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Our Abpromise guarantee covers the use of ab24525 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ICC/IF||Use at an assay dependent concentration.|
|IHC-Fr||Use at an assay dependent concentration. PubMed: 22919071|
Immunohistochemistry (Frozen sections) analysis of mouse heart tissue sections labelling Vimentin with ab24525. Tissue was fixed in 4% paraformaldehyde in PBS at 4°C overnight, and cyroprotected with 30% sucrose in PBS. The samples were embedded in optimal cutting temperature (OCT) compound, snap-frozen in liquid nitrogen, and stored at −80°C. Paraformaldehyde-fixed cryosections (10 μm) were labeled with the primary antibodies Anti-Vimentin (ab24525) and Anti-GFP (ab6662) and secondary Alexa Fluor-conjugated antibodies. Nuclei were labeled with 0.2 μg/ml DAPI.
Immunocytochemistry/ Immunofluorescence analysis of neuron/glial cultures labeling Vimentin with ab24525 (green) and GFAP with ab7260 (red). Vimentin is the sole cytoplasmic intermediate filament subunit expressed in fibroblasts, microglial and endothelial cells. The flattened cells in the middle of the image which appear green are fibroblasts. Astrocytes may express primarily GFAP, or both GFAP and vimentin, and so appear red (GFAP only) or golden yellow (GFAP and Vimentin). In cells which express both GFAP and vimentin, the two proteins assemble to produce heteropolymer filaments.
Rat cerebral cortex cultures stained with chicken antibody to vimentin ab24525 (green) and rabbit antibody to GFAP (red). Note flattened fibroblastic cells are mostly green (i.e. vimentin positive, GFAP negative), while clearly astrocytic cells, express both vimentin and GFAP and therefore appear golden or orange. Certain other cells express predominantly GFAP and therefore appear red.
Immunohistochemistry (Frozen sections) analysis of mouse dorsal skin wound labelling Vimentin with ab24525 (red) at 1/200 dilution. Dissected wounds were fixed for 2 hrs in 4% PFA and infused with 10% sucrose before cryo-embedding. For cryo-mounting, sucrose-infused wounds were mounted in optimal cutting medium (O.C.T) and frozen in a bath of dry ice and ethanol. 7 µm sections were subsequently cut on a cryo-stat for immuno-histochemistry studies. Cryo-sections were permeabilized in 0.5% Triton-x-100 before blocking with 10% normal goat serum. DAPI nuclear staining.
Western blot of Rat whole brain extract,HeLa,SH-SY5Y,HEK293, and NIH/3T3 cells probed with ab24525,showing a single strong band at ~ 50kDa.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat brain tissue sections labeling Vimentin with ab24525 (undiluted). Cells were fixed with paraformaldehyde and permeabilized with 0.3% Triton X-100. Cells were blocked with 5% normal donkey serum for 1 hour at 22°C, followed by incubation with ab24525 in 5% donkey serum 0.3%tritonx-100 in kpbs for 18 hours at 4°C. A polyclonal donkey anti-chicken AMCA conjugated secondary antibody was used at 1/500 dilution.
Immunohistochemical analysis of murine eye tissue, staining Vimentin (red) with ab24525.
Tissue was fixed in paraformaldehyde, blocked for 1 h at room temperature in 0.5% BSA, 0.2% Tween-20 plus 5% goat serum. Samples were incubated with primary antibody (1/200) for 1 hour at room temperature. A Cy3-conjugated goat anti-chicken IgG (1/200) was used as the secondary antibody.
ab24525 staining Vimentin in rat smooth muscle cells from mesenteric artery by Immunocytochemistry/ Immunofluorescence. Cells were fixed with 4% paraformaldehyde in physiological saline solution (PSS) 4 min at 4°C and permeabilized with 0.3% Triton x100 before blocking with 2% BSA was done for 30 minutes at 20°C. Samples were incubated with primary antibody (1/300: in PSS with 2%BSA and 0.3% Triton X-100) for 14 hours at 4°C. An Abcam`s ab6875, goat anti-chicken IgY Texas Red was used as secondary antibody at 1/400 dilution.