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Immune analysis detection kit for polypeptide related to IgA nephropathy

A detection kit and immunoassay technology, applied in analytical materials, biological testing, biological material analysis, etc., can solve the problems of dynamic monitoring of unfavorable disease treatment effects, unfavorable repeated puncture of kidney biopsy, infection, etc., to achieve clinical detection and detection Stable and specific effect

Active Publication Date: 2019-07-23
THE FIRST AFFILIATED HOSPITAL OF SUN YAT SEN UNIV +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, as we all know, renal biopsy is an invasive operation with absolute and relative contraindications. Complications such as bleeding at the puncture site, hematoma, and infection may also occur after surgery; Dynamic monitoring of disease treatment effects, therefore, finding a non-invasive, stable and reliable laboratory detection index is an important direction in the field of IgA nephropathy diagnosis and research

Method used

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  • Immune analysis detection kit for polypeptide related to IgA nephropathy
  • Immune analysis detection kit for polypeptide related to IgA nephropathy
  • Immune analysis detection kit for polypeptide related to IgA nephropathy

Examples

Experimental program
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Embodiment 1

[0021] Example 1 Preparation of Detection Kit

[0022] The detection kit is preferably an enzyme-linked immunosorbent assay (ELISA) detection kit.

[0023] A. Preparation of microtiter plates with specific antibodies pre-coated with polypeptide marker antigens:

[0024] A1. Preparation of Polyclonal Antibody to Polypeptide Marker Antigen:

[0025] The artificially synthesized polypeptide marker antigen (sequence EETKENEGFTVTAEGK) was coupled to the carrier protein keyhole limpet hemocyanin (KLH), and then the conjugate was used as an immunogen to immunize rabbits. The rabbit antiserum after immunization was collected and the titer was determined.

[0026] A2. Preparation of ELISA plates with pre-coated antibodies:

[0027] The prepared polyclonal antibody to the polypeptide marker antigen was diluted to 10 ng / μL with coating buffer, and then coated in a 96-well microtiter plate at an amount of 100 μL / well, and the plate was washed 3 times after overnight at 4°C. Add 200 μL...

Embodiment 2

[0047] Example 2 ELISA detection method of polypeptide marker antigen

[0048] Samples: serum from patients with IgA nephropathy and healthy people.

[0049] Standard curve production:

[0050] 1. Take out the microtiter plate coated with the specific antibody of the polypeptide marker antigen and equilibrate at room temperature for 30 minutes.

[0051] 2. Dilute the peptide marker antigen with phosphate buffer to a mass concentration of 250ng / mL, 125ng / mL, 62.5ng / mL, 31.25ng / mL, 15.6ng / mL, 7.8ng / mL and 0ng / mL according to 100 μL / well was added to a 96-well plate, and incubated at 37°C for 1 hour.

[0052] 3. Cleaning: pour out the liquid in the wells gently, add 1× washing solution at 200 μL / well, and wash 3 times with an interval of 3 minutes between each time.

[0053] 4. Hybridization: Add the working concentration of HRP-labeled enzyme-labeled mouse anti-polypeptide marker antigen monoclonal antibody (diluted at 1:5000) at 100 μL / well, and incubate at 37°C for 1 hour.

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Abstract

The invention relates to the technical field of biological detection, in particular to an immune analysis detection kit for polypeptide related to IgA nephropathy. The kit comprises the following components: an enzyme label plate coated with a specific antibody of Pep16, a protein standard of Pep16, an enzyme-labeled mouse anti-Pep16 monoclonal antibody, 10x washing liquid, a color developing agent and a reaction termination liquid. The kit disclosed by the invention is stable, rapid, sensitive and accurate in detection, strong in specificity and high in sensitivity, and can quantitatively detect the content of the polypeptide Pep16 related to IgA nephropathy.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to an immunoassay detection kit for polypeptides related to IgA nephropathy. Background technique [0002] IgA nephropathy is a primary glomerular disease characterized by a large amount of granular IgA deposition in the glomerular mesangial area by renal puncture immunofluorescence examination, and is a common cause of chronic kidney disease in children, with 20% to 30% of cases reported After 15 to 30 years, it develops into end-stage renal disease, which requires a lot of medical resources for dialysis or kidney transplantation to maintain life, which brings huge economic pressure and heavy psychological burden to the children's families and society. [0003] The clinical manifestations of IgA nephropathy are ever-changing, and pathological biopsy of the kidney has become the "gold standard" for its diagnosis. However, as we all know, renal biopsy is an invasive ope...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/58G01N33/577
CPCG01N33/68G01N33/581G01N33/577G01N2800/347
Inventor 饶春宝陆小梅
Owner THE FIRST AFFILIATED HOSPITAL OF SUN YAT SEN UNIV