Anti-Cytokeratin 5 抗体 [EP1601Y] - Cytoskeleton Marker (ab52635)

IHC image of Cytokeratin 5 staining in a section of formalin-fixed paraffin-embedded normal human skin* performed on a Leica Biosystems BOND® RX instrument. 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 ab52635, 0.1ug/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

Blocking buffer: 5% NFDM /TBST.

The lysates were prepared in 1%SDS Hot lysis method.

Observed MW: 62kDa

Blocking/diluting buffer and concentration: 5% NFDM/TBST

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human cervical carcinoma tissue sections labeling Cytokeratin 5 with Purified ab52635 at 1:200 dilution. Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used. PBS instead of the primary antibody was used as the negative control.

Immunocytochemistry/ Immunofluorescence analysis of A431 (Human epidermoid carcinoma epithelial cell) cells labeling Cytokeratin 6 with Purified ab52635 at 1/100 dilution. Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1/200 (2.5 μg/ml). ab150077 Goat anti rabbit IgG(Alexa Fluor^® 488) was used as the secondary antibody at 1/1000 dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

Blocking and diluting buffer: 5% NFDM/TBST.

The lysates were prepared in 1%SDS Hot lysis method.

Intracellular Flow Cytometry analysis of A431 (human epidermoid carcinoma) cells labelling Cytokeratin 5 with purified ab52635 at 1/20 (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. An Alexa Fluorr^®488-conjugated goat anti-rabbit IgG (1/2000) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies. 

Colocalization of KRT5, KRT6 and KRT17 in HSC3 cells

Immunocytochemistry in HSC3 (human oral squamous carcinoma cell line) cells. Scale bar, 10 μm.

(Taken from Figure S3 of Khanom et al)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat skin tissue sections labeling Cytokeratin 5 with Purified ab52635 at 1:200 dilution. Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used. PBS instead of the primary antibody was used as the negative control.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse skin tissue sections labeling Cytokeratin 5 with Purified ab52635 at 1:200 dilution. Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) secondary antibody was used. PBS instead of the primary antibody was used as the negative control.

Unpurified ab52635 showing positive staining in squamous cell cervical carcinoma tissue.

A431 cells stained with unpurified ab52635 at 1/100 - 1/250

Human transitional urinary bladder carcinoma stained with unpurified ab52635 at 1/100 - 1/250 dilution.

Unpurified ab52635 showing positive staining in basal cell breast carcinoma tissue.

Unpurified ab52635 showing negative staining in ductal breast carcinoma tissue.

Unpurified ab52635 showing negative staining in stomach adenocarcinoma tissue.

Unpurified ab52635 showing positive staining in normal tonsil squamous cells tissue.

Unpurified ab52635 showing positive staining in squamous cell lung carcinoma tissue.