Revvity, 64LRRKS1PEH, HTRF Human Phospho-LRRK2 (Ser1292) Detection Kit, 10,000 Assay Points

Overview
Leucine-Rich Repeat Kinase 2 (LRRK2) is a multi-domain protein that phosphorylates a subset of Rab GTPases. The LRRK2/Rab pathway operates at the interface of vesicular trafficking, lysosomal functioning, and autophagy, and thus plays a crucial role in immune response and neuroinflammation. Ser1292 autophosphorylation serves as an indicator of LRRK2 kinase activity. Pathogenic LRRK2 mutations, such as the common mutation G2019S, increase LRRK2 kinase activity, resulting in a toxic hyperactive protein that contributes to the Parkinson’s Disease (PD) phenotype. Consequently, LRRK2 inhibitors and degraders are considered promising therapeutic agents.
How it works
Phospho-LRRK2 (Ser1292) assay principle

The Phospho-LRRK2 (Ser1292) assay measures LRRK2 when phosphorylated at Ser1292. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The assay uses 2 antibodies, one labeled with a donor fluorophore and the other with an acceptor. The first antibody was selected for its specific binding to the phosphorylated motif on the protein, and the second for its ability to recognize the protein independently of its phosphorylation state. Protein phosphorylation leads to an immune-complex formation involving both labeled antibodies, and which brings the donor fluorophore into close proximity to the acceptor, thereby generating a FRET signal. Its intensity is directly proportional to the concentration of phosphorylated protein present in the sample and provides a means of assessing the protein's phosphorylation state under a no-wash assay format.

 
Phospho-LRRK2 (Ser1292) two-plate assay protocol

The two-plate protocol involves culturing cells in a 96-well plate before lysis, then transferring lysates into a 384-well low volume detection plate before the addition of Activation buffer followed by Phospho-LRRK2 (Ser1292) HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored. Application-Cell Signaling
Automation Compatible-Yes
Brand-HTRF
Detection Modality-HTRF
Lysis Buffer Compatibility-Lysis Buffer 4
Molecular Modification-Phosphorylation
Product Group-Kit
Sample Volume-16 µL
Shipping Conditions-Shipped in Dry Ice
Target-LRRK2
Target Class-Phosphoproteins
Target Species-Human
Technology-TR-FRET
Therapeutic Area-Neuroscience
Unit Size-10,000 assay points