Cytokeratins comprise a diverse group of intermediate filament proteins (IFPs) that are expressed as pairs in both keratinized and non-keratinized epithelial tissue. Cytokeratins play a critical role in differentiation and tissue specialization and function to maintain the overall structural integrity of epithelial cells. Cytokeratins have been found to be useful markers of tissue differentiation which is directly applicable to the characterization of malignant tumors. For example, Cytokeratins 10 and 13 are expressed highly in a subset of squamous cell carcinomas while Cytokeratin 18 is expressed in a majority of adenocarcinomas and basal cell carcinomas. Cytokeratin 18 contains two major phosphorylation sites on Ser 33 and Ser 52. Phosphorylation of Ser 18 is essential for the association of Cytokeratin 18 with 14-3-3 proteins and is involved in keratin organization and distribution.
Background References
1. Kuramoto G. et. al. Preventive effect of oral mucosal epithelial cell sheets on intrauterine adhesions. Hum Reprod 30:406-16 (2015).
2. Huang YS. et. al. RGD surface functionalization of the hydrophilic acrylic intraocular lens material to control posterior capsular opacification. PLoS One 9:e114973 (2014).
Immunofluorescence analysis of paraffin-embedded human kidney tissue labeling Cytokeratin 18 (HA720139F) and Vimentin (EM0401).
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 10% negative goat serum for 1 hour at room temperature, washed with PBS. And then probed with the primary antibodies Cytokeratin 18 (HA720139F, green) at 1/50 dilution and Vimentin (EM0401, red) at 1/1,000 dilution overnight at 4 ℃, washed with PBS.
iFluor™ 594 conjugate-Goat anti-Mouse IgG (HA1126) was used as the secondary antibody at 1/1,000 dilution. DAPI was used as nuclear counterstain.
Immunocytochemistry analysis of SK-Br-3 cells labeling Cytokeratin 18 with Rabbit anti-Cytokeratin 18 antibody (HA720139F) at 1/100 dilution.
Cells were fixed in 4% paraformaldehyde for 15 minutes, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes, and then blocked with 1% BSA for 30 minutes at room temperature. Cells were then incubated with Rabbit anti-Cytokeratin 18 antibody (HA720139F) at 1/100 dilution in 1% BSA overnight at 4 ℃. Nuclear DNA was labelled in blue with DAPI.
Beta tubulin (M1305-2, red) was stained at 1/200 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) were used as the secondary antibody at 1/800 dilution.
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