The PCSK9(D374T)-LDLR TR-FRET Assay Kit is designed to measure the inhibition of the high affinity PCSK9 mutant (D374T) binding to LDLR in a homogeneous 384 reaction format. The PCSK9 Assay Kit assay requires no time-consuming washing steps, making it especially suitable for high throughput screening applications. The assay procedure is straightforward and simple; a sample containing europium-labeled (Eu) LDLR ectodomain, dye-labeled acceptor, biotin-labeled PCSK9(D374T), and an inhibitor is incubated for two hours. Then, the fluorescence intensity is measured using a fluorescence reader. The kit contains enough purified proteins, Eu-donor, dye-labeled acceptor and assay buffer to perform 384 reactions.
Need us to run inhibitor screens or profile your compounds against PCSK9(D374T)-LDLR? Check out our PCSK9 Screening Services.
*The concentration of LDLR-Eu and PCSK9 are lot-specific and will be indicated on the tubes containing the protein
UniProt #
Q8NBP7
Background
PCSK9 is a crucial player in the regulation of plasma cholesterol homeostasis. It binds to the ectodomain of hepatic low-density lipid receptor family members: low density lipoprotein receptor (LDLR), very low density lipoprotein receptor (VLDLR), apolipoprotein E receptor (LRP1/APOER) and apolipoprotein receptor 2 (LRP8/APOER2), and promotes their degradation. PCSK9 acts via a non-proteolytic mechanism to enhance the degradation of the hepatic LDLR through a clathrin LDLRAP1/ARH-mediated pathway. May prevent the recycling of LDLR from endosomes to the cell surface or direct it to lysosomes for degradation. The D374T mutation results in higher affinity of PCSK9 for LDLR.
References
1. Chan, J.C. et al. (2009). Proc. Natl. Acad. Sci. USA, 106, 9820-9825. 2. Liang, H., et al. (2012). J. Pharmacol. Exp. Ther. 340 2289-236.
Storage/Stability
At least 6 months from date of receipt when stored as directed.
Instructions for Use
See assay kit data sheet for detailed protocol.
Applications
Useful for screening small molecular inhibitors for drug discovery and HTS applications.
Shipping Temperature
-80°C
Notes
MATERIALS OR INSTRUMENTS REQUIRED BUT NOT SUPPLIED: Fluorescent microplate reader capable of measuring Time Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Adjustable micropipettor and sterile tips
Disclaimers and Limitations
BPS Bioscience assay kits are validated using the listed components according to our specific protocol. Any deviations to kit components or protocols, such as using different proteins, cell/tissue lysates, or buffers (including using the same component from other commercial sources) will void any warranties on the performance of the assay kit and is not recommended.
End-users should immediately report any issues to [email protected] upon receipt of the kit, such as missing components, damaged vials, no dry ice, etc.