96-well microplate

B7-H3 (CD276) Human ELISA

KT1016
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Description

B7 family of immunoregulatory proteins is composed of ten members: B7-1 (CD80), B7-2 (CD86), B7-H1 (PD-L1), B7-DC (PD-L2), B7-H2, B7-H3, B7-H4, B7-H5 (VISTA), B7-H6 and B7-H7. B7-H3 (or CD276) is a 316aa long type I transmembrane protein. B7-H3 shares 20-27% amino acid identity with other B7 family ligands. Glycosylated B7-H3 protein has a molecular weight of approximately 100 kDa (1). A B7-H3 'soluble' isoform has been detected in plasma and B7-H3 is also expressed on exosomes (2).

B7-H3 is ubiquitously expressed by cells in the non-hematopoietic compartment, such as fibroblasts and epithelial cells, it can be induced on T cells and NK cells. Although B7-H3 expression is elevated in tumors, B7-H3 is also constitutively expressed at higher levels in the liver than in other healthy tissue (3). B7-H3 mRNA is expressed in most normal tissues, but the B7-H3 protein expression is limited in normal tissues, because of its posttranscriptional regulation by miRNAs (4). Specifically, miRNA-29 suppresses B7-H3 expression in normal tissues by targeting the B7-H3 3'-untranslated mRNA region.

Little is known about the receptors for these B7-family ligands and their downstream signaling. TLT2 (triggering receptor expressed on myeloid cells-like transcript 2) has been characterized as a putative receptor for B7-H3, but it is dispensable for T cell response, suggesting that other unknown receptors should bind B7-H3 (5). Although receptors remain unidentified, soluble B7-H3 has been shown to bind to CD4+ T, CD8+ T, NK and NKT cells and the extent of its binding is increased upon T cell activation.

B7-H3 inhibits the activation and function of T cells, potently suppressing the proliferation, cytokine production and cytotoxicity of activated T cells. It also inhibits natural killer (NK) cell activation and has a proinflammatory role leading to cytokine release from monocytes and/or macrophages. However, B7-H3 was initially characterized as a co-stimulatory molecule required for optimal promotion of T cell proliferation and cytokine production, which makes the exact role of B7-H3 controversial.

The B7-H3 [CD276] (human) ELISA Kit (Prod. No. KT1016) detects the soluble human B7-H3 (CD276) protein that is ectopically expressed in various cancers (6) and its levels in serum of patients with cancer suggests it may have potential as a non-invasive biomarker for diagnosis, prognosis and/or treatment response.

Type

Sandwich type ELISA

Other names

4Ig-B7-H3, B7 homolog 3, Costimulatory molecule, CD_antigen: CD276

Principle of Method

This assay is a sandwich Enzyme Linked-Immunosorbent Assay (ELISA) for quantitative determination of human B7-H3 [CD276] in cell culture supernatants, serum and plasma. A monoclonal antibody specific for human B7-H3 [CD276] 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, B7-H3 [CD276] is recognized by the addition of a monoclonal biotinylated antibody specific for human B7-H3 [CD276] (DET). After removal of excess biotinylated monoclonal 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 B7-H3 [CD276] in the samples.

Sample Types

Serum, Plasma or Cell Culture Supernatant

Species crossreactivity

This ELISA is specific for the measurement of natural and recombinant human B7-H3 [CD276].

Sample Volume

Serum and Plasma : 0.50 µL (actual amount of sample loaded in well after dilution)

Standard Curve

B7-H3 Standard Curve

Calibration Range: 0.3125 ng/ml – 20 ng/ml

Limit of Detection

<0.3 ng/ml

Intra-assay (Within-Run) C.V.

4.79%

Inter-assay (Run-to-Run) C.V.

4.70%

Resources

References to Summary

  1. B7-H3 in Cancer - Beyond Immune Regulation: F. Flem-Karlsen, et al.; Trends Cancer 6, 401 (2018)

 

  1. B7-H3 in Medulloblastoma-Derived Exosomes; A Novel Tumorigenic Role: I.J. Purvis, et al.; Int. J. Mol. Sci. 19, 7050 (2020)

 

  1. Potential Therapeutic Targets of B7 Family in Colorectal Cancer: C. Wang C. et al.; Immunol. 11, 681 (2020)

 

  1. Regulation of cancer immune escape: The roles of miRNAs in immune checkpoint proteins: Q. Yang, et al.; Cancer Lett. 431, 73 (2018)

 

  1. Triggering receptor expressed on myeloid cell-like transcript 2 (TLT-2) is a counter-receptor for B7-H3 and enhances T cell responses: M. Hashiguchi, et al.; PNAS 105, 10495 (2008)

 

  1. The Role of CD276 in Cancers: S. Liu, et al.; Front. Oncol. 11, 654684 (2021)