Immunofluorescence analysis of FAK [pY397] was done on 80% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 15 minutes; permeabilized with 0.25% Triton™ X-100 for 10 minutes followed by blocking with 5% BSA for 1 hour at room temperature. The cells were incubated with ABfinity™ FAK [pY397] Recombinant Rabbit Monoclonal Antibody (700255) at 1µgmL in 1% BSA and incubated for 3 hours at room temperature and then labeled with Alexa Flour® 488 Goat Anti-Rabbit IgG Secondary Antibody (A11008) at a dilution of 1:400 for 30 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (S36938). F-actin (Panel c: red) was stained with Alexa Fluor 594 Phalloidin (A12381). Panel d is a merged image showing cytoplasmic of FAK [pY397]. Panel e shows no primary antibody. The images were captured at 20X magnification.
|Tested species reactivity||Human, Mouse|
|Published species reactivity||Rat, Mouse, Human, Not Applicable|
|Host / Isotype||Rabbit / IgG|
|Immunogen||A peptide corresponding to amino acids 390-401 of Q05397.|
|Contains||0.09% sodium azide|
|Storage Conditions||Maintain refrigerated at 2-8°C for up to 1 month. For long term storage store at -20°C|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||1-3 µg/10^6 cells|
|Immunofluorescence (IF)||1-2 ug/ml|
|Immunohistochemistry (Paraffin) (IHC (P))||1:10-1:1000|
|Western Blot (WB)||1:500-1:2000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
This antibody is predicted to react with mouse, rat, primate, bovine, equine, canine, chicken, opossum, zebra finch, zebrafish, Xenopus and pufferfish based on sequence homology.
ABfinity™ recombinant antibodies are rabbit monoclonal antibodies, unmatched for producing superior results. ABfinity™ antibodies are developed by immunizing animals, screening for functionality, cloning the immunogen-specific antibody genes into high-level mammalian expression vectors, produced on a large scale, and purified with Protein A.
ABfinity™ monoclonal antibodies resemble rabbit monoclonals isolated from serum or produced by hybridomas, but demonstrate greater specificity and sensitivity. Because ABfinity™ recombinant antibodies are derived from cloned DNA sequences of the heavy and light antibody chains, they are not susceptible to cell-line drift or lot-to-lot variation, thus allowing for peak specificity and performance.
Intact IgG appears on a non-reducing gel as ~150 kDa band and upon reduction generating a ~25 kDa light chain band and a ~50 kDa heavy chain.
Focal Adhesion Kinase (FAK) is a 125 kDa non-receptor protein tyrosine kinase that acts as a substrate for Src and is a key element of integrin signaling. FAK plays a central role in cell spreading, differentiation, migration, cell death and acceleration of the G1 to S phase transition of the cell cycle. Regulation of FAK includes phosphorylation at multiple tyrosine and serine residues. Increased FAK tyrosine phosphorylation occurs upon integrin engagement with fibronectin.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
|Not Applicable||Not Cited||
HSP70 Inhibition Limits FAK-Dependent Invasion and Enhances the Response to Melanoma Treatment with BRAF Inhibitors.
700255 was used in immunohistochemistry - paraffin section to determine limits on FAK-dependent invasion and enhancement of the response to melanoma treatment with BRAF inhibitors due to HSP70 inhibition
|Budina-Kolomets A,Webster MR,Leu JI,Jennis M,Krepler C,Guerrini A,Kossenkov AV,Xu W,Karakousis G,Schuchter L,Amaravadi RK,Wu H,Yin X,Liu Q,Lu Y,Mills GB,Xu X,George DL,Weeraratna AT,Murphy ME||Cancer research (76:2720)||2016|
Focal adhesion kinase overexpression and its impact on human osteosarcoma.
700255 was used in immunohistochemistry - paraffin section and western blot to determine the impact on human osteosarcoma due to focal adhesion kinase overexpression
|Ren K,Lu X,Yao N,Chen Y,Yang A,Chen H,Zhang J,Wu S,Shi X,Wang C,Sun X||Oncotarget (6:31085)||2015|
Mechanical Loading Stimulates Expression of Collagen Cross-Linking Associated Enzymes in Periodontal Ligament.
700255 was used in immunohistochemistry (paraffin) to test the effects of mechanical loading on the expression of collagen-modifying enzymes and subsequent tissue changes in periodontal ligament.
|Kaku M,Rosales Rocabado JM,Kitami M,Ida T,Akiba Y,Yamauchi M,Uoshima K||Journal of cellular physiology (231:926)||2016|
Spatiotemporal dynamics of traction forces show three contraction centers in migratory neurons.
700255 was used in immunocytochemistry to study how traction force is generated and regulated during neuronal migration.
|Jiang J,Zhang ZH,Yuan XB,Poo MM||The Journal of cell biology (209:759)||2015|
||Loading-related regulation of transcription factor EGR2/Krox-20 in bone cells is ERK1/2 protein-mediated and prostaglandin, Wnt signaling pathway-, and insulin-like growth factor-I axis-dependent.||Zaman G,Sunters A,Galea GL,Javaheri B,Saxon LK,Moustafa A,Armstrong VJ,Price JS,Lanyon LE||The Journal of biological chemistry (287:3946)||2012|
Matrix stiffening and ß1 integrin drive subtype-specific fibroblast accumulation in lung cancer.
700255 was used in western blot to elucidate the mechanical and soluble mitogenic cues that promote the accumulation of tumor-associated fibroblasts in non-small cell lung cancer subtypes.
|Puig M,Lugo R,Gabasa M,Giménez A,Velásquez A,Galgoczy R,Ramírez J,Gómez-Caro A,Busnadiego Ó,Rodríguez-Pascual F,Gascón P,Reguart N,Alcaraz J||Molecular cancer research : MCR (13:161)||2015|
|Mouse||Not Cited||Loading-related regulation of transcription factor EGR2/Krox-20 in bone cells is ERK1/2 protein-mediated and prostaglandin, Wnt signaling pathway-, and insulin-like growth factor-I axis-dependent.||Zaman G,Sunters A,Galea GL,Javaheri B,Saxon LK,Moustafa A,Armstrong VJ,Price JS,Lanyon LE||The Journal of biological chemistry (287:3946)||2012|