Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EP1628Y] to Cytokeratin 8 - Cytoskeleton Marker
- Suitable for: WB, IP, Flow Cyt, IHC-P, ICC/IF
- Knockout validated
- Reacts with: Mouse, Human
Product nameAnti-Cytokeratin 8 antibody [EP1628Y] - Cytoskeleton Marker
See all Cytokeratin 8 primary antibodies
DescriptionRabbit monoclonal [EP1628Y] to Cytokeratin 8 - Cytoskeleton Marker
Tested applicationsSuitable for: WB, IP, Flow Cyt, IHC-P, ICC/IFmore details
Species reactivityReacts with: Mouse, Human
Synthetic peptide within Human Cytokeratin 8 aa 300-400 (C terminal). The exact sequence is proprietary.
Database link: P05787
- IHC-P: Human breast adenocarcinona, ovarian carcinoma, breast carcinoima, colon adenocarcinoma, endometrial carcinoma and thyroid carcinoma tissue; mouse liver tissue; ICC/IF: HT-29 and HeLa cells; WB: HeLa, A431 and HaCaT cell lysates; Human breast cancer lysates and Mouse colon lysate; Flow cyt: HeLa cells.
Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.
Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
Dissociation constant (KD)KD = 4.60 x 10 -10 M Learn more about KD
Storage bufferpH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
Concentration information loading...
PurityProtein A purified
- Anti-Cytokeratin 8 antibody [EP1628Y] (Alexa Fluor® 488) (ab192467)
- Anti-Cytokeratin 8 antibody [EP1628Y] (Alexa Fluor® 647) (ab192468)
- Anti-Cytokeratin 8 antibody [EP1628Y] (HRP) (ab193094)
- Anti-Cytokeratin 8 antibody [EP1628Y] (PE) (ab209297)
- Anti-Cytokeratin 8 antibody [EP1628Y] (Alexa Fluor® 405) (ab210139)
- Anti-Cytokeratin 8 antibody [EP1628Y] - BSA and Azide free (ab217173)
KO cell lines
KO cell lysates
Our Abpromise guarantee covers the use of ab53280 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||1/10000. Detects a band of approximately 52 kDa (predicted molecular weight: 54 kDa).
For unpurified use at 1/25,000 - 1/50,000.
For unpurified use at 1:70.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
|IHC-P||1/250. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.|
|ICC/IF||1/100 - 1/500.|
FunctionTogether with KRT19, helps to link the contractile apparatus to dystrophin at the costameres of striated muscle.
Tissue specificityObserved in muscle fibers accumulating in the costameres of myoplasm at the sarcolemma membrane in structures that contain dystrophin and spectrin. Expressed in gingival mucosa and hard palate of the oral cavity.
Involvement in diseaseCirrhosis
Sequence similaritiesBelongs to the intermediate filament family.
modificationsPhosphorylation on serine residues is enhanced during EGF stimulation and mitosis. Ser-74 phosphorylation plays an important role in keratin filament reorganization.
O-glycosylated. O-GlcNAcylation at multiple sites increases solubility, and decreases stability by inducing proteasomal degradation.
O-glycosylated (O-GlcNAcylated), in a cell cycle-dependent manner.
Cellular localizationCytoplasm. Nucleus, nucleoplasm. Nucleus matrix.
- Information by UniProt
- CARD2 antibody
- CK 8 antibody
- CK-8 antibody
All lanes : Anti-Cytokeratin 8 antibody [EP1628Y] - Cytoskeleton Marker (ab53280) at 1/10000 dilution
Lane 1 : A431 cell lysate
Lane 2 : MCF7 cell lysate
Lane 3 : Wild-type HeLa cell lysate
Lane 4 : KRT8 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 54 kDa
Lanes 1 - 4: Merged signal (red and green). Green - ab53280 observed at 55 kDa. Red - loading control, ab8245 observed at 37 kDa.
ab53280 was shown to react with Cytokeratin 8 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab255400 (knockout cell lysate ab263785) was used. Wild-type and Cytokeratin 8 knockout samples were subjected to SDS-PAGE. ab53280 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 10000 (For unpurified use at 1/25,000 - 1/50,000) dilution and 1 in 20000 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 in 20000 dilution for 1 hour at room temperature before imaging.
Immunocytochemistry/ Immunofluorescence analysis of HT-29 (Human colorectal adenocarcinoma epithelial cell) cells labeling Cytokeratin 9 with Purified ab53280 at 1:500 dilution. Cells were fixed in 100% Methanol and permeabilized with None. 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.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human thyroid carcinoma tissue sections labeling Cytokeratin 8 with Purified ab53280 at 1:250 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 at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.
Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling Cytokeratin 8 with purified ab53280 at 1:20 dilution (red). Cells were fixed with 4% Paraformaldehyde. A Goat anti rabbit IgG (Alexa Fluor® 488) secondary antibody was used at 1:2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).
All lanes : Anti-Cytokeratin 8 antibody [EP1628Y] - Cytoskeleton Marker (ab53280) at 1/10000 dilution (purified)
Lane 1 : A431 (Human epidermoid carcinoma epithelial cell) whole cell lysates
Lane 2 : Human breast cancer lysates
Lane 3 : HaCaT (Human skin keratinocyte) whole cell lysates
Lane 4 : Mouse colon lysates
Lysates/proteins at 20 µg per lane.
All lanes : Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution
Predicted band size: 54 kDa
Blocking and diluting buffer: 5% NFDM/TBST.
ab53280 (purified) at 1:20 dilution (0.2μg) immunoprecipitating Cytokeratin 8 in HeLa whole cell lysate.
Lane 1 (input): HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate, 10μg
Lane 2 (+): ab53280 & HeLa whole cell lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab53280 in HeLa whole cell lysate
For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1:1000 dilution.
Blocking and diluting buffer: 5% NFDM/TBST.
Panx1-/- mice have normal mammary gland epithelial differentiation at lactation
Immunofluorescent analysis of luminal epithelial marker keratin 8 (green) and myoepithelial marker keratin14 (red) revealed a similar staining pattern in Panx1-/- mice compared to control mice during lactation. Paraffin-embedded tissue samples.
Hoescht (blue) denotes nuclei. N = 6. Scale bars = 50 um.
Immunocytochemistry/Immunofluorescence analysis of HeLa (human cervix adenocarcinoma) labelling Cytokeratin 8 with purified ab53280 at 1/500. Cells were fixed with 4% Paraformaldehyde and permeabilised by 0.1% tritonX-100. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody (Ab150077). Nuclei counterstained with DAPI (blue).
Unpurified ab53280 (1:250) staining human Cyotkeratin 8 in human breast adenocarcinoma tissue by immunohistochemistry using paraffin embedded tissue.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse liver tissue sections labeling Cytokeratin 8 with Purified ab53280 at 1:250 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 at 1:0 dilution. PBS instead of the primary antibody was used as the negative control.
Anti-Cytokeratin 8 antibody [EP1628Y] - Cytoskeleton Marker (ab53280) at 1/50000 dilution (unpurified) + A431 cell lysate at 10 µg
Goat anti-Rabbit HRP labeled at 1/2000 dilution
Predicted band size: 54 kDa
Observed band size: 52 kDa why is the actual band size different from the predicted?
Unpurified ab53280 staining Cytokeratin 8 in Mouse liver tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formalin and blocked with 10% serum for 20 minutes at 23°C; antigen retrieval was by heat mediation in a citrate buffer. Samples were incubated with primary antibody (1/75 in TBS + 1% BSA) for 1 hour at 23°C. A Biotin-conjugated Goat anti-rabbit IgG polyclonal (1/500) was used as the secondary antibody.
Immunofluorescent staining of HeLa cells using unpurified ab53280 (1:100).
Image: Courtesy of Dr. Shaohua Li, UMDNJ-Robert Wood Johnson Medical School
Sample: mouse embryonic stem cell-differentiated embryoid bodies (EBs)
Fix in 3%PFA in PBS for 30 min at RTIncubate in 7.5% sucrose-PBS for 3h at RTIncubate in 15% sucrose-PBS at 4 degree Celsius overnightEmbed the EBs in tissue-Tek OCT compoundCut frozen sections to 4-20 µm thickness
Primary antibody 1: Rabbit anti cytokeratin 8 (unpurified ab53280), 1:100
Primary antibody 2: Rat anti-perlecan, 1:100
Secondary antibody 1: Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 488) pre-adsorbed (ab150081), 1:200
Secondary antibody 2: Goat polyclonal Secondary Antibody to Rat IgG - H&L (Cy5®) pre-adsorbed (ab150081), 1:200
Nuclei were counterstained with DAPI
Fluorescent immunohistochemical analysis of paraffin-embedded human ovarian carcinoma tissue using unpurified ab53280. Green-CK8 red-PI.
Fluorescent immunohistochemical analysis of paraffin-embedded human breast carcinoma tissue using unpurified ab53280. Green-CK8 red-PI
Fluorescent immunohistochemical analysis of paraffin-embedded human colonic adenocarcinoma tissue using unpurified ab53280. Green-CK8 red-PI.
Fluorescent immunohistochemical analysis of paraffin-embedded human endometrial carcinoma tissue using unpurified ab53280. Green-CK8 red-PI.
Overlay histogram showing HeLa cells stained with unpurified ab53280 (red line). The cells were fixed with 2% PFA (room temperature, 30 min) and then permeabilized with 1% FACS permeabilizing solution for 30 min. The cells were then incubated in 3% FBS in 1X PBS followed by the antibody (ab53280, 1/20 dilution) for 1 hour at room temperature. The cells were then incubated for 30 min at room temperature with the secondary antibody. An isotype control antibody (black line) was used and an unlabelled sample (blue line) was also used as a control.
Equilibrium disassociation constant (KD)
Learn more about KD
Click here to learn more about KD
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
ab53280 has been referenced in 59 publications.
- Pein M et al. Metastasis-initiating cells induce and exploit a fibroblast niche to fuel malignant colonization of the lungs. Nat Commun 11:1494 (2020). PubMed: 32198421
- Li X et al. The Metastasis Potential Promoting Capacity of Cancer-Associated Fibroblasts Was Attenuated by Cisplatin via Modulating KRT8. Onco Targets Ther 13:2711-2723 (2020). PubMed: 32280245
- Parfitt GJ Immunofluorescence Tomography: High-resolution 3-D reconstruction by serial-sectioning of methacrylate embedded tissues and alignment of 2-D immunofluorescence images. Sci Rep 9:1992 (2019). PubMed: 30760855
- Hu T et al. Developing a Novel and Convenient Model for Investigating Sweat Gland Morphogenesis from Epidermal Stem Cells. Stem Cells Int 2019:4254759 (2019). PubMed: 30863451
- Ezz MA et al. TLR2/4 signaling pathway mediates sperm-induced inflammation in bovine endometrial epithelial cells in vitro. PLoS One 14:e0214516 (2019). PubMed: 30995239