The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration.
1/1000 - 1/10000. Detects a band of approximately 195 kDa (predicted molecular weight: 189 kDa).
1/500 - 1/1000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Application notesIs unsuitable for Flow Cyt or IP.
FunctionBinds to activated CDC42 but does not stimulate its GTPase activity. It associates with calmodulin. Could serve as an assembly scaffold for the organization of a multimolecular complex that would interface incoming signals to the reorganization of the actin cytoskeleton at the plasma membrane. May promote neurite outgrowth.
Tissue specificityExpressed in the placenta, lung, and kidney. A lower level expression is seen in the heart, liver, skeletal muscle and pancreas.
DomainRegions C1 and C2 can either interact with nucleotide-free CDC42, or interact together, depending on the phosphorylation state of Ser-1443. When Ser-1443 is not phosphorylated, C1 and C2 interact, which prevents binding of nucleotide-free CDC42 and promotes binding of GTP-bound CDC42. Phosphorylation of Ser-1443 prevents interaction between C1 and C2, which opens the structure of the C-terminus and allows binding and sequestration of nucleotide-free CDC42 on both C1 and C2.
Post-translational modificationsPhosphorylation of Ser-1443 by PKC prevents interaction between C1 and C2, allowing binding of nucleotide-free CDC42. Ser-1443 phosphorylation enhances the ability to promote neurite outgrowth.
Immunocytochemistry/Immunofluorescence analysis of HeLa (Human epithelial cell line from cervix adenocarcinoma) labeling IQGAP1 with Purified ab109292 at 1/500 dilution (5 µg/ml). Cells were fixed with 100% methanol. ab150077 Goat anti rabbit IgG(Alexa Fluor® 488) at 1/1000 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI. PBS was used instead of the primary antibody as the negative control.
Western blot - Anti-IQGAP1 antibody [EPR5221] (ab109292)
Predicted band size : 189 kDa
Lane 1: Wild-type HAP1 cell lysate (20 µg) Lane 2: IQGAP1 knockout HAP1 cell lysate (20 µg) Lane 3: HEK293 cell lysate (20 µg) Lane 4: HeLa cell lysate (20 µg) Lanes 1 - 4: Merged signal (red and green). Green - ab109292 observed at 190 kDa. Red - loading control, ab7291, observed at 52 kDa. ab109292 was shown to specifically react with IQGAP1 when IQGAP1 knockout samples were used. Wild-type and IQGAP1 knockout samples were subjected to SDS-PAGE. ab109292 and ab7291 (loading control to alpha tubulin) were diluted 1/1000 and 1/2000 and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.
Western blot - IQGAP1 antibody [EPR5221] (ab109292)
All lanes : Anti-IQGAP1 antibody [EPR5221] (ab109292) at 1/1000 dilution
Lane 1 : HeLa cell lysate Lane 2 : Cos-1 cell lysate Lane 3 : HUVEC cell lysate Lane 4 : Jurkat cell lysate