Detection of endogenous LRP6 expressed on human cells by monoclonal antibodies specific for the native conformation

J Immunol Methods. 2010 Jan 31;352(1-2):153-60. doi: 10.1016/j.jim.2009.11.016. Epub 2009 Nov 26.

Abstract

LRP6 is a cell surface molecule that plays a critical role in the Wnt signaling pathway, and is implicated in numerous human diseases. Studies of cellular signaling mediated by LRP6 have relied on overexpression experiments, due to the lack of good monoclonal antibodies (mAbs) reactive with native LRP6 ectodomain. By using native recombinant LRP6 ectodomain fragment produced in mammalian expression system, we succeeded in developing a panel of anti-human LRP6 mAbs. Selected mAbs were capable of staining endogenous LRP6 on cell surface by using flow cytometry and immunofluorescence microscopy, and enriching detergent-solubilized LRP6 from cell lysate by immunoprecipitation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Monoclonal*
  • Cell Separation
  • Epitopes
  • Flow Cytometry
  • HeLa Cells
  • Humans
  • LDL-Receptor Related Proteins / agonists
  • LDL-Receptor Related Proteins / biosynthesis
  • LDL-Receptor Related Proteins / genetics
  • LDL-Receptor Related Proteins / immunology*
  • Low Density Lipoprotein Receptor-Related Protein-6
  • Mice
  • Mice, Inbred BALB C
  • Microscopy, Fluorescence
  • Protein Conformation
  • Protein Structure, Tertiary / genetics
  • Recombinant Proteins / genetics
  • Recombinant Proteins / immunology*
  • Recombinant Proteins / metabolism
  • Signal Transduction / drug effects
  • Signal Transduction / genetics
  • T Cell Transcription Factor 1 / metabolism
  • Transfection
  • Wnt Proteins / pharmacology
  • Wnt3 Protein

Substances

  • Antibodies, Monoclonal
  • Epitopes
  • LDL-Receptor Related Proteins
  • LRP6 protein, human
  • Low Density Lipoprotein Receptor-Related Protein-6
  • Lrp6 protein, mouse
  • Recombinant Proteins
  • T Cell Transcription Factor 1
  • Wnt Proteins
  • Wnt3 Protein