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Test strip for detecting cerebral hemorrhage 31-kDa Occludin after thrombolysis as well as preparation method and application of test strip

A 31-kdaoccludin and test strip technology, which is applied to measurement devices, instruments, disease diagnosis, etc., can solve the problem that the detection method of nano-enzyme detection test strips does not have universal applicability, The problem of low content in serum has achieved the effect of good market application value, good detection specificity and high detection sensitivity

Pending Publication Date: 2020-12-29
THE SECOND PEOPLES HOSPITAL OF SHENZHEN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Vascular recanalization therapy (thrombolysis or thrombectomy) is currently the only effective means of acute stroke treatment, but there is a huge risk of cerebral hemorrhage, resulting in a treatment rate of less than 5%
[0004] The 31-kDa Occludin protein degradation fragment, due to its small molecular weight and low content in serum, has strict requirements for its detection specificity and sensitivity. The detection of diseases by traditional colloidal gold and other immunochromatographic techniques is greatly limited. At the same time, The time window from the onset of stroke to recanalization therapy is less than 6 hours, and a short-term bedside detection is required, which is difficult to fulfill based on proteomics detection
However, these nanozymes in the prior art do not have universal applicability. For different target detection, it is necessary to find suitable nanozymes. At present, there is no nanozyme detection test suitable for the detection of 31-kDa Occludin protein degradation fragments. Paper strips and corresponding detection methods

Method used

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  • Test strip for detecting cerebral hemorrhage 31-kDa Occludin after thrombolysis as well as preparation method and application of test strip
  • Test strip for detecting cerebral hemorrhage 31-kDa Occludin after thrombolysis as well as preparation method and application of test strip
  • Test strip for detecting cerebral hemorrhage 31-kDa Occludin after thrombolysis as well as preparation method and application of test strip

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 Synthesis and Preparation of Anti-31-kDa Occludin Protein Antibody-Nanozyme Conjugate Complex

[0027] 1. Fe 3 o 4 Synthesis of nanozymes

[0028] Nanozyme was synthesized by hydrothermal method: Dissolve 0.3g FeCl3·6H2O in 20mL ethylene glycol, stir to dissolve, then add 1.5g sodium acetate, after the mixture is completely dissolved, seal it in an autoclave, and react at 200°C for 14 hour, the product obtained was magnetically separated, the supernatant was discarded, the precipitate was washed 3 times with ethanol, and then dried and stored at 50-60°C in an electric constant temperature drying oven to obtain Fe 3 o 4 nanozyme.

[0029] 34 Synthesis of Nanozyme-Anti-31-kDa Occludin Antibody Conjugate Complex

[0030] (1) Prepare 1 mL of activator, which is a mixture of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide EDC and N-hydroxysuccinimide NHS, and the EDC The ratio of the amount of substance to the NHS is 1:1;

[0031] (2) Add the Fe prepared by t...

Embodiment 2

[0034] Example 2 Preparation of Detecting 31-kDa Occludin Protein Nanozyme Test Strip

[0035] The preparation for detecting the 31-kDa Occludin protein nanozyme test strip comprises the following steps:

[0036] (1) Sample pad pretreatment: Soak the glass fiber membrane in the sample pad buffer solution to completely soak it, soak it for 5 minutes, take it out, drain and dry it for later use; the sample pad buffer solution is 1% BSA-PBS, 1% Tween- 20. The preparation method of the 1% BSA-PBS is as follows: 1 g of bovine serum albumin BSA is dissolved in 100 mL of phosphate buffer solution PBS, the pH of the phosphate buffer solution is 7.4, and the molar concentration of the phosphate buffer solution is It is 10mmol / L.

[0037] (2) Pretreatment of the reaction pad: soak another piece of glass fiber membrane in the buffer of the reaction pad until it is completely soaked; soak for 5 minutes, take it out and dry it and dry it for later use; the buffer of the sample pad is 1% B...

Embodiment 3

[0041] Example 3 Optimization of preparation process conditions for detecting 31-kDa Occludin protein nanozyme test strips

[0042] 1. Exploration of the optimal surface active type and concentration in the reaction pad buffer

[0043] (1) Preparation of different types and concentrations of reaction pad buffers and reaction pad pretreatment

[0044] A suitable surfactant can often enhance the reaction signal and eliminate false positives during the entire reaction process. On the contrary, if the surface activity is not suitable, it will often lead to problems such as confusion of test results and inability to distinguish between positive and negative. Therefore, adding a suitable surfactant to the reaction pad is very important for the detection performance of the 31-kDa Occludin protein nanozyme test strip. This experiment mainly explores the surfactant suitable for the detection of 31-kDa Occludin protein nanozyme. %, 0.75%, and 1% concentration gradients were added to ...

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Abstract

The invention discloses a test strip for detecting cerebral hemorrhage 31-kDa Occludin after thrombolysis. The test strip is prepared by sequentially pasting a sample pad, a reaction pad, a nitrocellulose membrane and an absorption pad on a PVC bottom card, wherein the nitrocellulose membrane adopts a goat anti-rabbit antibody line as a quality control line C, an IgG capture antibody of rabbit anti-31-kDa Occludin protein is adopted as a detection line T on the nitrocellulose membrane, the reaction pad is sprayed with a Fe3O4 nano-enzyme-anti-31-kDa Occludin protein antibody coupling compound,and the reaction pad is pretreated by adopting 1-2% of Casein-PBS and 0.5-1% of Triton X-100. The test strip is good in specificity and high in sensitivity, is a prediction product which is low in price, high in accuracy, rapid in detection and simple to operate and is used for predicting cerebral hemorrhage risk after cerebral apoplexy thrombolysis or thrombus taking, and has good market application value.

Description

technical field [0001] The invention belongs to the field of immunochromatography detection, in particular to the detection of cerebral hemorrhage after thrombolysis or thrombectomy in stroke, in particular a protein fragment of cerebral hemorrhage after thrombolysis or thrombectomy in stroke and its application. Background technique [0002] For nearly 20 years from 1990 to 2017, stroke has been ranked the first cause of death among Chinese nationals. Its five characteristics of high morbidity, high mortality, high disability, high recurrence rate and heavy economic burden seriously threaten human health. Vascular recanalization therapy (thrombolysis or thrombectomy) is currently the only effective means of acute stroke treatment, but there is a huge risk of cerebral hemorrhage, resulting in a treatment rate of less than 5%. How to clarify the etiology of cerebral hemorrhage caused by thrombolysis or thrombectomy, early identification of risk warning signs, and provide rap...

Claims

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Application Information

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IPC IPC(8): G01N33/558G01N33/543
CPCG01N33/558G01N33/54346G01N2800/2871
Inventor 刘文兰徐迹廖子君雷雪萌李梦娜张圆
Owner THE SECOND PEOPLES HOSPITAL OF SHENZHEN
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