The mucins are a family of highly glycosylated, secreted proteins with a basic structure consisting of a variable number of tandem repeats (VNTRs). Membrane-associated and secretory Mucins are high molecular weight glycoproteins of the glycocalyx (polysaccharide biofilm) that protects mucosal epithelium from particulate matter and microorganisms. Epithelial Mucins are large, secreted and cell surface glycoproteins crucial for adhesion modulation, signaling and epithelial cell protection. The number of repeats is highly polymorphic and varies among different alleles. The Mucin family consists of Mucins 1-4, Mucin 5 (AC and B), Mucins 6-8, Mucins 11-13 and Mucins 15-17. The Mucin 16 protein (also commonly referred to as CA125), encoded for by the gene MUC16, is a very high molecular weight tumor antigen consisting of three domains: a carboxy terminal domain, an extracellular domain and an amino terminal domain. Mucin 16, an ovarian cancer-associated antigen, is used as a marker to monitor the progress of epithelial ovarian cancer. It is a hydrophilic membrane-associated protein that may be involved in vitamin A functions.
Background References
1. Vitiazeva V et al. The O-Linked Glycome and Blood Group Antigens ABO on Mucin-Type Glycoproteins in Mucinous and Serous Epithelial Ovarian Tumors. PLoS One 10:e0130197 (2015).
2. Garg G et al. Novel treatment option for MUC16-positive malignancies with the targeted TRAIL-based fusion protein Meso-TR3. BMC Cancer 14:35 (2014).
Tissue Specificity
Expressed in corneal and conjunctival epithelia (at protein level). Overexpressed in ovarian carcinomas and ovarian low malignant potential (LMP) tumors as compared to the expression in normal ovarian tissue and ovarian adenomas.
Post-translational Modification
Heavily O-glycosylated; expresses both type 1 and type 2 core glycans.; Heavily N-glycosylated; expresses primarily high mannose and complex bisecting type N-linked glycans.; May be phosphorylated. Phosphorylation of the intracellular C-terminal domain may induce proteolytic cleavage and the liberation of the extracellular domain into the extracellular space.; May contain numerous disulfide bridges. Association of several molecules of the secreted form may occur through interchain disulfide bridges providing an extraordinarily large gel-like matrix in the extracellular space or in the lumen of secretory ducts.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1611-73) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
ICC staining of CA125/MUC16 in HepG2 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1611-73, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
ICC staining of CA125/MUC16 in Hela cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1611-73, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
ICC staining of CA125/MUC16 in SKOV-3 cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1611-73, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
Immunohistochemical analysis of paraffin-embedded human endometrium tissue with Rabbit anti-CA125/MUC16 antibody (ET1611-73) at 1/1,000 dilution.
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 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1611-73) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Flow cytometric analysis of CA125/MUC16 was done on Hela cells. The cells were fixed, permeabilized and stained with the primary antibody (ET1611-73, 1/50) (red). After incubation of the primary antibody at room temperature for an hour, the cells were stained with a Alexa Fluor 488-conjugated Goat anti-Rabbit IgG Secondary antibody at 1/1000 dilution for 30 minutes.Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
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