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CN-122011186-A - Monoclonal antibody combination for detecting human IL-6 protein and application

CN122011186ACN 122011186 ACN122011186 ACN 122011186ACN-122011186-A

Abstract

The invention relates to the field of biological detection, in particular to a monoclonal antibody combination for detecting human IL-6 protein and application thereof. The combination comprises monoclonal antibodies 3A10 and 2G1, and the amino acid sequences of the complementarity determining regions of the light and heavy chain variable regions are respectively shown in SEQ ID NO. 1-12. The invention defines the complete variable region sequence and the coding nucleotide sequence of the antibody. The combination specifically recognizes human IL-6 recombinant and natural proteins, and has no cross reaction with IL-11. The double-antibody sandwich ELISA kit constructed based on the combination takes 3A10 as a coating antibody and 2G1 as a labeling antibody, and has high sensitivity and high specificity. The invention solves the problems of insufficient specificity and low sensitivity of the existing reagent, provides a reliable core raw material for the standardized detection of IL-6, and is suitable for preparing a kit, a test strip and an antibody chip.

Inventors

  • QIN KUN
  • LU SHUAI
  • LI YUE
  • SUN YUTIAN
  • ZHONG MING
  • SU DU

Assignees

  • 北京溯本源和生物科技有限公司

Dates

Publication Date
20260512
Application Date
20260414

Claims (10)

  1. 1. A monoclonal antibody combination for detecting human IL-6 protein, wherein the monoclonal antibody combination comprises monoclonal antibody 3a10 and monoclonal antibody 2G1; The heavy chain variable region of the monoclonal antibody 3A10 comprises three complementarity determining regions, and the amino acid sequences of the complementarity determining regions are respectively shown as SEQ ID NO.1-SEQ ID NO. 3; The light chain variable region of the monoclonal antibody 3A10 comprises three complementarity determining regions, and the amino acid sequences of the complementarity determining regions are respectively shown in SEQ ID NO.4-SEQ ID NO. 6; the heavy chain variable region of the monoclonal antibody 2G1 comprises three complementarity determining regions, and the amino acid sequences of the complementarity determining regions are respectively shown in SEQ ID NO.7-SEQ ID NO. 9; The light chain variable region of the monoclonal antibody 2G1 comprises three complementarity determining regions, and the amino acid sequences of the complementarity determining regions are respectively shown in SEQ ID NO.10-SEQ ID NO. 12.
  2. 2. The monoclonal antibody combination for detecting human IL-6 protein according to claim 1, wherein the amino acid sequence of the heavy chain variable region of the monoclonal antibody 3A10 is shown as SEQ ID NO.13, and the amino acid sequence of the light chain variable region of the monoclonal antibody 3A10 is shown as SEQ ID NO. 14.
  3. 3. The monoclonal antibody combination for detecting human IL-6 protein according to claim 2, wherein the amino acid sequence of the heavy chain variable region of the monoclonal antibody 2G1 is shown as SEQ ID NO.15, and the amino acid sequence of the light chain variable region of the monoclonal antibody 2G1 is shown as SEQ ID NO. 16.
  4. 4. The monoclonal antibody combination for detecting human IL-6 protein according to claim 3, wherein the nucleotide sequence encoding the heavy chain variable region of the monoclonal antibody 3A10 is shown in SEQ ID NO.17 and the nucleotide sequence encoding the light chain variable region of the monoclonal antibody 3A10 is shown in SEQ ID NO. 18.
  5. 5. The monoclonal antibody combination for detecting human IL-6 protein according to claim 4, wherein the nucleotide sequence encoding the heavy chain variable region of the monoclonal antibody 2G1 is shown as SEQ ID NO.19 and the nucleotide sequence encoding the light chain variable region of the monoclonal antibody 2G1 is shown as SEQ ID NO. 20.
  6. 6. The monoclonal antibody combination for detecting human IL-6 protein according to claim 5, wherein the monoclonal antibody combination specifically recognizes human IL-6 recombinant protein and human IL-6 native protein.
  7. 7. Use of a combination of monoclonal antibodies according to claim 1 for the preparation of a means for detecting human IL-6 protein for in vitro detection of human IL-6 protein in a sample selected from any one of serum, plasma, cell culture supernatant and tissue homogenate, and not for diagnosis of a disease.
  8. 8. The use of claim 7, wherein the means comprises reagents, kits, test strips and antibody chips.
  9. 9. The use according to claim 8, wherein the kit comprises a double antibody sandwich ELISA kit.
  10. 10. The use according to claim 9, wherein the ELISA kit employs monoclonal antibody 3a10 as the coating antibody and monoclonal antibody 2G1 as the labeling antibody.

Description

Monoclonal antibody combination for detecting human IL-6 protein and application Technical Field The invention relates to the field of biological detection, in particular to a monoclonal antibody combination for detecting human IL-6 protein and application thereof. Background Human Interleukin-6 (IL-6) is a multifunctional cytokine belonging to the IL-6 cytokine family, which also includes other cytokines such as Interleukin-11 (IL-11) that share gp130 as a signal transduction chain. IL-6 protein space structure is in typical four alpha helical bundle configuration, can regulate cell growth and differentiation of various tissues, and can secrete IL-6 when various cells such as mononuclear/macrophages, T lymphocytes, B lymphocytes, fibroblasts, endothelial cells and the like are stimulated. When the body is in an infectious, tissue damaging or inflammatory state, the expression level of IL-6 is obviously increased under the induction of Toll-like receptor signals and inflammatory factors such as IL-1, TNF-alpha and the like, and the content of the IL-6 in serum of a healthy individual is usually kept at a lower level. In addition, IL-6 is also involved in hematogenesis, bone metabolism and cancer progression and is determined to play a key role in the leading of the transition from innate immunity to acquired immunity. In a variety of disease states, abnormal changes in IL-6 levels are closely related to the occurrence and progression of the disease. For example, in chronic inflammatory diseases such as rheumatoid arthritis, an increased continuous IL-6 can promote the amplification of inflammatory response and tissue damage, IL-6 is often used as an index reflecting the intensity of inflammation in patients with severe infection or systemic inflammatory response syndrome, and in some tumor microenvironments, IL-6 is also associated with tumor cell survival and altered immunoregulatory status. Therefore, the IL-6 is reliably detected, so that the method has reference value for disease monitoring and provides important basis for related mechanism research. Currently, detection of IL-6 protein levels is mostly accomplished by immunological methods, and common sample types include serum, plasma, cell culture supernatants, tissue homogenates, and the like. Chemiluminescent immunoassay and enzyme-linked immunosorbent assay (ELISA) are relatively common technical routes, wherein the double-antibody sandwich ELISA is relatively mature in operation, convenient for quantitative analysis and relatively wide in application in cytokine detection. However, in practical application, the existing IL-6 detection reagent still has the problems of insufficient antibody specificity, easy occurrence of cross reaction and low sensitivity, and is difficult to meet the requirement of developing a standardized reagent. Therefore, obtaining the human IL-6 monoclonal antibody with high affinity, high specificity and definite sequence, and screening out the optimal pairing antibody combination is the key for developing high-performance standardized ELISA reagents. Disclosure of Invention First, the technical problem to be solved In view of the above-mentioned shortcomings and disadvantages of the prior art, the present invention provides a monoclonal antibody combination for detecting human IL-6 protein and application thereof, which solves the technical problems of insufficient antibody specificity, easy cross reaction and low sensitivity in the existing IL-6 detection reagent, and difficult to satisfy the development requirement of standardized reagent. (II) technical scheme In order to achieve the above purpose, the main technical scheme adopted by the invention comprises the following steps: in a first aspect, the invention provides a monoclonal antibody combination for detecting human IL-6 protein, the monoclonal antibody combination comprising monoclonal antibody 3a10 and monoclonal antibody 2G1; The heavy chain variable region of the monoclonal antibody 3A10 comprises three complementarity determining regions, and the amino acid sequences of the complementarity determining regions are respectively shown as SEQ ID NO.1-SEQ ID NO. 3; The light chain variable region of the monoclonal antibody 3A10 comprises three complementarity determining regions, and the amino acid sequences of the complementarity determining regions are respectively shown in SEQ ID NO.4-SEQ ID NO. 6; the heavy chain variable region of the monoclonal antibody 2G1 comprises three complementarity determining regions, and the amino acid sequences of the complementarity determining regions are respectively shown in SEQ ID NO.7-SEQ ID NO. 9; The light chain variable region of the monoclonal antibody 2G1 comprises three complementarity determining regions, and the amino acid sequences of the complementarity determining regions are respectively shown in SEQ ID NO.10-SEQ ID NO. 12. In some embodiments, the amino acid sequence of the heavy chain variable region of the m