function of trophoblast cells and endothelial cells, and can promote angiogenesis. Detecting the blood PLGF level of pregnant women can be used clinically to identify the oxygen supply pressure of placental syncytiotrophoblast cells, and to predict, identify and monitor treatment of preeclampsia. [0003] So far, the methods used to detect plac...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/74G01N33/543G01N33/532G01N21/76
CPCG01N33/74G01N33/54326G01N33/532G01N21/76
Inventor 周义正唐静陈星星
Owner NINGBO AUCHEER BIOTECHNOLOGY CO LTD
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