Biologic sensor for detecting human beta interferon and its special polypeptide

A biosensor and interferon-beta technology, applied in the field of biosensors and their special peptides, can solve the problems of denaturation and inactivation, high price, and decline in biological activity, and achieve good selectivity and good reproducibility

Inactive Publication Date: 2008-07-16
INST OF CHEM CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the sensing element is an antigen or antibody molecule immobilized on the surface of the sensor chip, and most proteins such as antigen or antibody are very sensitive to the environment, and their immobilization proc

Method used

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  • Biologic sensor for detecting human beta interferon and its special polypeptide
  • Biologic sensor for detecting human beta interferon and its special polypeptide
  • Biologic sensor for detecting human beta interferon and its special polypeptide

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0032] The preparation of the biosensor is carried out as follows:

[0033] Through the self-assembly of 1,2-ethanedithiol on the surface of the wafer gold electrode, the surface of the electrode is brought with mercapto groups; under weakly alkaline conditions, the mercapto groups are reacted with diepoxy 1,4-butanediol diglycidyl ether, and the gold electrode The surface is modified by epoxy groups; the small molecule recognition element antisense peptide AS-IFN 1 or AS-IFN 3 can be modified to the surface of the gold electrode through the reaction of its amino group and epoxy groups, and a biosensor is obtained. Its structure is as figure 1 In the figure, (a) AS-IFN 1 immobilized wafer; (b) AS-IFN 3 immobilized wafer; (c) blank wafer.

[0034] In order to control the loading amount of the polypeptide, its resonance frequency was measured before and after immobilization, and the amount of bound substance was calculated according to the Sauerbray equation. For a 10 MHz quar...

Embodiment 1

[0041] Embodiment 1, the preparation of biosensor

[0042] Take a 10MHz AT-cut piezoelectric quartz wafer with a diameter of 12.0mm, double-sided gold-plated electrodes, and an electrode diameter of 6.0mm. With Piranha solution (concentrated sulfuric acid: 30% H 2 o 2 =3:1, v / v), and then carefully washed with ethanol and three-distilled water respectively, and dried for later use. Immerse the cleaned wafer in 5% ethanol solution of ethanedithiol, keep it in the dark for 2 hours, wash it with ethanol and triple distilled water, and then immerse it in 1mL 1,4-butanediol diglycidyl ether and 4mL pH 9.5 0.1mol / LNa 2 CO 3 In the mixed solution composed of buffer solution, react at 40° C. for 4 hours, stop the reaction, rinse with water, and obtain a modified wafer. The antisense peptide AS-IFN1 was dissolved in a pH 10.0 buffer to make a 2 mg / ml solution, which was dropped onto the surface of the modified chip electrode with a micro-syringe, and reacted at 4°C for 20 hours. ...

Embodiment 2

[0044] Embodiment 2, the specificity experiment of biosensor

[0045] The blank wafer was fixed in the QCM flow cell with an O-ring rubber seal, and the modified side was in contact with the solution and the other side was placed in the gas phase. The mobile phase is 20mmol / L phosphate buffer, pH 7.4, and the flow rate is 30-90μL / min. Inject 100 μL of human beta interferon, bovine serum albumin (BSA), human serum albumin (HSA) and chicken ovalbumin (CEA) protein solutions with different concentrations through the six-port injection valve, and record the sensing spectrum, such as figure 2 Shown (mobile phase: 20mmol / L phosphate buffer (pH 7.4); flow rate: 60μL / min; protein concentration: 1.0mg / mL; injection volume: 100μL). The results showed that human beta interferon and other three proteins had no non-specific adsorption on the blank chip.

[0046] The human interferon-beta biosensors with AS-IFN 1 and AS-IFN3 as ligands were fixed in the QCM flow cell with an O-shaped rub...

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Abstract

The invention discloses a biological sensor for detecting the human interferon Beta and the dedicated polypeptide thereof. The amino acid sequence of the polypeptide provided by the invention is SEQ No.1, for example. The biological sensor for detecting the human interferon Beta of the invention includes an identifying element and a transducer, wherein, the identifying element is the polypeptide which at least contains the SEQ No.1 amino acid sequence or the derivative thereof. The invention designs and synthesizes an antisense peptide which has the specific interaction with the human interferon Beta, and a biological sensor is prepared by using the antisense peptide, thus realizing the detection of the human interferon Beta. The biological sensor of the invention shows the obvious special combination of the human interferon Beta and has good selectivity; the signal response and the sample size of the human interferon Beta show a linear relation, the reproducibility is good, and the invention can be used for the quantitative detection of human interferon Beta.

Description

technical field [0001] The invention relates to a biosensor for detecting human beta interferon and its special polypeptide. Background technique [0002] Human beta interferon (hIFN-β) is a glycoprotein with a molecular weight of 20kD composed of 166 amino acids. It is mainly secreted by fibroblasts induced by viruses and double-stranded RNA in vivo, and can also be cultured by prokaryotic or eukaryotic cells. expression produced. As a bioengineering product, hIFN-β belongs to type I interferon, which has broad-spectrum anti-virus, anti-tumor and immunomodulatory effects. It is mainly used clinically for multiple sclerosis, hepatitis B, hepatitis C and various Tumor treatment. At present, the methods for detecting human beta interferon mainly include antiviral activity detection method, ultraviolet spectrophotometry, isoelectric focusing method, SDS-PAGE, and HPLC method [Liu Changnuan, Zhang Yi, Rao Chunming, Lin Jianwei, Gao Kai, Zhang Mingfang, Wang Junzhi: Research o...

Claims

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Application Information

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IPC IPC(8): G01N33/68
Inventor 赵睿罗佳章群丹刘国诠
Owner INST OF CHEM CHINESE ACAD OF SCI
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