The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration.
Use at an assay dependent concentration.
Use a concentration of 1 µg/ml. Detects a band of approximately 265 kDa (predicted molecular weight: 263 kDa).
1/200 - 1/600. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Fibronectins bind cell surfaces and various compounds including collagen, fibrin, heparin, DNA, and actin. Fibronectins are involved in cell adhesion, cell motility, opsonization, wound healing, and maintenance of cell shape. Involved in osteoblast compaction through the fibronectin fibrillogenesis cell-mediated matrix assembly process, essential for osteoblast mineralization. Participates in the regulation of type I collagen deposition by osteoblasts. Anastellin binds fibronectin and induces fibril formation. This fibronectin polymer, named superfibronectin, exhibits enhanced adhesive properties. Both anastellin and superfibronectin inhibit tumor growth, angiogenesis and metastasis. Anastellin activates p38 MAPK and inhibits lysophospholipid signaling.
Plasma FN (soluble dimeric form) is secreted by hepatocytes. Cellular FN (dimeric or cross-linked multimeric forms), made by fibroblasts, epithelial and other cell types, is deposited as fibrils in the extracellular matrix. Ugl-Y1, Ugl-Y2 and Ugl-Y3 are found in urine.
Ugl-Y1, Ugl-Y2 and Ugl-Y3 are present in the urine from 0 to 17 years of age.
Sulfated. It is not known whether both or only one of Thr-2064 and Thr-2065 are/is glycosylated. Forms covalent cross-links mediated by a transglutaminase, such as F13A or TGM2, between a glutamine and the epsilon-amino group of a lysine residue, forming homopolymers and heteropolymers (e.g. fibrinogen-fibronectin, collagen-fibronectin heteropolymers). Phosphorylated by FAM20C in the extracellular medium. Proteolytic processing produces the C-terminal NC1 peptide, anastellin.
Immunocytochemistry/ Immunofluorescence - Anti-Fibronectin antibody (ab23750)Shen H. et al PLoS Genet. 2016 Aug 19;12(8):e1006244. doi: 10.1371/journal.pgen.1006244. eCollection 2016 Aug.
Immunofluorescence assay was performed on paired NFs (normal fibroblasts; left) and CAFs (cancer-associated fibroblasts; right) from sample ID 1 using anti-fibronectin (red).
We isolated CAFs from 15 human lung carcinomas and their corresponding counterpart NFs from their matched non-malignant adjacent tissues, taken at least 10 cm from the outer tumor margin. NFs and CAFs were maintained in 1:1 mixture of DMEM and F12 medium supplemented with 400 ng/ml hydrocortisone, 200 ng/ml insulin, 15% FBS and 1% penicilin/streptomycin. Cells were maintained in 5% CO2 incubator at 37°C.
For immunofluorescence assay, samples were stained with fibronectin after blocking with bovine serum albumin. Samples were incubated with Alexa Fluor 568-conjugated secondary antibody (red). Microscopic observation was performed under a fluorescence microscope.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Fibronectin antibody (ab23750)This image is courtesy of an anonymous abreview.
ab23750 staining Fibronectin in Rat skin tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with Immunoblock® (5 µg/ml) for 30 minutes at 25°C; antigen retrieval was by heat mediation in a citrate buffer. Samples were incubated with primary antibody (1/500 in diluent) for 1 hour 30 minutes at 25°C. An undiluted Goat anti-rabbit HRP/DAB polydetector polyclonal was used as the secondary antibody.
Immunohistochemistry (Frozen sections) - Anti-Fibronectin antibody (ab23750)This image is courtesy of an anonymous abreview.
ab23750 staining fibronectin in mouse liver tissue by immunohistochemistry (frozen sections). Cells were formaldehyde fixed and permeabilized in 0.2% Triton X-100 prior to blocking in 2% BSA for 30 minutes at 20°C. The primary antibody was diluted 1/200 and incubated with the sample for 9 hours at 4°C. Alexa fluor® 488 goat polyclonal to rabbit Ig, diluted 1/200, was used as the secondary antibody.
Immunocytochemistry/ Immunofluorescence - Anti-Fibronectin antibody (ab23750)This image is courtesy of an anonymous abreview.
ab23750 staining Fibronectin in mouse bone marrow cells by Immunocytochemistry/ Immunofluorescence. Cells were formaldehyde fixed, blocked in 2% BSA for 45 minutes at 20°C. The primary antibody was diluted 1/200 and incubated with sample for 9 hour at 4°C. An Alexa Fluor® 488 conjugated goat polyclonal to rabbit IgG, was used undiluted as secondary. The nuclei were stained with DAPI.
ab23750 (2ug/ml) staining Fibronectin in human aorta using an automated system (DAKO Autostainer Plus). Using this protocol there is strong staining Fibronectin in the cytoplasm and extracellularly in the connective tissue. Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer EDTA pH 9.0 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.
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Vermeulen M et al. Development of a Cytocompatible Scaffold from Pig Immature Testicular Tissue Allowing Human Sertoli Cell Attachment, Proliferation and Functionality. Int J Mol Sci19:N/A (2018).
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