Anti-Vimentin antibody [EPR3776] - Cytoskeleton Marker – Chicken IgY (Chimeric)

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [EPR3776] - Cytoskeleton Marker – Chicken IgY (Chimeric) (AB323238)

ab14734 staining VDAC1 in HEK293 (positive) and HEK293-VDAC1 KO (negative) cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Tween 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 for 2 hours at room temperature with ab14734 at 1µg/ml (shown in green) and ab186735, anti-TOMM20 antibody [EPR15581-54] - Mitochondrial Marker (shown in magenta) and ab323238, anti-Vimentin antibody [EPR3776] - Cytoskeleton Marker – Chicken IgY Chimeric (shown in yellow) . Cells were then incubated with ab150117, Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution; ab150084, Goat Anti-Rabbit IgG H&L (Alexa Fluor® 594) preadsorbed at 1/1000 dilution; and ab150175, Goat Anti-Chicken IgY H&L (Alexa Fluor® 647) preadsorbed, at 1/1000 dilution. Nuclear DNA was labelled with DAPI (shown in blue). Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [EPR3776] - Cytoskeleton Marker – Chicken IgY (Chimeric) (AB323238)

Immunofluorescence staining of VIM in sections of formalin-fixed paraffin-embedded human kidney (positive) and human skeletal muscle (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 ab323238 at 1/500 dilution and then incubated for 1 hour with ab150173 Goat Anti-Chicken IgG 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. *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Vimentin antibody [EPR3776] - Cytoskeleton Marker – Chicken IgY (Chimeric) (AB323238)

ab323238 staining VIM in Hap1 (positive) and Hap1-VIM KO (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 ab323238 at 1ug/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.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Vimentin antibody [EPR3776] - Cytoskeleton Marker – Chicken IgY (Chimeric) (AB323238)

Immunofluorescence staining of VIM in sections of formalin-fixed paraffin-embedded mouse ovary (positive) and mouse skeletal muscle (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 ab323238 at 1/500 dilution and then incubated for 1 hour with ab150173 Goat Anti-Chicken IgG 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.