Anti-Src antibody [GD11] - BSA and Azide free

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Src antibody [GD11] - BSA and Azide free (AB269563)

ab231081 staining Src in wild-type HAP1 cells (top panel) and SRC knockout HAP1 cells (bottom panel). The cells were fixed with 4% formaldehyde (10min), 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 ab231081 at 5μg/ml and ab6046 (Rabbit polyclonal to beta tubulin - loading control) at 1/1000 dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with ab150117 (Goat secondary antibody to Mouse IgG (Alexa Fluor^® 488) at 2 μg/ml (colored green) and ab150084 (Goat secondary antibody to Rabbit IgG (Alexa Fluor^® 594) at 2 μg/ml (pseudo color red). Nuclear DNA was labelled in blue with DAPI.

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

This image was produced using the same antibody clone but in a different formulation ab231081, PBS and sodium azide.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Src antibody [GD11] - BSA and Azide free (AB269563)

IHC image of Src staining in a section of formalin-fixed paraffin-embedded human normal testis* performed on a Leica BOND^TM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20mins. The section was then incubated with ab231081, 1ug/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

This image was produced using the same antibody clone but in a different formulation ab231081, PBS and sodium azide.

• WB

Lab

Western blot - Anti-Src antibody [GD11] - BSA and Azide free (AB269563)

ab231081 was shown to specifically react with Src in wild type HAP1 cells. No band was observed when Src knockout samples were used. Wild-type and Src knockout samples were subjected to SDS-PAGE. ab231081 and ab181602 (Rabbit anti-GAPDH loading control) were incubated overnight at 4°C at a 1ug/ml concentration and 1/10000 dilution respectively. 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 1/20000 dilution for 1 hour at room temperature before imaging.

This image was produced using the same antibody clone but in a different formulation ab231081, PBS and sodium azide.

All lanes:

Western blot - Anti-Src antibody [GD11] (<a href='/en-us/products/primary-antibodies/src-antibody-gd11-ab231081'>ab231081</a>)

Lane 1:

HAP1 whole cell lysate at 20 µg

Lane 2:

HAP1 Src KO whole cell lysate at 20 µg

Lane 3:

A431 whole cell lysate at 20 µg

Lane 4:

Mouse testes whole tissue lysate at 20 µg

Lane 5:

Rat testes whole tissue lysate at 20 µg

Predicted band size: 60 kDa

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