![]() IL-17 has been widely investigated in human medicine during the last decade. The receptor IL-17RA/RC can be recognized by the homodimers of IL-17A or IL-17F and by the heterodimer formed by IL-17 and IL-17F. ![]() The IL-17 receptor family includes IL-17RA, IL-17RB, IL-17RC, IL-17RD and IL-17RE, among which, IL-17RA is a common subunit, and each of the remaining subunits can form a heterodimer with IL-17RA. The receptors for IL-17 are widely distributed on various types of tissue cells, especially on epithelial cells and immune cells. There are various types of IL-17-secreting cells, including T helper 17 lymphocytes (Th17 cells), IL-17-secreting CD8 + cytotoxic T lymphocytes (Tc17 cells), γδ TCR + T lymphocytes (γδ T cells), natural killer T cells (NKT cells), and two subsets of innate lymphoid cells (ILCs), i.e., lymphoid tissue–inducer cells (LTi cells) and RORγt + NCR − ILCs. The commonly denoted “IL-17” refers to IL-17A, while IL-17E is also named IL-25. The IL-17 family consists of six members: IL-17A, IL-17B, IL-17C, IL-17D, IL-17E, and IL-17F. Interleukin-17 (IL-17), which was first named cytotoxic T-lymphocyte-associated antigen 8 (CTLA-8), is a type of pro-inflammatory cytokine that is mainly produced by T lymphocytes. ![]() The monoclonal antibody mAb H8 prepared in this study may be a potential tool for the detection of cIL-17A and beneficial for investigating the pathogenesis of various IL-17-associated diseases. Immunofluorescence and immunohistochemistry assays showed that mAb H8 could strongly recognize both the eukaryotically expressed and natural cIL-17A. Western blot analysis showed that the mAb H8 could recognize the eukaryotically expressed cIL-17A in the supernatant of transfected HEK293T cells. Our results showed that the maximum amount of soluble protein could be obtained directly in the supernatant when the recombinant expression cells were induced by isopropyl-β-d-thiogalactoside (IPTG) at a concentration of 0.3 mmol/L at 16 ☌ for 42 h. Finally, the supernatants of two hybridoma cell lines showing positive reaction with the recombinant fusion protein and negative reaction with fusion tags of PET 32a were collected for western blot, immunofluorescence (IF) and immunohistochemistry (IHC) analysis. Recombinant fusion protein obtained under optimized conditions was used to immunize BALB/c mice for preparing monoclonal antibodies. The coding sequence (CDS) region of cIL-17A was cloned from the peripheral blood mononuclear cells (PBMCs) of dairy goats and then inserted into the expression vector PET 32a and transformed into competent TransB (DE3) cells. The aim of this study was to obtain purified protein caprine IL-17A (cIL-17A) as an antigen for preparing an IL-17A-specific monoclonal antibody (mAb). Interleukin-17 (IL-17), the characteristic cytokine secreted by T helper 17 lymphocytes (Th17 cells), plays a pivotal role in host defense and many inflammatory or autoimmune diseases.
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