Anti-alpha smooth muscle Actin antibody [EPR5368] – Chicken IgY (Chimeric) – BSA and Azide Free

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-alpha smooth muscle Actin antibody [EPR5368] – Chicken IgY (Chimeric) – BSA and Azide Free (AB322917)

ab321866 staining ACTA2 in HeLa (positive) and A431 (negative) cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-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 overnight at 4°C with ab321866 at 1µg/ml (shown in green) and ab202272, Alexa Fluor® 594 Rabbit monoclonal [EP1332Y] to alpha Tubulin - Microtubule Marker (shown in pseudocolour magenta). Cells were then incubated with ab150173, Goat Anti-Chicken IgY H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution. Nuclear DNA was labelled with DAPI (shown in blue).

Also suitable in cells fixed with 4% paraformaldehyde (10 min).

Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

This data was developed using the same antibody clone in a different buffer formulation.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-alpha smooth muscle Actin antibody [EPR5368] – Chicken IgY (Chimeric) – BSA and Azide Free (AB322917)

Immunofluorescence staining of ACTA2 in sections of formalin-fixed paraffin-embedded human colon (positive) and human cerebellum (negative)*. Performed on a Leica BOND. The section was pre-treated using heat mediated antigen retrieval with sodium citrate (pH6.0) for 20 minutes. The section was then incubated at room temperature for 1 hour withab321866 at 1/500 dilution and then incubated for 1 hour with ab150173 Goat Anti-Chicken IgY H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green). Nuclear DNA was labelled with DAPI (shown in blue). The section was then mounted using Dako Fluorescence Mounting Medium®. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). For other IHC staining systems (automated and non-automated), customers should optimize variable parameters such as antigen retrieval conditions, antibody concentrations and incubation times. This data was developed using the same antibody clone in a different buffer formulation. *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-alpha smooth muscle Actin antibody [EPR5368] – Chicken IgY (Chimeric) – BSA and Azide Free (AB322917)

Immunofluorescence staining of ACTA2 in sections of formalin-fixed paraffin-embedded mouse stomach (positive) and mouse brain (negative)*. Performed on a Leica BOND. The section was pre-treated using heat mediated antigen retrieval with sodium citrate (pH6.0) for 20 minutes. The section was then incubated at room temperature for 1 hour with ab321866 at 1/500 dilution and then incubated for 1 hour with ab150173 Goat Anti-Chicken IgY H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (shown in green). Nuclear DNA was labelled with DAPI (shown in blue). The section was then mounted using Dako Fluorescence Mounting Medium®. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). This data was developed using the same antibody clone in a different buffer formulation. For other IHC staining systems (automated and non-automated), customers should optimize variable parameters such as antigen retrieval conditions, antibody concentrations and incubation times.