United Kingdom (UK)
Recombinant monoclonal antibody to Galectin 9. Manufactured using AbAb’s Recombinant Platform with variable regions (i.e. specificity) from the hybridoma RG9-35.
UniProt Accession Number of Target Protein: O08573
Alternative Name(s) of Target: ecalectin; Gal-9; galectin 9; Galectin-9; HUAT; lectin, galactose binding, soluble 5; LGALS9A; tumor antigen HOM-HD-21; urate transporter/channel protein.
Immunogen: This antibody was raised by immunising rats with recombinant mouse galectin-9.
Specificity: This antibody is specific for murine galectin-9, a 40 kDa S-type lectin that is expressed by various cell types, including lymphocytes, thymocytes, macrophages, dendritic cells, astrocytes, mast cells, eosinophils, fibroblasts, epithelial cells, and endothelial cells. Galectin-9 binds to β-galactosides and can serve as a ligand for TIM-3 (CD366). The protein is implicated in both innate and adaptive immune responses, specifically, induction of cytokine secretion by macrophages, bactericidal functions, promotion of dendritic cell maturation, regulatory T cell expansion, and negative regulation of Th1, Th17, NK, and cytotoxic T cells.
Application Notes: When this antibody was first generated and characterised, it was shown to bind with galectin-9 (Gal-9) but not with either galectin-1 or galectin-3. Furthermore, it was shown to be capable of blocking Tim-3-Fc binding to Gal-9, and inhibiting Gal-9-induced Th1 cell death in vitro (Fukushima et al., 2008; PMID: 18087160). This antibody has been used in FACS analyses. For instance, to describe a novel mechanism where Tim-3 binds HMGB-1 on virus-specific CD8+ Treg cells, suppressing proliferation of CD8+ T effector cells during acute adenoviral infection (Dolina et al., 2014; PMID: 24677194); and to indicate a crucial role for Tim-3 in phagocytosis of apoptotic cells and cross-presentation (Nakayama et al., 2009; PMID: 19224762). In addition, this antibody has been used in vivo to study the roles of Gal-9 in the development of experimental allergic conjunctivitis in mice (Fukushima et al., 2008; PMID: 18087160) and to demonstrate that constitutive expression of Gal-9 plays an immunosuppressive role in corneal allografts (Shimmura-Tomita et al., 2013; PMID: 23667648). This antibody was used in a cold immunoprecipitation (IP) assay to explore the binding of Gal-Nab1 and Gal-Nab2 to native murine gal-9, using a protein extract from murine thymus as the source of native Gal-9; RG9-35, was one of the antibodies used to isolate immune complexes formed between primary antibodies and proteins extracted from a mouse thymus for further analysis (Lhuillier et al., 2018; PMID: 30204750).
Antibody first published in:
Fukushima et al. Roles of galectin-9 in the development of experimental allergic conjunctivitis in mice. Int Arch Allergy Immunol. 2008;146(1):36-43. PMID:18087160
Note on publication:
Describe the original generation of this antibody and its subsequent use in in vitro and in vivo functional assays to study the roles of galectin-9 in the development of experimental allergic conjunctivitis in mice.
Immunofluorescence staining of fixed NIH3T3 cells with anti-Galectin 9 antibody RG9-35 (Ab01088) Immunofluorescence analysis of paraformaldehyde fixed RAW264.7 cells on Shi-fix™ coverslips stained with the chimeric rabbit IgG version of RG9-35 (Ab01088-23.0) at 10 µg/ml for 1h followed by Alexa Fluor® 488 secondary antibody (2 µg/ml), showing membrane staining. The nuclear stain is DAPI (blue). Panels show from left-right, top-bottom Ab01088-23.0, DAPI, merged channels and an isotype control. The isotype control was an unknown specificity antibody (Ab00178-23.0) followed by staining with Alexa Fluor® 488 secondary antibody.
Flow cytometry using the anti-Galectin 9 antibody RG9-35 (Ab01088). NIH3T3 cells were fixed using 2% PFA and stained with anti-unknown specificity antibody (Ab00178-23.0; isotype control, black line) or the rabbit IgG1 version of RG9-35 (Ab01088-23.0, blue line) at a dilution of 1:100 for 1h at RT. After washing, the bound antibody was detected using a goat anti-rabbit IgG AlexaFluor® 488 antibody at a dilution of 1:1000 and cells analyzed using a FACSCanto flow-cytometer.