Immunofluorescent analysis of GTPase Activating Protein (green) showing positive staining in the cytoplasm of C2C12 cells (right) compared with a negative control in the absence of primary antibody (left). Formalin-fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 5-10 minutes, blocked with 3% BSA-PBS for 30 minutes at room temperature and probed with a GTPase Activating Protein monoclonal antibody (Product # MA4-001) in 3% BSA-PBS at a dilution of 1:200 and incubated overnight at 4 °C in a humidified chamber. Cells were washed with PBST and incubated with a DyLight 488-conjugated goat-anti-mouse IgG (H+L) secondary antibody in PBS at room temperature in the dark. F-actin (red) was stained with a flourescent red phalloidin and nuclei (blue) were stained with DAPI for 5-10 minutes in the dark. Images were taken at a magnification of 60x.
|Tested species reactivity||Bovine, Human, Mouse, Non-human primate, Rat|
|Published species reactivity||Not Applicable|
|Host / Isotype||Mouse / IgG2a|
|Immunogen||Full length recombinant human GAP.|
|Contains||0.05% sodium azide|
|Storage Conditions||-20° C, Avoid Freeze/Thaw Cycles|
|Tested Applications||Dilution *|
|Immunocytochemistry (ICC)||5 µg/ml|
|Immunofluorescence (IF)||5 µg/ml|
|Immunoprecipitation (IP)||1-5 µg|
|Western Blot (WB)||10 µg/ml|
* 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.
|Miscellaneous PubMed (MISC)||See 1 publications below|
MA4-001 detects GTPase activating protein (GAP) from human, mouse, rat, bovine and non-human primate tissues.
MA4-001 has been successfully used in Western blot, immunofluorescence and immunoprecipitation procedures. By Western blot, this antibody recognizes a single 120 kDa protein representing ras GAP from RS-2 cell lysate. Immunofluorescence staining of GAP in mouse fibroblast cells with MA4-001 results in diffuse cytoplasmic staining. Following the addition of PDGF, immunofluorescence staining shows that some GAP rapidly translocates to the plasma membrane. MA4-001 immunoprecipitates GAP that is complexed with the GAP related p190.
The MA4-001 antigen is full length recombinant human GAP. Epitope mapping studies suggest that this antibody binds a portion of GAP that contains the src homology regions SH2 and SH3.
Reconstitute with PBS.
GTPase Activating Protein (GAP) is capable of stimulating the hydrolysis of GTP by ras p21 proteins, though GAP has little effect on the oncogenic forms of ras. It is also known that several tyrosine kinases such as platelet derived growth factor receptor and epidermal growth factor receptor are involved in the tyrosine phosphorylation of GAP. It has therefore been suggested that GAP may provide a connection or link between growth factor receptors and the ras p21 family.
IP-MS enrichment of RASA1 (LFQ intensity): RASA1 was enriched 137-fold from BT549 lysate compared to background proteins, using the optimized IP-MS workflow with Pierce MS-Compatible Magnetic IP Kit protein A/G (Part No. 90409) and RASA1 antibody (Part No. MA4-001). The STRING database (www.string-db.org) was used to identify the protein interactor list. See more information on IP-MS verification of antibody selectivity. IP-MS validation info.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
|Not Applicable||Not Cited||
Molecular cloning of cDNAs encoding the GAP-associated protein p190: implications for a signaling pathway from ras to the nucleus.
MA4-001 was used in immunoprecipitation and western blot to clone the GAP-associated protein p190 and to investigate its functional properties
|Settleman J,Narasimhan V,Foster LC,Weinberg RA||Cell (69:539)||1992|