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A kit for quantitatively detecting b-type natriuretic peptide and a detection method for b-type natriuretic peptide

A technology for quantitative detection and natriuretic peptide, which is applied in biological testing, measuring devices, material inspection products, etc., can solve the problems of not being able to quickly provide inspection results, low degree of automation, and radioactive contamination, and achieve a high degree of automation and improve Sensitivity, effects of various immune sites

Active Publication Date: 2017-09-29
HEBEI TEWENTE BIOTECH DEV CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The RIA method has radioactive pollution, while the IRMA method is time-consuming and laborious, and the degree of automation is not high
At present, the commonly used laboratory detection method at home and abroad is the electrochemiluminescence method, which has accurate results, but cannot quickly provide inspection results

Method used

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  • A kit for quantitatively detecting b-type natriuretic peptide and a detection method for b-type natriuretic peptide
  • A kit for quantitatively detecting b-type natriuretic peptide and a detection method for b-type natriuretic peptide
  • A kit for quantitatively detecting b-type natriuretic peptide and a detection method for b-type natriuretic peptide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1 Kit for quantitative detection of B-type natriuretic peptide

[0023] 1. Preparation of immunomagnetic beads and detection of captured B-type natriuretic peptide

[0024] 1. Preparation of nano-immunomagnetic beads

[0025] (1) Antibody preparation and purification: 6-week-old healthy BALB / c female mice were immunized with a mixture of B-type natriuretic peptide fragments aar6-18, aar10-22 and aar11-26, according to conventional hybridoma technology and limits Preparation and screening of monoclonal antibody cell lines by dilution method, and then the specific antibody cell lines against B-type natriuretic peptide were proliferated and cultured, injected into mice to prepare ascites, and the obtained ascites was purified by octanoic acid-ammonium sulfate precipitation; similarly New Zealand white rabbits were immunized with the immune antigen, and the anti-B-type natriuretic peptide serum was purified by ammonium sulfate precipitation, and the polyclonal anti...

experiment example 1

[0033] Experimental example 1 Determination of capture rate of immunomagnetic beads

[0034] The capture rate determination of the immunomagnetic beads of Example 1:

[0035]Take a blood sample containing 20 pg / mL BNP in a centrifuge tube, take 0.1 mg of immunomagnetic beads prepared with a polyclonal antibody / magnetic bead coupling ratio of 60 mg / g in the above centrifuge tube, and set up a control group. After reacting on the Dynal-MIX1 rotary mixer at room temperature for 30 minutes (10 r / min), remove it, insert the centrifuge tube into the magnetic stand for separation for 2 minutes, and suck out the supernatant. Add 1 mL of PB (-0.02% Tween 20) to wash twice, suck out the washing solution after separation, and finally resuspend the magnetic beads with 1 mL of PB, and mix the supernatant, washing solution, magnetic bead resuspension and the BNP of the control group to test. Calculate its capture efficiency (CE) according to the formula:

[0036] CE(%)=(C1-C2-C3) / C1*100%...

experiment example 2

[0039] Experimental example 2 Sensitivity detection of simulated blood samples and drawing of standard curve

[0040] The initial concentration of the simulated contaminated BNP blood sample was 4608 pg / mL, and it was diluted according to 2n (n=0, 1, 2...) times (4608pg / mL, 2304 pg / mL, 1152 pg / mL, 576 pg / mL, 288 pg / mL, 144 pg / mL, 72 pg / mL, 36 pg / mL, 18 pg / mL, 9 pg / mL, 4.5 pg / mL, 2.25 pg / mL, 1 pg / mL) and 0 pg / mL, For the determination of BNP, 1 mL each was added to a centrifuge tube, and 0.1 mg of self-made immunomagnetic beads were added to each tube, reacted on a Dynal-MIX1 rotary mixer for 30 min (10 r / min), and then removed, and the centrifuge tube Insert the magnetic frame to separate for 2 min, wash with PB twice, suck out 100 μL of the supernatant, add the sample to the coated 96-well enzyme plate, incubate at 45°C for 30 min; wash the plate three times, add the enzyme-labeled secondary antibody, 45 Incubate at ℃ for 20 minutes; wash the plate 3 times, add fluorescent s...

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Abstract

The present invention relates to a kit for quantitatively detecting B-type natriuretic peptide, which includes immunomagnetic beads and a double-antibody sandwich fluorescence immuno-ELISA kit; the present invention also relates to a detection method for B-type natriuretic peptide, which uses the present invention kit for detection. The kit of the invention has high detection accuracy, strong specificity and high degree of automation. The kit and detection method of the invention are particularly suitable for rapid screening of B-type natriuretic peptide, and have broad application prospects.

Description

technical field [0001] The invention belongs to the field of immune analysis and detection, and in particular relates to a kit for quantitatively detecting B-type natriuretic peptide, and also relates to a detection method for B-type natriuretic peptide using the kit. Background technique [0002] B-type natriuretic peptide (BNP) is a marker of heart failure and is closely related to congestive heart failure. It is mainly derived from ventricular myocytes. The increase in left ventricular volume or pressure load is to stimulate BNP secretion and The main factor of release, the increase of BNP level has been regarded as an independent prognostic indicator of the degree of heart failure and the mortality rate of patients with chronic heart failure. Therefore, BNP has become one of the hotspots of cardiovascular disease research in the world. [0003] At present, the detection of BNP mainly includes radioimmunoassay (RIA), highly sensitive immunoradiometric assay (IRMA), enzym...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/68G01N33/577G01N33/543
CPCG01N33/54326G01N33/54346G01N33/577G01N33/6893G01N2333/58G01N2800/325G01N2800/52
Inventor 邢志祺齐颖颖
Owner HEBEI TEWENTE BIOTECH DEV CO LTD
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