Monoclonal antibody composition for quantitatively detecting coxiella burnetii I strain

文档序号：1542533 发布日期：2020-01-17 浏览：26次中文

阅读说明：本技术 一种用于定量检测贝氏柯克斯体i相株的单抗组合物 (Monoclonal antibody composition for quantitatively detecting coxiella burnetii I strain ) 是由熊小路张平平杨瑞馥焦俊赵勇王津周冬生于 2019-10-28 设计创作，主要内容包括：本发明提供一种用于贝氏柯克斯体I相株定量检测的单克隆抗体,该单克隆抗体克结合上转发光技术进行检测,跟传统的贝氏柯克斯体检测方法相比,具有准确定量、零背景干扰,检测结果稳定,检测简便快速的特点。(The invention provides a monoclonal antibody for quantitatively detecting a coxiella burnetii I phase strain, which is detected by combining an up-conversion luminescence technology and has the characteristics of accurate quantification, zero background interference, stable detection result and simple and quick detection compared with the traditional coxiella burnetii detection method.)

1. The test paper for detecting the coxiella burnetii I strain is characterized by comprising a combination pad and an analysis membrane connected with the combination pad; the combination pad is coated with UCP-monoclonal antibody 10 compound, and the UCP-monoclonal antibody 10 compound is obtained by labeling the UCP with the UCP monoclonal antibody 10;

the analysis membrane is provided with a detection band and a quality control band which are separated from each other, and the detection band is coated with a monoclonal antibody 10;

the monoclonal antibody 10 is a monoclonal antibody composed of a heavy chain and a light chain, both of which are composed of a variable region and a constant region, both of which are composed of a determinant complementary region and a framework region, both of which are composed of CDR1, CDR2 and CDR 3; the amino acid sequence of CDR1 of the heavy chain of the monoclonal antibody 10 is shown in the 26 th to 33 th positions of SEQ ID No. 4; the amino acid sequence of CDR2 of the heavy chain of the monoclonal antibody 10 is shown in 51-60 th position of SEQ ID No. 4; the amino acid sequence of CDR3 of the heavy chain of the monoclonal antibody 10 is shown in the 99 th to 106 th positions of SEQ ID No. 4; the amino acid sequence of CDR1 of the light chain of the monoclonal antibody 10 is shown in 26 th to 32 th positions of SEQ ID No. 8; the amino acid sequence of CDR2 of the light chain of the monoclonal antibody 10 is shown in the 50 th to 52 th positions of SEQ ID No. 8; the amino acid sequence of CDR3 of the light chain of monoclonal antibody 10 is shown in 89-97 th position of SEQ ID No. 8.

2. The test paper of claim 1, wherein the amino acid sequence of the heavy chain variable region of mab 10 is position 1-117 of seq id No. 4; the amino acid sequence of the variable region of the light chain of the monoclonal antibody 10 is1 st to 107 th in a sequence 8.

3. The test paper of claim 1 or 2, wherein the amino acid sequence of the heavy chain of the monoclonal antibody 10 is sequence 4; the amino acid sequence of the light chain of the monoclonal antibody 10 is sequence 8.

4. The strip of any one of claims 1 to 3, wherein said control band is coated with a second antibody that specifically binds to said UCP-mAb 10 complex.

5. A monoclonal antibody, which is the mab 10 of any of claims 1-3.

6. The monoclonal antibody of claim 5, which is a murine monoclonal antibody.

7. A biomaterial related to the monoclonal antibody as claimed in claim 5 or 6, said biomaterial being any one of B1) to B16):

B1) a nucleic acid molecule encoding the monoclonal antibody of claim 5 or 6;

B2) an expression cassette comprising the nucleic acid molecule of B1);

B3) a recombinant vector comprising the nucleic acid molecule of B1);

B4) a recombinant vector comprising the expression cassette of B2);

B5) a recombinant microorganism comprising the nucleic acid molecule of B1);

B6) a recombinant microorganism comprising the expression cassette of B2);

B7) a recombinant microorganism containing the recombinant vector of B3);

B8) a recombinant microorganism containing the recombinant vector of B4);

B9) a transgenic animal cell line comprising the nucleic acid molecule of B1);

B10) a transgenic animal cell line comprising the expression cassette of B2);

B11) a transgenic animal cell line containing the recombinant vector of B3);

B12) a transgenic animal cell line containing the recombinant vector of B4);

B13) a transgenic plant cell line comprising the nucleic acid molecule of B1);

B14) a transgenic plant cell line comprising the expression cassette of B2);

B15) a transgenic plant cell line comprising the recombinant vector of B3);

B16) a transgenic plant cell line comprising the recombinant vector of B4).

8. The biomaterial of claim 7, wherein: B1) the nucleic acid molecule is a gene encoding the monoclonal antibody of claim 5 or 6.

9. The biomaterial of claim 8, wherein: the gene is the DNA molecule described in the following A) or B):

A) the coding sequence of CDR1 of the heavy chain of the monoclonal antibody 10 is shown as 76-99 th position of SEQ ID No. 3; the coding sequence of the CDR2 of the heavy chain of the monoclonal antibody 10 is shown as the 151 th-180 th position of SEQ ID No. 3; the coding sequence of the CDR3 of the heavy chain of the monoclonal antibody 10 is shown as the 295-318 bit of SEQ ID No. 3;

the coding sequence of CDR1 of the light chain of the monoclonal antibody 10 is shown in 79-96 th position of SEQ ID No. 7; the coding sequence of the CDR2 of the light chain of the monoclonal antibody 10 is shown as 148-156 bit of SEQ ID No. 7; the coding sequence of CDR3 of the light chain of monoclonal antibody 10 is shown as 267-291 bit of SEQ ID No. 7;

B) DNA having 90% or more identity to the DNA molecule defined in A) and encoding the monoclonal antibody.

10. A composition for detecting coxiella burnetii strain I, wherein the active ingredient of the composition is mab 10 of any one of claims 1-3.

Technical Field

The invention relates to a monoclonal antibody composition for quantitatively detecting coxiella burnetii I phase strain.

Background

Coxiella burnetii (Coxiella burnetii) is an important zoonosis-pathogenic bacterium of Q fever (Qfever), gram-negative staining, obligate intracellular parasitism, and small short rod shape or small ball rod shape. Phase change exists in the growth process of Coxiella burnetii, the phase I strain is usually a virulent strain separated from Q fever patients or infected animals, and partial antigen of LPS of the phase I strain is lost after tens or hundreds of generations of laboratory artificial passage, so that an attenuated strain, namely the phase II strain is formed. The Coxiella burnetii phase I strain contains phase I and phase II antigens and can induce animals to generate phase I and phase II antibodies, while the phase II strain is mainly phase II antigen and can only induce phase II antibodies.

The clinical symptoms of acute Q fever mainly include acute fever, headache and muscular soreness, and are often accompanied by pneumonia, hepatitis and the like. Chronic Q fever is often complicated with serious diseases such as endocarditis and osteomyelitis. Q fever has no specific symptoms and signs in clinic and lacks specific clinical diagnosis markers, so that Q fever is difficult to distinguish from other febrile infectious diseases and has high misdiagnosis rate. Serological detection of coxiella burnetii specific antibodies is the most common method, including Complement Fixation (CF), enzyme-linked immunosorbent assay (ELISA), indirect Immunofluorescence (IFA), Microaggregation (MA), etc.; the molecular biological detection method comprises Polymerase Chain Reaction (PCR), real-time fluorescence quantitative PCR, recombinase polymerase amplification Reaction (RPA), loop-mediated isothermal amplification (Lamp), DNA dot hybridization and the like. The serological detection method has strong specialization, high biological safety requirement and high technical requirement, and patients with acute disease often cannot detect the antibody and the separation of the pathogen from the blood is very difficult; the genome needs to be extracted in advance during nucleic acid detection, the operation is time-consuming and labor-consuming, professional instruments and equipment are needed, the operation needs to be carried out in a professional laboratory, and meanwhile, the nucleic acid detection method cannot judge whether the thalli survive or not, so that the result judgment and the infection risk evaluation are easy to be complicated.

With the development of related disciplines such as chemistry and materials science and the continuous expansion of the social application field, in recent years, the immunodiagnosis technology is developing towards the stable, high-flux, high-sensitivity and wide-field direction. For example, the Up-Converting Phosphor Technology (UPT) and the like, it has strong tolerance to various complex detection samples on the basis of improving the sensitivity of the traditional immunochromatography, and makes it possible to directly carry out quantitative detection on trace-level antibodies or antigens in the serum of patients or animals.

Disclosure of Invention

The invention provides a monoclonal antibody composition for quantitatively detecting coxiella burnetii I phase strain.

The invention provides test paper for detecting coxiella burnetii, which comprises a combination pad and an analysis membrane connected with the combination pad; the combination pad is coated with UCP-monoclonal antibody 10 compound, and the UCP-monoclonal antibody 10 compound is obtained by labeling the UCP with the UCP monoclonal antibody 10;

the analysis membrane is provided with a detection band and a quality control band which are separated from each other, and the detection band is coated with a monoclonal antibody 10;

the monoclonal antibody 10 is a monoclonal antibody composed of a heavy chain and a light chain, both of which are composed of a variable region and a constant region, both of which are composed of a determinant complementary region and a framework region, both of which are composed of CDR1, CDR2 and CDR 3; the amino acid sequence of CDR1 of the heavy chain of the monoclonal antibody 10 is shown in the 26 th to 33 th positions of SEQ ID No. 4; the amino acid sequence of CDR2 of the heavy chain of the monoclonal antibody 10 is shown in 51-60 th position of SEQ ID No. 4; the amino acid sequence of CDR3 of the heavy chain of the monoclonal antibody 10 is shown in the 99 th to 106 th positions of SEQ ID No. 4; the amino acid sequence of CDR1 of the light chain of the monoclonal antibody 10 is shown in 26 th to 32 th positions of SEQ ID No. 8; the amino acid sequence of CDR2 of the light chain of the monoclonal antibody 10 is shown in the 50 th to 52 th positions of SEQ ID No. 8; the amino acid sequence of CDR3 of the light chain of the monoclonal antibody 10 is shown in 89-97 th position of SEQ ID No. 8; .

Wherein, the amino acid sequence of the heavy chain variable region of the monoclonal antibody 10 is 1-117 th position in SEQ ID No. 4; the amino acid sequence of the variable region of the light chain of the monoclonal antibody 10 is1 st to 107 th in SEQ ID No. 8.

Wherein, the amino acid sequence of the heavy chain of the monoclonal antibody 10 is SEQ ID No. 4; the amino acid sequence of the light chain of the monoclonal antibody 10 is SEQ ID No. 8.

Wherein the quality control band is coated with a second antibody specifically binding to the UCP-mAb 10 complex.

The invention also provides a monoclonal antibody, which is the monoclonal antibody 10.

The monoclonal antibody is a murine monoclonal antibody.

A biomaterial related to said monoclonal antibody, said biomaterial being any one of B1) to B16):

B1) a nucleic acid molecule encoding the monoclonal antibody of claim;