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Ratio-type ECL biosensor as well as preparation method and application thereof

A biosensor, the technology of SEQIDNO.8-SEQIDNO.10, is used in biochemical equipment and methods, microbial determination/inspection, resistance to vector-borne diseases, etc. And other issues

Active Publication Date: 2021-08-20
JIANGSU INST OF NUCLEAR MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, due to the limited walking space of single-legged DNA Walker, the walking efficiency cannot be improved

Method used

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  • Ratio-type ECL biosensor as well as preparation method and application thereof
  • Ratio-type ECL biosensor as well as preparation method and application thereof
  • Ratio-type ECL biosensor as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0062] Au-g-c 3 N 4 Preparation:

[0063] 50 μl of HAUCL 4 (0.01M) solution and 4 ml of G-C 3 N 4 (0.15mg ml -1 Mixed together, ultrasonic stirring for 10 minutes, then stirring for 1 hour. Then, 100 μl of fresh NABH is injected into the mixed solution. 4 (0.01 m) solution was treated in an ice bath for 20 minutes. A 50 μl of mass fraction was continuously added to the mixed solution of 1% sodium citrate solution, and the mixed solution was stirred at 25 ° C for 20 minutes. Finally, in order to remove excess Aunps, sodium citrate and NABH 4 , The above mixed solution was washed with ultrapure water with ultrapure water after centrifugation of 12000 rpm, and the sonication was treated 3 times to give pure Au-G-C. 3 N 4 Re-dispensed it in 1 ml h 2 O is preserved at 4 ° C for the next experiment.

[0064] Pei-Ru @ Ti 3 C 2 @ AUNPS-S7 probe synthesis Follow these steps:

[0065] 1, 10 mg of three (4,4'-dicarboxylic acid-2,2'-bispyridine) ruthenium (II) dichloride [RU (DCBPY) 3 2+ EDC...

Embodiment 1

[0073] Example 1 Composition of the ratio ECL biosensor and used to detect SARS COV-2RDRP genes

[0074] I. Comparison ECL biosensor construction method, including the following steps:

[0075] 1. Clean the glass carbon electrode (GCE). 10 μL of Au-g-C 3 N 4 Fixed on GCE to get AU-G-C 3 N 4 / GCE and dry at room temperature. Subsequently, Au-g-c will be obtained. 3 N 4 / GCE was soaked in a TDNAS (1 μM) solution containing 10 mm TCEP in a 10 mM TCEP to reduce the S-S bond. Next, the modified GCE was soaked in a 60 μl of MCH (100 μm) solution for 1 hour to prevent non-specific adsorption. In each of the above steps, the unconnected substance is required to be washed with a PBS solution. Will construct a good electrode (MCH / TDNAS / AU-G-C 3 N 4 / GCE) is stored in a fridge in 4 ° C for use with the next experiment.

[0076] 2, put MCH / TDNAS / AU-G-C 3 N 4 / GCE incubated in an entropy drive mixture of 37 ° C for 40 min, entropy-driven reaction mixture comprising different concen...

Embodiment 2

[0105] Example 2 Detect SARS-COV-2RDRP with a ratio ECL biosensor

[0106] 1. Under the optimum conditions of Example 1, the SARS-COV-2RDRP gene (detection method is seen in Example 1) was detected by the ratio ECL biosensor obtained in Example 1. Figure 7 A describes that as the SARS-COV-2RDRP concentration is increased from 10AM to 10 pm, the ECL signal strength decreases at 460 nm, and increases at 620 nm. Figure 7 A also describe the change in ECL signal strength at 460 nm has a good linear relationship, ie y = -502.82LGC (RDRP) -986.17, r 2 = 0.9944.620 nm ECL signal strength change also exhibits Y = 805.79LGC (RDRP) +14704.28 excellent linear relationship, R 2 = 0.9950. In order to obtain higher reliability, the increase in ECL (460 nm) / ECl (620 nm) of ECl (460 nm) / ECl (620 nm) is evaluated. Figure 7 B shows. According to the detection limit (LOD) = 3σ / k, the LOD is 7.8AM.

[0107] 2, the selectivity and stability of the ratio ECL biosensor (in Example 1)

[0108] In o...

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Abstract

The invention constructs a ratio electrochemical luminescence (ECL) biosensor, which is used for detecting the RNA-dependent RNA polymerase (RdRp) gene of severe acute respiratory syndrome coronavirus 2 (SARS-COV-2). The SARS-COV-2RdRp gene participates in the entropy-driven cyclic amplification reaction, then a piece of Bandage DNA is output, and can be combined with the other two single chains S1 and S2 to form a double-chain DNAWalker, and the next cyclic reaction is started. After the walking process of the double-foot DNA Walker is completed, a hairpin structure at the top of a DNA tetrahedron (TDNAs) is removed. Then, a PEI-Ru@Ti3C2@AuNPs-S7 probe is used for being specifically combined with a TDNAs hairpin part cut off from the surface of Au-g-C3N4, and signal change is achieved through ECL-RET (electrochemiluminescence resonance energy transfer).

Description

Technical field [0001] The present invention relates to the field of biotechnology, and more particularly to a ratio ECL biosensor and a preparation method thereof. Background technique [0002] The outbreak of the 2019 new coronavirus disease (COVID-19) caused by severe acute respiratory syndrome crosent virus 2 (SARS-COV-2) has jeopardized the health of the people of the world. Before developing an effective vaccine, in order to prevent the spread of new coronavirus diseases, the COVID-19 is first accurately diagnosed, especially for diagnosis of non-symptomatic patients. At present, there are two main strategies for the detection of new coronavirus diseases in the market. One strategy is to detect SARS-COV-2RNA using a polymerase chain reaction (PCR) technology. However, the PCR's operation is more complicated. It is necessary to design different primers and temperatures, low accuracy, usually false results. Another strategy is based on antibody serological and immunological t...

Claims

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Application Information

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IPC IPC(8): C12Q1/6825C12Q1/682C12Q1/70C12N15/11
CPCC12Q1/6825C12Q1/682C12Q1/701Y02A50/30
Inventor 张凯范振强丁月娣谢敏浩
Owner JIANGSU INST OF NUCLEAR MEDICINE
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