The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 - 5 µg/ml. Predicted molecular weight: 43 kDa.
Use a concentration of 5 µg/ml.
Use 1µg for 106 cells. ab170192-Mouse monoclonal IgG2b, is suitable for use as an isotype control with this antibody.
Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
FunctionMay play a role in the immune system processes. Protects cells from FAS-, TNF alpha- and FADD-induced apoptosis without increasing expression of the inhibitors of apoptosis BCL2 and BCLXL. Seems to activate an inhibitory pathway that prevents CASP8 activation following FAS stimulation, rather than blocking apoptotic signals downstream. May inhibit FAS-induced apoptosis by preventing CASP8 processing through CFLAR up-regulation.
Tissue specificityExpressed in lymph nodes, peripheral blood leukocytes, lung, thymus and kidneys. Very weak expression detected in spleen, liver, heart, and salivary gland. Expressed in lymphoid cell lines such as Jurkat cells, CemT4 cells, MolT-4 cells, HB11.19 cells, and Reh cells. No expression detected in nonhematopoietic cell lines, including HepG2 cells, 293 cells and Hela cells.
ICC/IF image of ab56487 stained SHSY5Y cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab56487, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
IHC image of ab56487 staining in human lymphoma formalin fixed paraffin embedded tissue section, performed on a Leica BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab56487, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
Flow Cytometry - Anti-FAIM3 antibody (ab56487)
Overlay histogram showing Jurkat cells stained with ab56487 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab56487, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.