Principle of Method
This assay is a sandwich Enzyme Linked-Immunosorbent Assay (ELISA) for quantitative determination of human ICOSL in cell culture supernatants, serum and plasma. A monoclonal antibody specific for ICOSL has been precoated onto the 96-well microtiter plate. Standards (STD) and samples are pipetted into the wells for binding to the coated antibody. After extensive washing to remove unbound compounds, ICOSL is recognized by the addition of a biotinylated monoclonal antibody specific for ICOSL (DET). After removal of excess biotinylated antibody, streptavidin-peroxidase (STREP-HRP) is added. Following a final washing, peroxidase activity is quantified using the substrate 3,3’,5,5’-tetramethylbenzidine (TMB). The intensity of the color reaction is measured at 450nm after acidification and is directly proportional to the concentration of ICOSL in the samples.
Inducible T-cell costimulatory protein (ICOS, also called CD278, AILIM, H4), a member of the CD28 family of costimulatory receptors, has a role in the generation and maintenance of germinal centers (GCs) in lymphatic organs, induction of thymus-dependent (TD) antibody (Ab) responses, and antibody class switching (1, 2). ICOS has a low expression on naïve T cells but is rapidly induced on activated T cells (1, 2, 3). ICOS binds to the inducible co-stimulator ligand (ICOSL, also called CD275, B7-H2, B7h, B7RP-1) that is found in professional APCs such as dendritic cells (DCs), B lymphocytes, various non-hematopoietic cells such as endothelial cells (ECs), as well as some cancer cells (2, 4). ICOS activated by ICOSL induces T cell proliferation, survival and differentiation and co-induces the secretion of IL-4, IL-5, IL-6, IL-10, IL-21, TNF-α, and interferon gamma (IFN-γ) (whereas CD28 induces IL-2 production) (5). Therefore, ICOS enhances Th1, Th2, and Th17 function largely through augmented production of these effector cytokines. ICOSL is upregulated by TNF-α and other inflammatory mediators and has an important co-stimulation role in EC (endothelial cells)-mediated T-cell activation, especially in reactivation of effector/memory T cells on the endothelium, which promote the homing of immune cells into inflamed tissue (5, 6). As seen in diverse types of costimulatory molecules in the CD28 family in T cells, ICOS is able to deliver a reverse signal through ICOSL that has a direct effect on dendritic cells (7). ICOS/ICOSL is involved in some hematologic malignancies such as myeloma or lymphoma. ICOS and its ligand ICOSL have been shown to play diverse roles in mediating autoimmunity as well as enhancing the development/activity of regulatory T cells (8). ICOS has a dual role in oncogenesis: i) the costimulatory signal of ICOS/ICOSL clearly participates in an antitumor T-cell response; ii) the ICOS signaling also exhibits pro-tumoral features, which are related to the induction of Treg immunosuppressive effects (9, 10, 11). In humans, homozygous ICOS deficiency results in common variable immunodeficiency (CVID), a condition characterized by aberrantly low serum gammaglobulin concentration (12). ICOS/ICOSL pathway is necessary for the optimal therapeutic effect of anti-CTLA-4, thus implicating this pathway as a target for future combinatorial strategies to improve the efficacy of anti-CTLA-4 therapy (13). ICOSL sheds from the cell membrane and the soluble form of ICOSL (sICOSL), found in plasma and sera, can be a potential biomarker for autoimmune diseases and some cancers (8).
Sandwich type ELISA
ICOS Ligand, inducible co-stimulator ligand, CD_antigen: CD275, B7-related protein 1 (B7RP-1), B7 Homologue 2 (B7-H2)
Serum, Plasma or Cell Culture Supernatant
This ELISA is specific for the measurement of natural and recombinant human ICOSL [CD275].
Serum and Plasma : 0.25 µL (actual amount of sample loaded in well after dilution)
Calibration Range: 0.0625 ng/ml – 4 ng/ml
Limit of Detection
Intra-assay (Within-Run) C.V.
Inter-assay (Run-to-Run) C.V.
References to Summary
(1) The past, present, and future of costimulation blockade in organ transplantation: P.M. Schroder, et al.; Curr. Opin. Organ Transplant. (Epub ahead of print) (2019)
(2) Characterization of H4: a mouse T lymphocyte activation molecule functionally associated with the CD3/T cell receptor: V. Redoglia, et al.; Eur. J. Immunol. 26, 2781 (1996)
(3) ICOS is an inducible T-cell co-stimulator structurally and functionally related to CD28: A. Hutloff, et al.; Nature 397, 263 (1999)
(4) Inducible Co-Stimulator (ICOS) as a potential therapeutic target for anti-cancer therapy: F. Amatore, et al.; Expert Opin. Ther. Targets 22, 343 (2018)
(5) ICOS co-stimulation: friend or foe? D.J. Wikenheiser & J.S. Stumhofer; Front. Immunol. 7, 304 (2016)
(6) B7h, a novel costimulatory homolog of B7.1 and B7.2, is induced by TNFalpha: M.M. Swallow, et al.; Immunity 11, 423 (1999)
(7) Reverse signaling using an inducible costimulator to enhance immunogenic function of dendritic cells: G. Tang, et al.; Cell Mol. Life Sci. 66, 3067 (2009)
(8) Increased expression of soluble inducible costimulator ligand (ICOSL) in patients with systemic lupus erythematosus: M. Her, et al.; Lupus 18, 501 (2009)
(9) Melanoma cells express ICOS ligand to promote the activation and expansion of T- regulatory cells: N. Martin-Orozco, et al.; Cancer Res. 70, 9581 (2010)
(10) The inducible costimulator augments Tc17 cell responses to self and tumor tissue: M.H. Nelson, et al.; J. Immunol. 194, 1737 (2015)