The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Peptide ELISA: Antibody detection limit dilution 1/8000.
WB: Use at a concentration of 0.3 - 1 µg/ml. Detects a band of approximately 37 kDa (predicted molecular weight: 52 kDa).
Not yet tested in other applications.
Optimal dilutions/concentrations should be determined by the end user.
Isoform 1 functions as extracellular chaperone that prevents aggregation of nonnative proteins. Prevents stress-induced aggregation of blood plasma proteins. Inhibits formation of amyloid fibrils by APP, APOC2, B2M, CALCA, CSN3, SNCA and aggregation-prone LYZ variants (in vitro). Does not require ATP. Maintains partially unfolded proteins in a state appropriate for subsequent refolding by other chaperones, such as HSPA8/HSC70. Does not refold proteins by itself. Binding to cell surface receptors triggers internalization of the chaperone-client complex and subsequent lysosomal or proteasomal degradation. Secreted isoform 1 protects cells against apoptosis and against cytolysis by complement. Intracellular isoforms interact with ubiquitin and SCF (SKP1-CUL1-F-box protein) E3 ubiquitin-protein ligase complexes and promote the ubiquitination and subsequent proteasomal degradation of target proteins. Promotes proteasomal degradation of COMMD1 and IKBKB. Modulates NF-kappa-B transcriptional activity. Nuclear isoforms promote apoptosis. Mitochondrial isoforms suppress BAX-dependent release of cytochrome c into the cytoplasm and inhibit apoptosis. Plays a role in the regulation of cell proliferation.
Detected in blood plasma, cerebrospinal fluid, milk, seminal plasma and colon mucosa. Detected in the germinal center of colon lymphoid nodules and in colon parasympathetic ganglia of the Auerbach plexus (at protein level). Ubiquitous. Detected in brain, testis, ovary, liver and pancreas, and at lower levels in kidney, heart, spleen and lung.
Belongs to the clusterin family.
Isoform 1 is proteolytically cleaved on its way through the secretory system, probably within the Golgi lumen. Polyubiquitinated, leading to proteasomal degradation. Heavily N-glycosylated. About 30% of the protein mass is comprised of complex N-linked carbohydrate.
Secreted. Can retrotranslocate from the secretory compartments to the cytosol upon cellular stress and Nucleus. Cytoplasm. Mitochondrion membrane. Cytoplasm, cytosol. Microsome. Endoplasmic reticulum. Cytoplasmic vesicle, secretory vesicle, chromaffin granule. Isoforms lacking the N-terminal signal sequence have been shown to be cytoplasmic and/or nuclear. Secreted isoforms can retrotranslocate from the secretory compartments to the cytosol upon cellular stress. Detected in perinuclear foci that may be aggresomes containing misfolded, ubiquitinated proteins. Detected at the mitochondrion membrane upon induction of apoptosis.
Anti-Clusterin antibody (ab79280) at 0.3 µg/ml + Rat Brain lysate (in RIPA buffer) at 35 µg
Predicted band size : 52 kDa Observed band size : 37 kDa (why is the actual band size different from the predicted?) The observed molecular weight corresponds to earlier findings in literature with different antibodies (Miyata et al, Circulation. 2001 Sep 18;104(12):1407-12.; PMID: 11560857).