Use of an immune composition in the preparation of reagents for non-invasive detection of membranous nephropathy

An immune composition, technology for membranous nephropathy, applied in the direction of measuring devices, instruments, scientific instruments, etc., to achieve high detection accuracy, avoid damage, and easy sampling

Inactive Publication Date: 2020-06-09
苏庆宁 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, clinical practice shows that the positive rate of serum PLA2R antibody in patients with spontaneous membranous nephropathy is only 68.5%, and 31.5% of patients with idiopathic membranous nephropathy are still negative for serum PLA2R antibody, so it is difficult to effectively detect

Method used

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  • Use of an immune composition in the preparation of reagents for non-invasive detection of membranous nephropathy
  • Use of an immune composition in the preparation of reagents for non-invasive detection of membranous nephropathy
  • Use of an immune composition in the preparation of reagents for non-invasive detection of membranous nephropathy

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0108] Example 1 Separation

[0109] Such as figure 1 As indicated, the urine sample collected from the subject was separated as follows:

[0110] Collect 40 ml of fresh urine samples from the subjects, centrifuge them at 1000-4000 rpm for 10 minutes, and collect the sediment (containing cells and cell debris). Resuspend the precipitate with PBS, centrifuge again at 1000-4000 rpm for 10 minutes, wash away the residual soluble protein, and collect the sediment. The sediment was resuspended with PBS and divided into three test samples I, II and III.

[0111] Wherein, the sample I to be tested was suspended and fixed with 0.5 ml of 4% paraformaldehyde for 30 minutes, and used for immunohistochemistry and immunofluorescence. Observational testing such as electron microscopy. Gel electrophoresis buffer was added to the test sample II for western blot detection (WesternBlot). Protease inhibitors were added to the sample III to be tested, stored at -80°C, and used for proteomics...

Embodiment 2

[0112] Example 2 Immunofluorescence detection

[0113] Such as figure 2 As indicated, the samples to be tested were subjected to immunofluorescence detection, as follows:

[0114] Use anti-human IgG4 antibody, anti-human PLA2R antibody, anti-human NPHS2 antibody and anti-human IgA antibody as the primary antibody, the source of antibody serum is mouse, rabbit or sheep, monoclonal antibody or polyclonal antibody;

[0115] Fluorescently labeled donkey anti-mouse, donkey anti-rabbit or donkey anti-sheep antibodies such as FITC, Cy3 and Cy5 are used as secondary antibodies;

[0116] The detection method is single-staining (one primary antibody), double-staining (two primary antibodies) or triple-staining (three primary antibodies);

[0117] The immunofluorescent primary antibody was diluted 100-200 times with phosphate buffered solution (PBS) containing 1% bovine serum albumin (BSA), and incubated with PBS (PBST) containing 0.2% Triton and 1% BSA for 15- 30 minutes, primary an...

Embodiment 3

[0123] Example 3 Detection of Urine IgG4 Immunofluorescence in Different Nephropathy Patients and Normal Subjects

[0124] According to the non-invasive detection method described above in the present invention, collected from normal healthy subjects, membranous nephropathy patients, focal segmental glomerulosclerosis (FSGS) patients, diabetic nephropathy (DN) patients, minimal change nephropathy The urine of (MCD) patients and IgA nephropathy patients were detected by IgG4 immunofluorescence, and the detection images are shown in Figure 5-11 .

[0125] Figure 5 It is the immunofluorescence detection image of a normal healthy subject, as shown in the figure, the normal human urine sediment IgG4 signal is negative.

[0126] Figure 6-7 Immunofluorescence detection images of patients with membranous nephropathy, Figure 6 is the IgG4 signal on intact cells, Figure 7 is the IgG4 signal on cell debris. As shown in the figure, IgG4 positive signals are mostly cell fragment...

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Abstract

The invention provides a use of an immune composition in the preparation of a reagent for noninvasive detection of membranous nephropathy. The immune composition comprises one or more of an anti-IgG4antibody, an anti-PLA2R antibody, an anti-Nephrin antibody and an anti-NPHS2 antibody, and the noninvasive detection of the membranous nephropathy comprises detection of the one or more of IgG4, PLA2R, Nephrin and NPHS2 in a biological sample collected from a subject. The immune composition is used for preparing the reagent for the noninvasive detection of membranous nephropathy, and a detection result highly consistent with renal biopsy can be obtained through the noninvasive detection, so the reagent has high detection accuracy for the membranous nephropathy; and the detection is noninvasive. The invention provides a rapid and noninvasive detection method for the differential diagnosis of the membranous nephropathy.

Description

technical field [0001] The invention relates to the detection of membranous nephropathy, in particular to the use of an immune composition in the preparation of reagents for non-invasive detection of membranous nephropathy. Background technique [0002] The prevalence of chronic kidney disease (CKD) in my country is 10.8%. Membranous nephropathy (MN) is a group of diseases characterized by deposition of immune complexes under the epithelial cells of the glomerular basement membrane and thickening of the basement membrane. common causes of symptoms. Membranous nephropathy accounts for about 6.0%-28.8% of primary glomerular diseases, becoming the second cause of primary glomerular diseases after IgA nephropathy, and also an important cause of end-stage renal disease (ESRD) . In 2016, a large-scale study by the team of Academician Hou Fanfan of Nanfang Hospital showed that the proportion of membranous nephropathy reached 23.4%, second only to IgA nephropathy (28.1%). [0003]...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/533
CPCG01N33/533
Inventor 不公告发明人
Owner 苏庆宁
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