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Method of detecting target sarcosine of prostatic cancer

A prostate cancer and sarcosine-labeled technology, which is applied in the detection of prostate cancer target sarcosine, in the field of detection systems based on cascade reactions, can solve the problems of detection sensitivity and specificity, high false positive rate, etc., and achieve improved detection Sensitivity, improvement of assembly efficiency, improvement of poor assembly efficiency

Inactive Publication Date: 2018-03-06
SHANGHAI NAT ENG RES CENT FORNANOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the curability of early prostate cancer, the existing early screening is mainly based on prostate-specific antigen (PSA), which has great problems in detection sensitivity and specificity, and has a high false positive rate

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] SOx and HRP were assembled with thiol DNA 1 and DNA 2 respectively: 100 μl SOx / HRP (2 μM) was incubated with SPDP at a ratio of 1:10 in 1 × PBS buffer for 2 hr at room temperature. Excess SPDP was washed by centrifugation (3000g, 10min) three times with 30 kD ultrafiltration tube, recovered by 1000g reverse ultrafiltration for 10 minutes, and quantified by ultraviolet light. SPDP-modified SOx / HRP were incubated with 10-fold excess of thiol-DNA 1 and DNA 2 in 1 × PBS buffer overnight at room temperature. The connection ratio of SOx, HRP and DNA was quantified by measuring the change of absorbance at 343 nm. Finally, excess DNA was removed by centrifugal ultrafiltration (3000g, 10min, three times), and recovered by 1000g reverse ultrafiltration for 10 minutes to obtain SOx-DNA complex I and HRP-DNA complex II.

[0029] DNA tubular origami synthesis: the replacement strand DNA 3, DNA 5-13 and the rest of the unsubstituted staple strands were mixed evenly, diluted to 200 nM,...

Embodiment 2

[0034] SOx and HRP were assembled with thiol DNA 1 and DNA 2 respectively: 100 μl SOx / HRP (2 μM) was incubated with SPDP at a ratio of 1:10 in 1 × PBS buffer for 2 hr at room temperature. Excess SPDP was washed by centrifugation (3000g, 10min) three times with 30 kD ultrafiltration tube, recovered by 1000g reverse ultrafiltration for 10 minutes, and quantified by ultraviolet light. SPDP-modified SOx / HRP were incubated with 10-fold excess of thiol-DNA 1 and DNA 2 in 1 × PBS buffer overnight at room temperature. The connection ratio of SOx, HRP and DNA was quantified by measuring the change of absorbance at 343 nm. Finally, excess DNA was removed by centrifugal ultrafiltration (3000g, 10min, three times), and recovered by 1000g reverse ultrafiltration for 10 minutes to obtain SOx-DNA complex I and HRP-DNA complex II.

[0035] DNA tubular origami synthesis: the replacement strand DNA 3, DNA 5-13 and the rest of the unsubstituted staple strands were mixed evenly, diluted to 200 n...

Embodiment 3

[0040] SOx and HRP were assembled with thiol DNA 1 and DNA 2 respectively: 100 μl SOx / HRP (2 μM) was incubated with SPDP at a ratio of 1:10 in 1 × PBS buffer for 2 hr at room temperature. Excess SPDP was washed by centrifugation (3000g, 10min) three times with 30 kD ultrafiltration tube, recovered by 1000g reverse ultrafiltration for 10 minutes, and quantified by ultraviolet light. SPDP-modified SOx / HRP were incubated with 10-fold excess of thiol-DNA 1 and DNA 2 in 1 × PBS buffer overnight at room temperature. The connection ratio of SOx, HRP and DNA was quantified by measuring the change of absorbance at 343 nm. Finally, excess DNA was removed by centrifugal ultrafiltration (3000g, 10min, three times), and recovered by 1000g reverse ultrafiltration for 10 minutes to obtain SOx-DNA complex I and HRP-DNA complex II.

[0041] DNA tubular origami synthesis: the replacement strand DNA 3, DNA 5-13 and the rest of the unsubstituted staple strands were mixed evenly, diluted to 200 n...

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PUM

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Abstract

The invention provides a method of detecting target sarcosine of prostatic cancer. The method comprises the following steps: assembling SOx and HRP with DAN separately; synthesizing tubular DNA fold paper; assembling an enzyme-DNA compound with two types of DNA fold paper separately; assembling an SOx-origami compound III and an HRP-origami compound IV and detecting sarcosine; separately connecting cascaded enzymes Sox and HRP to the two types of tubular DNA fold paper; and achieving orderly cascading between the two enzymes by means of interaction of strong basic groups of complementary arm chains extending from the two types of fold paper, thereby achieving a purpose of improving detecting sensitivity. The novel detection method of regulating the distance between SOx and HRP orderly is established by means of the tubular fold paper structure. By way of further assembly after assembly of the enzymes and fold paper separately, the defect of poor assembling efficiency as a result of steric-hinderance effect is improved, and the assembling efficiency of the two enzymes is improved.

Description

technical field [0001] The invention designs a method for detecting sarcosine, a target of prostate cancer, and specifically relates to using a DNA tubular origami structure to construct a detection system based on a cascade reaction, and to measure the content of sarcosine through a color reaction. The invention belongs to the field of biological sensor detection. Background technique [0002] With the aging of Chinese society, prostate cancer has shown explosive growth in China. Due to the curability of early prostate cancer, the existing early screening is mainly based on prostate-specific antigen (PSA), which has serious problems in detection sensitivity and specificity, and has a high false positive rate. The rising incidence of prostate cancer urgently requires the development of assays targeting other prostate cancer targets. Metabolomics studies have shown that sarcosine is involved in the development of prostate cancer, and its content is significantly increased, ...

Claims

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Application Information

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IPC IPC(8): C12Q1/28C12Q1/26
CPCC12Q1/26C12Q1/28G01N2800/342
Inventor 何丹农徐艳陈玮嘉王萍金彩虹
Owner SHANGHAI NAT ENG RES CENT FORNANOTECH
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