Human Predicted to work with:
Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) corresponding to Human Cytokeratin 18 conjugated to Keyhole Limpet Haemocyanin (KLH) (Cysteine residue). (Peptide available as ab26325, ab26326)
Hela, Jurkat and A431 cell lysates
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml.
Use 1µg for 106 cells. ab171870-Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.
Use a concentration of 5 µg/ml.
1/2000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Use a concentration of 1 µg/ml. Detects a band of approximately 48 kDa (predicted molecular weight: 48 kDa).
Involved in the uptake of thrombin-antithrombin complexes by hepatic cells (By similarity). When phosphorylated, plays a role in filament reorganization. Involved in the delivery of mutated CFTR to the plasma membrane. Together with KRT8, is involved in interleukin-6 (IL-6)-mediated barrier protection.
Expressed in colon, placenta, liver and very weakly in exocervix. Increased expression observed in lymph nodes of breast carcinoma.
Involvement in disease
Defects in KRT18 are a cause of cirrhosis (CIRRH) [MIM:215600].
Belongs to the intermediate filament family.
Phosphorylation at Ser-34 increases during mitosis. Hyperphosphorylated at Ser-53 in diseased cirrhosis liver. Phosphorylation increases by IL-6. Proteolytically cleaved by caspases during epithelial cell apoptosis. Cleavage occurs at Asp-238 by either caspase-3, caspase-6 or caspase-7. O-glycosylated at multiple sites; glycans consist of single N-acetylglucosamine residues.
ICC/IF image of ab24561 stained human HeLa cells. The cells were PFA fixed (10 min), permabilised in TBS-T (20 min) and incubated with the antibody (ab24561, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal goat serum / 0.3M glycine was used to quench autofluorescence and block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Cytokeratin 18 antibody (ab24561)This image is courtesy of an abreview submitted by Antibody Solutions Ltd.
ab24561 (1/2000) staining Cytokeratin 18 in paraffin-embedded Human liver tissue. Tissue underwent fixation in formaldehyde, peroxidase blocking, protein blocking and heat mediated antigen retrieval. The secondary antibody was goat anti rabbit/mouse conjugated to HRP. For further experimental details please refer to abreview.
Overlay histogram showing MCF7 cells stained with ab24561 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab24561, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (polyclonal) (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in MCF7 cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
- Anti-Cytokeratin 18 antibody (ab24561)
Cytokeratin 18 was immunoprecipitated using 0.5mg A431 whole cell extract, 5µg of Rabbit polyclonal to Cytokeratin 18 and 50µl of protein G magnetic beads (+). No antibody was added to the control (-). The antibody was incubated under agitation with Protein G beads for 10min, A431 whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation. Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab24561. Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697). Band: 48kDa; Cytokeratin 18