Detection of Human CTLA‑4 by Western Blot.
Western blot shows lysates of NS0 mouse myeloma cell line either mock transfected or transfected with human CTLA-4. PVDF membrane was probed with 0.5 µg/mL of Goat Anti-Human CTLA‑4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-386-PB) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). A specific band was detected for CTLA‑4 at approximately 50 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
Detection of CTLA‑4 in NS0 Mouse Cell Line Co-transfected with CTLA-4 and eGFP by Flow Cytometry.
NS0 mouse myeloma cell line co-transfected with human CTLA-4 and eGFP was stained with either (A) Goat Anti-Human CTLA‑4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-386-PB) or (B) Normal Goat IgG Control (Catalog # AB-108-C) followed by Allophycocyanin-conjugated Anti-Goat IgG Secondary Antibody (Catalog # F0108).
CTLA‑4 in Human Peripheral Blood Mononuclear Cells.
CTLA‑4 was detected in immersion fixed human peripheral blood mononuclear cells treated with treated with PMA and calcium ionomycin using Goat Anti-Human CTLA‑4 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-386-PB) at 15 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Goat IgG Secondary Antibody (red; Catalog # NL001) and counterstained with DAPI (blue). Specific staining was localized to cell surfaces. View our protocol for Fluorescent ICC Staining of Non-adherent Cells.
Preparation and Storage
Reconstitute at 0.2 mg/mL in sterile PBS.
Reconstitution Buffer Available
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
Stability & Storage
Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
12 months from date of receipt, -20 to -70 °C as supplied.
1 month, 2 to 8 °C under sterile conditions after reconstitution.
6 months, -20 to -70 °C under sterile conditions after reconstitution.
CTLA-4 and CD28, together with their ligands B7-1 and B7-2, constitute one of the dominant costimulatory pathways that regulate T- and B-cell responses. CTLA-4 and CD28 are structurally homologous molecules that are members of the immunoglobulin (Ig) gene superfamily. Both CTLA-4 and CD28 are composed of a single Ig V‑like extracellular domain, a transmembrane domain and an intracellular domain. CTLA-4 and CD28 are both expressed on the cell surface as disulfide-linked homodimers or as monomers. The genes encoding these two molecules are closely linked on human chromosome 2. CTLA-4 was originally identified as a gene that was specifically expressed by cytotoxic T lymphocytes. However, CTLA-4 transcripts have since been found in both Th1 and Th2, and CD4+ and CD8+ T cell clones. Whereas CD28 expression is constitutive on the surfaces of 95% of CD4+ T cells and 50% of CD8+ T cells and is down regulated upon T cell activation, CTLA-4 expression is upregulated rapidly following T cell activation and peaks approximately 24 hours following activation. Although both CTLA-4 and CD28 can bind to the same ligands, CTLA-4 binds to B7-1 and B7-2 with 20‑100‑fold higher affinity than CD28. The physiological role of CTLA-4 in T cell costimulation is currently being studied.
Lenschow, D.J. et al. (1996) Annu. Rev. Immunol. 14:233.
Hathcock, K.S. and R.J. Hodes (1996) Advances in Immunol. 62:131.
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