Probe and kit for improving pathogenic microorganism antigen detection sensitivity

A technology for detection of pathogenic microorganisms and antigens, applied in the field of probes, can solve problems such as the difficulty of detecting trace microbial infections, and achieve the effect of improving detection sensitivity and reducing detection background

Active Publication Date: 2021-08-13
BEIJING INST OF HEPATOLOGY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to solve the problem that the trace microbial infection mentioned in the background technology is difficult to de

Method used

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  • Probe and kit for improving pathogenic microorganism antigen detection sensitivity
  • Probe and kit for improving pathogenic microorganism antigen detection sensitivity
  • Probe and kit for improving pathogenic microorganism antigen detection sensitivity

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1 Design and preparation of double-stranded DNA probes

[0036] 1. Probe design and screening

[0037] The double-stranded DNA (such as 50bpDNA) with an endonuclease site used for linking antibodies in the present invention is composed of Cy5-B-CM-S and NH2-B-CM-AS, and Cy5-B-CM-S and NH2 A restriction endonuclease site was inserted in the middle of the DNA sequence of -B-CM-AS.

[0038] When no restriction site is inserted, the sequence is shown in SEQ ID No.3 and 4:

[0039] SEQ ID No. 3:

[0040] CM-S-5'-CCCCCATCTCATCCCTGCGTGTCACTGGTTCAAGGTTCTGGAG-3'

[0041] SEQ ID No.4:

[0042] CM-AS-5'-CTCCAGAACCTTGAACCAGTGACACGCAGGGATGAGATGG-3'.

[0043] After inserting a restriction endonuclease site into the above double-stranded DNA, the end of the CM-S oligonucleotide is modified with a fluorescent group, and the end of the CM-AS oligonucleotide is modified with an active amino group. The sequence is as shown in SEQ ID No. .1 and 2 are shown.

[0044] In the pre...

Embodiment 2

[0048] Example 2 Application of this probe to carry out hypersensitive detection of AFP in AFP-positive serum samples

[0049] The present invention takes AFP as an example to explain and illustrate the use method and technical effect of the probe in detail, but it is not limited to the antigen to be tested in the present invention. The probe can be applied to the detection of all single-molecule diagnostic antigens, and especially has a hypersensitive detection effect on trace antigens. The ultrasensitive detection principle lies in the structure design of double-stranded DNA in the present invention. In the present invention, the double-stranded DNA is linked to the detection antibody, which does not affect the structure of the detection antibody, nor does it affect the combination of antigen and antibody.

[0050] 1. Use the kit to label the coating and detection antibodies

[0051] 1. According to the ChromaLink Biotin One-Shot Antibody-Labeling Kit method, biotin was co...

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Abstract

The invention discloses a probe for improving antigen detection sensitivity of pathogenic microorganisms and a kit containing the probe. The probe is double-stranded DNA which contains a fluorescence label and is provided with a restriction enzyme site, and the double-stranded DNA consists of Cy5-B-CM-S and NH2-B-CM-AS; wherein the Cy5-B-CM-S is as shown in SEQ ID No. 1, and the NH2-B-CM-AS is as shown in SEQ ID No. 2. The probe is combined with a detection antibody of a to-be-detected antigen, so that the detection background can be greatly reduced, the detection sensitivity of the pathogenic microorganism antigen is improved, and a foundation is laid for research and development of a single-molecule diagnostic reagent of the pathogenic microorganism antigen. Meanwhile, the probe restriction enzyme design can realize rapid detection of antigens.

Description

technical field [0001] The invention relates to a probe, in particular to a probe for improving the detection sensitivity of pathogenic microorganism antigen and a kit containing the probe. Background technique [0002] The diagnosis of pathogenic microbial antigens has undergone significant changes in the past 30 years, from the early ELISA semi-quantitative to chemiluminescent quantitative, the accuracy has been greatly improved, and the sensitivity has also increased from micrograms per liter to nanograms. The conventional linear range of traditional ELISA is 10pg / ml~1000pg / ml. For a biological experiment, it is often necessary to take into account the samples of the normal control group and the disease group. The concentration distribution of the protein to be tested in the sample generally ranges from a few tenths of a pg to several thousand pg. Not equal, so the linear range of an ELISA kit cannot take into account the detection of high and low abundance proteins at th...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/576G01N33/543G01N33/536G01N33/53G01N21/64C12N15/11
CPCG01N21/6428G01N33/5306G01N33/536G01N33/54326G01N33/56983G01N33/56988G01N33/5761
Inventor 庞丽君乔录新马迎民陈德喜金荣华李文
Owner BEIJING INST OF HEPATOLOGY
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