VCP ELISA Kit (Human)

Intra-assay Precision (Precision within an assay) : 3 samples with low, middle and high level Valosin Containing Protein (VCP) were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays) : 3 samples with low, middle and high level Valosin Containing Protein (VCP) were tested on 3 different plates, 8 replicates in each plate. CV (%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%

Necessary for the fragmentation of Golgi stacks during mitosis and for their reassembly after mitosis. Involved in the formation of the transitional endoplasmic reticulum (tER) . The transfer of membranes from the endoplasmic reticulum to the Golgi apparatus occurs via 50-70 nm transition vesicles which derive from part-rough, part-smooth transitional elements of the endoplasmic reticulum (tER) . Vesicle budding from the tER is an ATP-dependent process. The ternary complex containing UFD1, VCP and NPLOC4 binds ubiquitinated proteins and is necessary for the export of misfolded proteins from the ER to the cytoplasm, where they are degraded by the proteasome. The NPLOC4-UFD1-VCP complex regulates spindle disassembly at the end of mitosis and is necessary for the formation of a closed nuclear envelope. Regulates E3 ubiquitin-protein ligase activity of RNF19A. Component of the VCP/p97-AMFR/gp78 complex that participates in the final step of the sterol-mediated ubiquitination and endoplasmic reticulum-associated degradation (ERAD) of HMGCR. Involved in endoplasmic reticulum stress-induced pre-emptive quality control, a mechanism that selectively attenuates the translocation of newly synthesized proteins into the endoplasmic reticulum and reroutes them to the cytosol for proteasomal degradation (PubMed:26565908) . Also involved in DNA damage response: recruited to double-strand breaks (DSBs) sites in a RNF8- and RNF168-dependent manner and promotes the recruitment of TP53BP1 at DNA damage sites (PubMed:22020440, PubMed:22120668) . Recruited to stalled replication forks by SPRTN: may act by mediating extraction of DNA polymerase eta (POLH) to prevent excessive translesion DNA synthesis and limit the incidence of mutations induced by DNA damage (PubMed:23042607, PubMed:23042605) . Required for cytoplasmic retrotranslocation of stressed/damaged mitochondrial outer-membrane proteins and their subsequent proteasomal degradation (PubMed:16186510, PubMed:21118995) . Essential for the maturation of ubiquitin-containing autophagosomes and the clearance of ubiquitinated protein by autophagy (PubMed:20104022, PubMed:27753622) . Acts as a negative regulator of type I interferon production by interacting with DDX58/RIG-I: interaction takes place when DDX58/RIG-I is ubiquitinated via 'Lys-63'-linked ubiquitin on its CARD domains, leading to recruit RNF125 and promote ubiquitination and degradation of DDX58/RIG-I (PubMed:26471729) . May play a role in the ubiquitin-dependent sorting of membrane proteins to lysosomes where they undergo degradation (PubMed:21822278) . May more particularly play a role in caveolins sorting in cells (PubMed:21822278, PubMed:23335559) . By controlling the steady-state expression of the IGF1R receptor, indirectly regulates the insulin-like growth factor receptor signaling pathway (PubMed:26692333) .

The test principle applied in this kit is Sandwich enzyme immunoassay. The microtiter plate provided in this kit has been pre-coated with an antibody specific to Valosin Containing Protein (VCP) . Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody specific to Valosin Containing Protein (VCP) . Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After TMB substrate solution is added, only those wells that contain Valosin Containing Protein (VCP), biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm +/- 10nm. The concentration of Valosin Containing Protein (VCP) in the samples is then determined by comparing the O.D. of the samples to the standard curve.

Reconstitution & Storage Instructions Western Blotting/Immunoblotting (WB/IB) Protocol Immunohistochemistry (IHC) Protocol Immunocytochemistry (ICC) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Blocking Peptide Competition Protocol (BPCP) Immunoprecipitation (IP) Protocol Antibody Array (AA) Protocol Reconstitution & Storage Instructions Reconstitution & Storage Instructions Western Blotting/Immunoblotting (WB/IB) Protocol Western Blotting/Immunoblotting (WB/IB) Protocol Immunohistochemistry (IHC) Protocol Immunohistochemistry (IHC) Protocol Immunocytochemistry (ICC) Protocol Immunocytochemistry (ICC) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Enzyme-Linked ImmunoSorbent Assay (ELISA) Protocol Blocking Peptide Competition Protocol (BPCP) Blocking Peptide Competition Protocol (BPCP) Immunoprecipitation (IP) Protocol Immunoprecipitation (IP) Protocol Antibody Array (AA) Protocol Antibody Array (AA) Protocol

Component Amount Anti-VCP Microplate 96 Wells (12 x 8 Well Strips) VCP Lyophilized Standard 2 x 20 ng 100X Biotinylated VCP Detector Antibody 120 uL Avidin/HRP Conjugate 120 uL Standard Diluent 1 x 20 mL Detector Antibody Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL 30X Wash Buffer 1 x 20 mL TMB Substrate 1 x 9 mL Stop Solution 1 x 6 mL Component Amount Component Amount Anti-VCP Microplate 96 Wells (12 x 8 Well Strips) Anti-VCP Microplate 96 Wells (12 x 8 Well Strips) VCP Lyophilized Standard 2 x 20 ng VCP Lyophilized Standard 2 x 20 ng 100X Biotinylated VCP Detector Antibody 120 uL 100X Biotinylated VCP Detector Antibody 120 uL Avidin/HRP Conjugate 120 uL Avidin/HRP Conjugate 120 uL Standard Diluent 1 x 20 mL Standard Diluent 1 x 20 mL Detector Antibody Diluent 1 x 12 mL Detector Antibody Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL Conjugate Diluent 1 x 12 mL 30X Wash Buffer 1 x 20 mL 30X Wash Buffer 1 x 20 mL TMB Substrate 1 x 9 mL TMB Substrate 1 x 9 mL Stop Solution 1 x 6 mL Stop Solution 1 x 6 mL

This assay has high sensitivity and excellent specificity for detection of Valosin Containing Protein (VCP) . No significant cross-reactivity or interference between Valosin Containing Protein (VCP) and analogues was observed.