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Mixture of affinity purified tyrosine phosphoproteins from chick embryo fibroblasts expressing activated pp60 src.
Our Abpromise guarantee covers the use of ab33333 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 a concentration of 1 µg/ml.|
|WB||Use a concentration of 0.5 - 2 µg/ml. Predicted molecular weight: 61 kDa. Use 20 µg per lane of 3T3 cell lysate as a positive control for mini gels.
Note: ab33333 recognizes 80kD and 85kD proteins in avian cells, but it recognizes an 80kD protein in rodent and human cells.
|IP||Use at 10 µg/mg of lysate.|
|ICC||Use a concentration of 10 µg/ml.|
Samples were incubated with primary antibody for 2 hours at 20ºC.
Blocking: 5% milk for 1 hour at 20ºC.
ab33333 staining Cortactin in wild-type HAP1 cells (top panel) and CTTN knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab33333 at 1μg/ml and ab7291 at 1μg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and goat secondary antibody to Mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in pseudo colour red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
Blocking was with 5% milk for 1 hour at 20°C.
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