CN-121991184-A - Monoclonal antibody for detecting rotavirus in specific type and application thereof

Abstract

The invention provides a monoclonal antibody for specifically detecting rotavirus and application thereof, which aims at epidemic strains of porcine rotavirus, refers to a gene sequence of VP6 protein published in GenBank, further optimizes epitope, obtains a wheel-purified recombinant VP6 protein with antigenicity for animal immunization, and successfully screens out the monoclonal antibody capable of specifically recognizing porcine rotavirus. The monoclonal antibody obtained by screening can effectively identify epidemic strains, has broad spectrum identification property, is superior to commercial antibodies, has higher antigen identification targeting property and low non-specificity. And the sequence background is clear, the large-scale amplification and production can be carried out, and the method has good commercial value.

Inventors

• ZHOU ZHIPING
• MAO LI
• LIU JI

Assignees

• 尼沃诺斯(苏州)生物工程有限公司

Dates

Publication Date

20260508

Application Date

20250224

Claims (6)

1. 1. An antibody or antigen-binding fragment, characterized in that the antibody or antigen-binding fragment comprises light chain variable regions LCDR1, LCDR2, LCDR3 and heavy chain variable regions HCDR1, HCDR2, HCDR3, wherein the amino acid sequence of LCDR1 is shown in SEQ ID No. 7, the amino acid sequence of LCDR2 is shown in SEQ ID No. 8, and the amino acid sequence of LCDR3 is shown in SEQ ID No. 9, the amino acid sequence of HCDR1 is shown in SEQ ID No. 4, the amino acid sequence of HCDR2 is shown in SEQ ID No. 5, and the amino acid sequence of HCDR3 is shown in SEQ ID No. 6.
2. 2. A nucleic acid molecule encoding an antibody or antigen-binding fragment of claim 1.
3. 3. An expression vector comprising the nucleic acid molecule of claim 2.
4. 4. A host cell comprising the nucleic acid molecule of claim 2, or the expression vector of claim 3.
5. 5. A composition comprising the antibody or antigen-binding fragment of claim 1.
6. 6. Use of the antibody or antigen binding fragment of claim 1 for in vitro detection of rotavirus or VP6 protein of rotavirus, said use being in the diagnosis of a non-disease.

Description

Monoclonal antibody for detecting rotavirus in specific type and application thereof Technical Field The invention belongs to the field of virus detection, and in particular relates to a monoclonal antibody for specifically detecting rotavirus and a rotavirus detection kit containing the antibody. Background Rotavirus (Rotavirus, RV) belongs to the family reoviridae, genus rotavirus, was first discovered in 1973 by Bishop et al, and was named rotavirus because it appears as a spoke of a wheel under electron microscopy. The complete rotavirus particle has a three-layer capsid whose genome is composed of 11 double stranded RNA segments, each gene segment encoding a protein, gene segment 6 being responsible for encoding the inner envelope structural protein VP6. The VP6 protein is the highest protein in rotavirus, is the virus group (subgroup) antigen protein, plays an important role in the assembly process of virus particles, plays a role in the assembly of inner shell protein genome RNA and the entry of outer shell protein into cells as a physical receptor, and is essential for the transcription of the virus, so that VP6 is widely applied to the detection of rotavirus. Since rotaviruses are RNA viruses with high variability and multiple serotypes (G-type and P-type) exist, and the infectivity of different serotypes on different hosts is also different, the results show that the A, B, C group of porcine rotaviruses all have different degrees of epidemic, but RVA is only detected in the group of pigs less than 10 weeks of age, and the genotypes are G2, G5 and G9. The disease is difficult to identify clinically with other bacterial and viral diarrhea diseases, and whether accurate diagnosis can be achieved is of great importance in treating infection of the virus. The existing reagent kit for detecting rotavirus is mainly used for clinical detection, and is mainly used for detecting rotavirus antigen, and no reagent kit for detecting RV antibody is available. Therefore, it is necessary to develop a colloidal gold test strip for rapidly, conveniently and specifically detecting rotavirus antibodies. In addition, VP6 protein commercialized monoclonal antibodies exist at present, but most of the prepared antigens are obtained by animal immunization of fully human rotaviruses, the combination level of the antigen and porcine rotaviruses is not high, and the large-scale use is limited due to the problems of commercial antibody supply, high price and the like. Disclosure of Invention In order to solve the problems, the invention further optimizes the antigen epitope based on the previous research (CN 110470829A), screens and obtains the rotavirus structural protein VP6 with antigenicity, and the gene sequence of the recombinant protein VP6 is shown as SEQ ID NO.1 （GSKDARDKIGGSGNDFQTGIGGSVENARGGARESQRNGIAPQSEGSSFDANSSDYIENWNLQNRRQGGNRSQPAHDNGSNAPKANIGRFSFPRVINSHAGSDGATTGSPNMTPAVANLGSFPQGGAIPVGPVFPPMPNWTEGSLITNYSPSREDN）. Furthermore, the fusion protein is taken as an immunogen, and a monoclonal antibody FK14 capable of specifically recognizing porcine rotavirus is obtained by screening, wherein the monoclonal antibody FK14 comprises at least one heavy chain variable region and at least one light chain variable region, the amino acid sequence of the heavy chain variable region is shown as SEQ ID NO.2, and the amino acid sequence of the light chain variable region is shown as SEQ ID NO. 3; further, the heavy chain variable region has an HCDR1 as shown in SEQ ID NO. 4, an HCDR2 as shown in SEQ ID NO. 5, and an HCDR3 as shown in SEQ ID NO. 6; the light chain variable region has LCDR1 as shown in SEQ ID NO. 7, LCDR2 as shown in SEQ ID NO. 8, and LCDR3 as shown in SEQ ID NO. 9. Further, the present invention provides an isolated nucleic acid encoding the monoclonal antibody described above. Further, the present invention provides an expression vector comprising an isolated nucleic acid according to the above. Further, the present invention provides a host cell comprising the isolated nucleic acid described above, or the expression vector described above. Further, the present invention provides an antibody drug conjugate comprising an antibody or antigen-binding fragment thereof prepared by the method described above, or an antibody or antigen-binding fragment thereof described above, and a ligand, preferably selected from the group consisting of a radioisotope, a fluorescent group, and a delivery vehicle. Further, the invention provides a pharmaceutical composition comprising the monoclonal antibody prepared by the method or the antibody drug conjugate and a pharmaceutically acceptable carrier. Furthermore, the invention provides the application of the monoclonal antibody prepared by the method, or the isolated nucleic acid, or the antibody drug conjugate or the drug composition in preparing drugs for preventing, treating and/or diagnosing diseases caused by rotaviruses. Furthermore, the invention provides a kit for detecting