TR-FRET Assays for Endogenous Huntingtin Protein Level in Mouse Cells

J Huntingtons Dis. 2014;3(3):253-9. doi: 10.3233/JHD-140104.

Abstract

High-throughput measurement of huntingtin (Htt) levels is useful for Huntington's disease research. For example, identification of genetic or chemical modifiers that reduce Htt levels by high-throughput screening provides promising strategy for HD drug discovery. In the human cells, high-throughput measurement of Htt levels has been established based on the Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) technology, using the 2B7/MW1 antibody pair. Unfortunately, application of this assay in the mouse cells has been problematic due to discrepancies between TR-FRET signals and Western-blots, possibly caused by non-specific antibody binding. Here we report TR-FRET assays that are able to detect endogenous Htt levels of the mouse striatal cell line (STHdh).

Keywords: Huntington's disease; TR-FRET; high-throughput technology; protein quantification.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cells, Cultured
  • Disease Models, Animal*
  • Fluorescence Resonance Energy Transfer / methods*
  • Gene Expression Profiling / methods
  • Huntingtin Protein
  • Immunoassay / methods*
  • Mice*
  • Nerve Tissue Proteins / analysis
  • Nerve Tissue Proteins / metabolism*
  • Neurons / metabolism*
  • Nuclear Proteins / analysis
  • Nuclear Proteins / metabolism*

Substances

  • Htt protein, mouse
  • Huntingtin Protein
  • Nerve Tissue Proteins
  • Nuclear Proteins