This HTRF kit enables the cell-based quantitative detection of LRRK2 protein phosphorylated at Serine 935 directly in your cell type of interest.

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  • Low sample consumption Low sample consumption
  • No-wash No-wash
  • Validated on PBMC Validated on PBMC

This HTRF kit enables the cell-based quantitative detection of LRRK2 protein phosphorylated at Serine 935 directly in your cell type of interest.

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Overview

The phospho-LRRK2 (Ser935) kit measures endogenous LRRK2 protein phosphorylated at Ser935 directly in your cell type of interest. LRRK2 is a significant target in neuroscience research involving Parkinson’s disease (PD) and neurodegenerative disorders in general.

Benefits

  • VALIDATED ON PATIENT PBMC'S
  • SPECIFICITY

Phospho-LRRK2 (Ser935) assay principle

The phospho-LRRK2 (Ser935) assay measures LRRK2 when phosphorylated at Ser935. Contrary to Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis or transfer. The phospho-LRRK2 (Ser935) assay uses 2 labeled antibodies: one with a donor fluorophore, the other one with an acceptor. The first antibody is selected for its specific binding to the phosphorylated motif on the protein, the second for its ability to recognize the protein independent of its phosphorylation state. Protein phosphorylation enables 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 (Ser935) assay principle

Phospho-LRRK2 (Ser935) 2-plate assay protocol

The 2 plate protocol involves culturing cells in a 96-well plate before lysis then transferring lysates to a 384-well low volume detection plate before adding phospho-LRRK2 (Ser935) HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.
Phospho-LRRK2 (Ser935) 2-plate assay protocol

Phospho-LRRK2 (Ser935) 1-plate assay protocol

Detection of phosphorylated LRRK2 (Ser935) with HTRF reagents can be performed in a single plate used for culturing, stimulation and lysis. No washing steps are required. This HTS designed protocol enables miniaturization while maintaining robust HTRF quality.
Phospho-LRRK2 (Ser935) 1-plate assay protocol

HTRF phospho-LRRK2 assay compared to Western Blot

Murine NIH-3T3 cells were cultured for 2 days until reaching 100% confluency. Following cell lysis, soluble fractions were then collected after a 10 min centrifugation. Serial dilutions of the cell lysate were performed in the supplemented lysis buffer and 16 µL of each dilution were dispensed and analyzed side-by-side by WB and by HTRF. The HTRF phospho-assay is 8-fold more sensitive than the Western Blot: only 2,500 cells are sufficient for minimal signal detection when using the HTRF phospho- LRRK2 assay while 20,000 cells are needed for a Western Blot signal.
HTRF phospho-LRRK2 assay vs Western Blot

Phospho LRRK2 detection in NIH 3T3 cells

Cells were cultured in 6-well plates and treated with increasing concentrations of CZC… and GSK… for 2 hours. Medium was removed and replaced with Cisbio lysis buffer for 1 hour. 16 µL of lysate were taken from each well and transferred to a 384-well plate for detection with HTRF reagents. The Cisbio phosphorylated LRRK2 assay enabled clear determination of both inhibitors’ IC50s, in agreement with literature.
Phospho LRRK2 detection in NIH 3T3 cells
Simplified pathway dissection with HTRF phospho-assays and CyBi-felix liquid handling

Analyse of PI3K/AKT/mTor translational control pathway - Application Notes

Cisbio lysis buffer compatibility

Cell Signaling: Biomarkers, Phospho- & total-protein Assays - Flyers

HTRF cellular phospho-protein assays

Physiologically relevant results fo fast flowing research - Flyers

Save time and money

Switch to HTRF assays - Flyers

Species compatibility

Cell Signaling: Biomarkers, Phospho- & total-protein assays - Flyers

Side-by-side comparison of HTRF, Western Blot, ELISA and AlphaScreen® SureFire®

Do all cell-based kinase assays perform similarly? - Posters

Universal HTRF® phospho-protein platform: from 2D, 3D, primary cells to patient derived tumor cells

Analysis of a large panel of diverse biological samples and cellular models - Posters

HTRF phospho assays reveal subtle drug induced effects in tumor-xenografts

Tumor xenograft analysis: HTRF versus Western blot - Application Notes

HTRF cell-based phospho-protein data normalization

Valuable guidelines for efficiently analyzing and interpreting results - Application Notes

HTRF phospho-total lysis buffer: a universal alternative to RIPA lysis buffers

Increased flexibility of phospho-assays - Application Notes

Exploring LRRK2 phosphorylation with HTRF® assays

LRRK2 kit, a game changer in neuroscience - Posters

Best practices for analyzing brain samples with HTRF® phospho assays for neurosciences

Insider Tips for successful sample treatment - Technical Notes

HTRF Alpha-tubulin Housekeeping kit

Properly interpret your compound effect - Application Notes

Optimize your HTRF cell signaling assays on tissues

HTRF and WB compatible guidelines - Technical Notes

Key guidelines to successful cell signaling experiments

Mastering the art of cell signaling assays optimization - Guides

HTRF phospho-assays reveal subtle drug-induced effects

Detailed protocol and direct comparison with WB - Posters

Best practices for analyzing tumor xenografts with HTRF phospho assays

Protocol for tumor xenograft analysis with HTRF - Technical Notes

How to run a cell based phospho HTRF assay

What to expect at the bench - Videos

Unleash the potential of your phosphorylation research with HTRF

Unmatched ease of use, sensitivity and specificity assays - Videos

Cisbio Product Catalog 2019

All your HTRF assays in one document! - Catalog

A guide to Homogeneous Time Resolved Fluorescence

General principles of HTRF - Guides

How HTRF compares to Western Blot and ELISA

Get the brochure about technology comparison. - Brochures

HTRF® cell signaling platform combined with iCell® Hepatocytes

A solution for phospho-protein analysis in metabolic disorders - Posters

Unleash the potential of your phosphorylation research with HTRF

A fun video introducing you to phosphorylation assays with HTRF - Videos

How to run a cell based phospho HTRF assay

3' video to set up your Phospho assay - Videos

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