Rat Valosin Containing Protein (VCP) ELISA Kit

Principle of the Assay

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 Rat VCP. Standards or samples are added to the appropriate microtiter plate wells then with a biotin-conjugated antibody specific to Rat 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 Rat 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 Rat VCP in the samples is then determined by comparing the OD of the samples to the standard curve.

For use with: Tissue homogenates and other biological fluids.

For Research Use Only. Not for use in diagnostic procedures.

Target Information

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) (PubMed:10930451, PubMed:12411482). Vesicle budding from the tER is an ATP-dependent process (PubMed:10930451, PubMed:12411482). 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 (PubMed:10930451, PubMed:12411482). 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. Plays a role in the regulation of stress granules (SGs) clearance process upon arsenite-induced response (By similarity). 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. 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. Together with SPRTN metalloprotease, involved in the repair of covalent DNA-protein cross-links (DPCs) during DNA synthesis. Involved in interstrand cross-link repair in response to replication stress by mediating unloading of the ubiquitinated CMG helicase complex. Required for cytoplasmic retrotranslocation of stressed/damaged mitochondrial outer-membrane proteins and their subsequent proteasomal degradation. Essential for the maturation of ubiquitin-containing autophagosomes and the clearance of ubiquitinated protein by autophagy. 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 (By similarity). May play a role in the ubiquitin-dependent sorting of membrane proteins to lysosomes where they undergo degradation (By similarity). May more particularly play a role in caveolins sorting in cells (By similarity). By controlling the steady-state expression of the IGF1R receptor, indirectly regulates the insulin-like growth factor receptor signaling pathway (By similarity).

GENE ID	116643
SWISS PROT	P46462
SYNONYMS	IBMPFD; p97; TERA; Transitional Endoplasmic Reticulum ATPase; 15S Mg(2+)-ATPase p97 subunit

Materials Supplied

Kit Components	96 Wells Quantity/Size
Pre-Coated Microplate	48T: 6 strips x 8 wells; 96T: 12 strips x 8 wells
Standard (Lyophilized)	48T: 1 vial; 96T: 2 vials
Biotinylated Antibody (100×)	48T: 60 μL; 96T: 120 μL
Streptavidin-HRP (100×)	48T: 60 μL; 96T: 120 μL
Standard/Sample Diluent Buffer	48T: 10 mL; 96T: 20 mL
Biotinylated Antibody Diluent	48T: 6 mL; 96T: 12 mL
HRP Diluent	48T: 6 mL; 96T: 12 mL
Wash Buffer (25×)	48T: 10 mL; 96T: 20 mL
TMB Substrate Solution	48T: 6 mL; 96T: 10 mL
Stop Reagent	48T: 3 mL; 96T: 6 mL
Plate Covers	48T: 1 piece; 96T: 2 pieces

Storage

Storage

*If the kit is opened, Store the whole kit at 4°C. If the kit is not used up in 1 week. Store the Pre-Coated Microplate, Standard ,Biotinylated Antibody and Streptavidin-HRP at -20°C, the rest reagents at 4°C, please used up within 6 months. *If the kit is not opened, store the whole kit: 4°C(short time storage, valid for 6 months); -20°C (long-term storage, valid for 1 year). Avoid repeated freeze-thaw cycles.

Performance Characteristics

REPEATABILITY	Intra-assay Precision (Precision within an assay)：CV% < 8% Three samples of known concentration were tested twenty times on 1 plate to assess intra-assay precision. Inter-assay Precision (precision between assays)：CV% < 10% Three samples of known concentration were tested in forty separate assays to assess inter-assay precision.
SENSITIVITY	The minimum detectable dose was 0.112ng/mL.
ASSAY RANGE	0.32–20ng/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for detection of Rat VCP. No significant cross-reactivity or interference between Rat VCP and analogues was observed.