Flow Cytometry Product Notes: Test Sizes: To accommodate multicolor flow cytometry, eBioscience is in the process of reducing test size volumes from 20 µl to 5 µl. Please check your antibody vial for the recommended test size. Fluorochrome Replacements: eBioscience is in the process of replacing all Alexa Fluor® 647 conjugated products with eFluor® 660 conjugated products.
Description
This 5C4 monoclonal antibody reacts with human Nef-associated factor-1 (Naf-1). This protein is also designated as A20-binding inhibitor of NFkB activation (ABIN-1) and TNFAIP3-interacting protein 1 (TNIP1). Naf-1 is predicted to consist of four leucine zippers and four coiled-coil domains. This protein exists as two isoforms, Naf1α and Naf1β, which are 72-kDa and 68-kDa, respectively, and is expressed in a variety of tissues, with greatest expression in peripheral blood lymphocytes, spleen, and skeletal muscle. Naf1 has also been detected in the Jurkat, Molt-4, and HL60 cell lines.
Naf-1 has been shown to play several biological roles. Naf-1 was originally identified associated with the Nef protein of HIV, and was found to enhance CD4 cell surface expression. Naf-1 has also been demonstrated to inhibit TNF-α- and IL-1-mediated NFkB-dependent gene expression by competing with IKK-γ. Finally, this adaptor protein has been found to associate constitutively with P-selectin glycoprotein ligand 1 (PSGL-1) in human leukocytes. Upon P-selectin binding to PSGL-1, Naf1 is phosphorylated at Tyr552 by Src family kinases, leading to PI3K recruitment and integrin activation.
This antibody recognizes both the unphosphorylated and phosphorylated forms of Naf1α and Naf1β.
Applications Reported
For research use only, not for diagnostic or therapeutic use. This 5C4 antibody has been reported for use in immunoprecipitation, immunoblotting (WB), and immunohistology staining of paraffin embedded tissue sections.
Applications Tested
The 5C4 antibody has been tested by western blot on lysates prepared from Jurkat cells. This antibody can be used at 1-5 ug/mL. It is recommended that the antibody be carefully titrated for optimal performace in the assay of interest.
References
Wang HB, Wang JT, Zhang L, Geng ZH, Xu WL, Xu T, Huo Y, Zhu X, Plow EF, Chen M, Geng JG. P-selectin primes leukocyte integrin activation during inflammation.
Nat Immunol. 2007 Aug;8(8):882-92. (5C4, WB, Pubmed )
Chang MO, Yamamoto N, Horiuchi S, Wu YF, Fujimoto M, Yamamoto N. Production and characterization of a monoclonal antibody specific to Nef-associated factor 1 (Naf1)/A20-binding inhibitor of NF-kappaB activation (ABIN-1). Hybridoma (Larchmt). 2005 Oct;24(5):248-57.
Fukushi M, Dixon J, Kimura T, Tsurutani N, Dixon MJ, Yamamoto N. Identification and cloning of a novel cellular protein Naf1, Nef-associated factor 1, that increases cell surface CD4 expression. FEBS Lett. 1999 Jan 8;442(1):83-8.
