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CN-122011185-A - Monoclonal antibody combination for detecting human IL-10 protein and application

CN122011185ACN 122011185 ACN122011185 ACN 122011185ACN-122011185-A

Abstract

The invention relates to the field of biological detection, in particular to a monoclonal antibody combination for detecting human IL-10 protein and application thereof. The antibody combination comprises 8E10 and 10D12, wherein the amino acid sequence of the complementarity determining region of the heavy chain variable region of the monoclonal antibody 8E10 corresponds to SEQ ID NO.1-SEQ ID NO.3, the CDR amino acid sequence of the light chain variable region corresponds to SEQ ID NO.4-SEQ ID NO.6, the CDR amino acid sequence of the heavy chain variable region of the monoclonal antibody 10D12 corresponds to SEQ ID NO.7-SEQ ID NO.9, and the CDR amino acid sequence of the light chain variable region corresponds to SEQ ID NO.10-SEQ ID NO.12. The antibody combination ensures the production repeatability of the raw materials, provides a reliable basis for preparing diagnosis tools such as ELISA kits with high sensitivity and high stability, and has wide application prospect.

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
20260413

Claims (10)

  1. 1. A monoclonal antibody combination for detecting human IL-10 protein, wherein the monoclonal antibody combination comprises monoclonal antibody 8E10 and monoclonal antibody 10D12; The heavy chain variable region of the monoclonal antibody 8E10 comprises three complementarity determining regions CDR-H1, CDR-H2 and CDR-H3, wherein the amino acid sequence of the CDR-H1 is shown as SEQ ID N0.1, the amino acid sequence of the CDR-H2 is shown as SEQ ID N0.2, and the amino acid sequence of the CDR-H3 is shown as SEQ ID NO. 3; The light chain variable region of the monoclonal antibody 8E10 comprises three complementarity determining regions CDR-L1, CDR-L2 and CDR-L3, wherein the amino acid sequence of the CDR-L1 is shown as SEQ ID NO.4, the amino acid sequence of the CDR-L2 is shown as SEQ ID NO.5, and the amino acid sequence of the CDR-L3 is shown as SEQ ID NO. 6; The heavy chain variable region of the monoclonal antibody 10D12 comprises three complementarity determining regions CDR-H1, CDR-H2 and CDR-H3, wherein the amino acid sequence of the CDR-H1 is shown as SEQ ID N0.7, the amino acid sequence of the CDR-H2 is shown as SEQ ID N0.8, and the amino acid sequence of the CDR-H3 is shown as SEQ ID NO. 9; The light chain variable region of the monoclonal antibody 10D12 comprises three complementarity determining regions CDR-L1, CDR-L2 and CDR-L3, wherein the amino acid sequence of the CDR-L1 is shown as SEQ ID NO.10, the amino acid sequence of the CDR-L2 is shown as SEQ ID NO.11, and the amino acid sequence of the CDR-L3 is shown as SEQ ID NO. 12.
  2. 2. The monoclonal antibody combination for detecting human IL-10 protein according to claim 1, wherein the amino acid sequence of the heavy chain variable region of the monoclonal antibody 8E10 is shown in SEQ ID NO.13, and the amino acid sequence of the light chain variable region of the monoclonal antibody 8E10 is shown in SEQ ID NO. 14.
  3. 3. The monoclonal antibody combination for detecting human IL-10 protein according to claim 2, wherein the amino acid sequence of the heavy chain variable region of monoclonal antibody 10D12 is shown in SEQ ID No.15, and the amino acid sequence of the light chain variable region of monoclonal antibody 10D12 is shown in SEQ ID No. 16.
  4. 4. The monoclonal antibody combination for detecting human IL-10 protein according to claim 3, wherein the nucleotide sequence encoding the heavy chain variable region of the monoclonal antibody 8E10 is shown in SEQ ID NO.17 and the nucleotide sequence encoding the light chain variable region of the monoclonal antibody 8E10 is shown in SEQ ID NO. 18.
  5. 5. The monoclonal antibody combination for detecting human IL-10 protein according to claim 4, wherein the nucleotide sequence encoding the heavy chain variable region of the monoclonal antibody 10D12 is shown in SEQ ID NO.19 and the nucleotide sequence encoding the light chain variable region of the monoclonal antibody 10D12 is shown in SEQ ID NO. 20.
  6. 6. The monoclonal antibody combination for detecting human IL-10 protein according to claim 5, wherein the monoclonal antibody combination specifically recognizes human IL-10 recombinant protein and human IL-10 native protein.
  7. 7. Use of a combination based on the monoclonal antibody of claim 1 for the preparation of a tool for detecting human IL-10 protein.
  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 8E10 as the coating antibody and monoclonal antibody 10D12 as the labeling antibody.

Description

Monoclonal antibody combination for detecting human IL-10 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-10 protein and application thereof. Background Interlukin-10 (IL-10) is an important pleiotropic immunoregulatory cytokine that plays a central role in regulating inflammation and maintaining immune homeostasis in the body. Human IL-10 is encoded by an IL-10 gene located on the long arm of chromosome 1, which is a soluble homodimer with a native bioactive form of about 36kDa, consisting of two monomers. Each monomer has a typical six alpha helical structure and is conformationally stable by intrachain disulfide bonds. IL-10 belongs to the class II cytokine family and mediates signal transduction through its specific receptor IL-10R (belonging to the class II cytokine receptor family, CRF 2), which is classified according to the structural characteristics of the cytokine receptor. The biological effects of IL-10 begin with the binding of IL-10 homodimers to the cell surface hetero-tetrameric IL-10 receptor complex (IL-10R), thereby activating a signaling pathway with JAK1/TYK2-STAT3 as a core, regulating the expression of various immune-related genes. IL-10 acts on the immune system as a double-edged sword which, on the one hand, is a potent anti-inflammatory and immunosuppressive cytokine and, on the other hand, also has immunostimulatory properties. The different sources of IL-10, the target cell types to which it acts, and the secretion site and time are key factors in activating various signal transduction pathways, which can trigger the inhibition or activation of the signal pathways of immune cells, respectively, under different microenvironments. During the acute infection phase, IL-10 helps limit the intensity of inflammatory responses, prevent immune-mediated tissue damage, and promote inflammation regression and tissue repair after pathogen clearance. However, when IL-10 expression or signaling continues to be enhanced, it is possible to suppress an effective anti-infective immune response, induce immune tolerance and immune escape, promote long-term survival of pathogens, and thus promote the formation of chronic or latent infections. For example, IL-10 inhibits pathogenic inflammatory responses in acute toxoplasma infection, whereas in chronic infections of toxoplasma, leishmania, epstein-Barr virus, HIV and hepatitis B virus, high expression is closely related to the inhibition of T cell function and enhanced replication of the pathogen. At the cellular level, IL-10 has a significant inhibitory effect on myeloid cells such as monocytes, macrophages and dendritic cells, which normally express IL-10R at a high level. IL-10 inhibits immune function by blocking the synthesis of pro-inflammatory cytokines (e.g., IL-1, IL-6, IFN-gamma, and TNF-alpha) in T cells, monocytes, and macrophages, as well as inhibiting the expression of cell surface molecules involved in antigen presentation and co-stimulation. At the same time, IL-10 may also exert immunopotentiating effects on lymphocyte populations under specific immune environments, such as promoting B cell survival and antibody production, or improving CD8 + T cell function in certain chronic infectious and oncological environments, representing an immunostimulatory aspect thereof. Given its key role in inflammatory regulation and maintenance of immune homeostasis, IL-10 has become an important biomarker in infectious disease progression assessment, sepsis immune state monitoring, autoimmune disease activity judgment, and tumor immune microenvironment research. The detection sample type of IL-10 is relatively wide, and clinically mainly comprises serum, plasma and whole blood, and the detection sample type can also detect the IL-10 level in peripheral blood mononuclear cell culture supernatant, tissue homogenate and inflammation-related body fluid (such as cerebrospinal fluid and hydrothorax and ascites) in the scientific research field. Currently, quantitative detection of IL-10 is mostly carried out by adopting an enzyme-linked immunosorbent assay (ELISA), a chemiluminescent immunoassay and a multi-factor combined detection platform. The double-antibody sandwich ELISA has the advantages of mature method, simple operation, relatively low cost, high detection flux, stable result, convenient standardization and the like, and is still one of the most widely applied technical means in cytokine detection. The method relies on two high-quality monoclonal antibodies which recognize different antigen epitopes as a capture antibody and a detection antibody respectively, and the detection performance (such as sensitivity and specificity) of the method is greatly dependent on the affinity, the epitope and the stability of the antibody. Therefore, developing monoclonal antibodies with different epitopes, high affinity, strong specificity