BD, 562697, BD Phosflow™ PE-CF594 Mouse Anti-Smad2 (pS465/pS467)/Smad3 (pS423/pS425)

Alternative Name: SMAD2, SMAD3; MADH2, MADH3; MAD homolog 2, MAD homolog 3
Reactivity: Human (QC Testing), Mouse (Tested in Development)
Isotype: Mouse IgG1, κ
Immunogen: Phosphorylated Human Smad2 Peptide
Application: Intracellular staining (flow cytometry) (Routinely Tested)
Vol. Per Test: 5 µl
RRID: AB_2737728
Storage Buffer: Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
Regulatory Status: RUO
Preparation And Storage: Preparation And Storage Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated with BD Horizon™ PE-CF594 under optimum conditions, and unconjugated antibody and free PE-CF594 were removed.
Recommended Assay Procedures: Recommended Assay Procedures This fluorescent antibody is suitable for intracellular staining of human lymphoid cell lines, peripheral blood mononuclear cells, and mouse splenocytes using BD Cytofix™ Fixation Buffer or BD Phosflow™ Lyse/Fix Buffer with BD Phosflow™ Perm Buffer III (see table, below). Prior to stimulation, cells were serum starved overnight at a density of 2-10X10^6 cells/mL in flat-bottom 96- or 6-well tissue culture plates or in loosely capped, round-bottom tubes containing approximately 100 mL of cells in suspension. Note: 1. Serum starvation for 2 hours following PBMC isolation was not sufficient to reduce basal phosphorylation of Smad2 and Smad3. 2. Do not mix or agitate untreated cells until just before the cells are ready to be fixed, since agitation of serum-starved mouse or human primary leukocytes prior to fixation increased Smad2/3 phosphorylation, even in the absence of exogenous TGF-β
Product Notices: Product Notices This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test). Please observe the following precautions: Absorption of visible light can significantly alter the energy transfer occurring in any tandem fluorochrome conjugate; therefore, we recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to prevent exposure of conjugated reagents, including cells stained with those reagents, to room illumination. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors. Texas Red is a registered trademark of Molecular Probes, Inc., Eugene, OR. CF™ is a trademark of Biotium, Inc. When excited by the yellow-green (561-nm) laser, the fluorescence may be brighter than when excited by the blue (488-nm) laser. This product is provided under an Agreement between BIOTIUM and BD Biosciences. The manufacture, use, sale, offer for sale, or import of this product is subject to one or more patents or pending applications owned or licensed by Biotium, Inc. This product, and only in the amount purchased by buyer, may be used solely for buyer’s own internal research, in a manner consistent with the accompanying product literature. No other right to use, sell or otherwise transfer (a) this product, or (b) its components is hereby granted expressly, by implication or by estoppel. This product is for research use only. Diagnostic uses require a separate license from Biotium, Inc. For information on purchasing a license to this product including for purposes other than research, contact Biotium, Inc., 3159 Corporate Place, Hayward, CA 94545, Tel: (510) 265-1027. Fax: (510) 265-1352. Email: btinfo@biotium.com. Because of the broad absorption spectrum of the tandem fluorochrome, extra care must be taken when using multi-laser cytometers, which may directly excite both PE and CF™594. Source of all serum proteins is from USDA inspected abattoirs located in the United States. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.