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Ischemic retinopathy detection biomarker, detection kit and application

A retinopathy and biomarker technology, applied in the field of biomedical detection, can solve the problem of no circRNA-Wdr37, and achieve the effect of reducing treatment and medical costs

Active Publication Date: 2020-11-13
南京医科大学眼科医院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Currently, there is no report of circRNA-Wdr37 as a biomarker for the diagnosis of ischemic retinopathy

Method used

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  • Ischemic retinopathy detection biomarker, detection kit and application
  • Ischemic retinopathy detection biomarker, detection kit and application
  • Ischemic retinopathy detection biomarker, detection kit and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 circRNA-Wdr37 ischemic retinopathy validation embodiment

[0037] circRNA microarray screening related circRNA ischemic retinopathy disease and verify it.

[0038] Step: Sample Preparation: Construction of mouse retinal ischemia reperfusion (IR) animal model, taking the experimental group (IR, n = 3) and control (Ctrl, n = 3) mice retinal tissue. RNA was extracted with Trizol (Invitrogen) reagent and preserved at -80 ° C spare.

[0039] Step two: Screening of differentially expressed circRNA:

[0040] American companies circRNA Aglient using cDNA microarray analysis circRNA related ischemic retinopathy; Anal specific steps: use of enzyme labeled fluorophore labeled circRNA, obtained with fluorescent probes for microarray hybridization, the labeling conditions in MAUI hybridization using the apparatus and chip hybridization; fluorescence intensity using GenePix 4000B scanner chip chip, and the experimental result into numerical data is saved; distribution of each s...

Embodiment 2

[0045] CircRNA-Wdr37 expression was detected in the blood cells and plasma in Example 2

[0046] The first step: separating serum and plasma samples

[0047] Were collected to obtain the experimental group (primary open-angle glaucoma, central vein occlusion and diabetic retinopathy) and control group (cataract) of each sample of each patient's blood 200 cases, the use of heparin tubes using serum was separated therefrom centrifugally and plasma RNA, for detecting circRNA. Centrifugation conditions of 4 ℃, 12,000rpm, 10min.

[0048] Step two: RNA extraction of serum and plasma samples

[0049] Respectively, in the isolated blood cells and plasma samples after the addition of TRIzol room temperature for 10min, the sample was sufficiently lysed (Note: If the next operation is not performed, a sample may be placed in long-term storage -70 ℃). 200μl of chloroform was added per 1ml TRIzol stand vigorous shaking after mixing at room temperature was naturally 3-5min phase. 4 ℃ 12,000rpm ...

Embodiment 3

[0075] Example 3 circRNA-Wdr37 feasibility embodiment as prognostic markers

[0076] First step; get blood sample to be inspected

[0077] Serum each disease in each of 200 patients with ischemic retinopathy (primary open-angle glaucoma, central vein occlusion and diabetic retinopathy) collected before and after treatment.

[0078] Step 2: Extract RNA from the blood sample to be inspected

[0079] a) serum samples or frozen serum samples taken 0.25ml transferred into a centrifuge tube, add 1ml Trizol, using a pipette by pipetting up and down repeatedly until the cells were completely lysed.

[0080] b) adding chloroform (+ Trizol Reagent sample liquid volume of the amount of 1 / 5 volume) centrifuge tube cap tightly, shake vigorously 15sec, 5min standing at room temperature;

[0081] c) 4 ℃ centrifugation, 12,000g × 15min, carefully removed from the centrifuge tube, then homogenate is divided into three, namely: the supernatant was colorless, white intermediate layer and a lower lay...

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Abstract

The invention belongs to the technical field of biomedical detection, and particularly relates to an ischemic retinopathy detection biomarker, a detection kit and application. The invention disclosesapplication of circRNA-Wdr37 in preparation of a diagnostic reagent for ischemic retinopathy and a diagnostic kit for ischemic retinopathy. Through high-throughput sequencing and quantitative PCR (Polymerase Chain Reaction) verification, the circRNA-Wdr37 is found to have remarkable correlation with ischemic retinopathy. The real-time fluorescence quantitative PCR technology is utilized, and the relative content of circRNA-Wdr37 in aqueous humor is detected; the detection biomarker and kit can be used for screening clinical ischemic retinopathy patients, and a theoretical basis is provided forearly intervention of ischemic retinopathy.

Description

Technical field [0001] The present invention is in the field of biomedical testing technology, particularly to an ischemic retinopathy detection of biomarkers, and detection kit applications, the development of human disease diagnosis and ischemic retinopathy in a subject the expression level according circRNA-Wdr37 judgment. Background technique [0002] Ischemic retinopathy is a common type of eye disease, mainly by central retinal vein occlusion (CRVO), branch retinal vein occlusion (BRVO), high tension glaucoma, diabetic retinopathy (the DR), ocular ischemic syndrome and aorta inflammation retinopathy and other retinal lesions caused by vascular perfusion. The performance of retinal ischemic, hypoxic-ischemic damage leading to retinal ganglion cells and other cell dysfunction, decreased visual function, it can seriously affect the quality of life of patients. In the early chronic ischemic retinopathy, and the patient may be no obvious symptoms, and the start when significant ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12Q1/6851C12N15/113A61K31/7088A61P27/02
CPCA61K31/7088A61P27/02C12Q1/6851C12Q1/6883C12Q2600/118C12Q2600/158C12Q2600/178C12Q2531/113C12Q2563/107C12Q2521/107
Inventor 蒋沁颜标姚牧笛张秋阳马严朱妍张荟颖
Owner 南京医科大学眼科医院
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