A Filter Retardation Assay Facilitates the Detection and Quantification of Heat-Stable, Amyloidogenic Mutant Huntingtin Aggregates in Complex Biosamples

  • Anne Ast
  • Franziska Schindler
  • Alexander Buntru
  • Sigrid Schnoegl
  • Erich E. WankerEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1780)


N-terminal mutant huntingtin (mHTT) fragments with pathogenic polyglutamine (polyQ) tracts spontaneously form stable, amyloidogenic protein aggregates with a fibrillar morphology. Such structures are detectable in brains of Huntington’s disease (HD) patients and various model organisms, suggesting that they play a critical role in pathogenesis. Heat-stable, fibrillar mHTT aggregates can be detected and quantified in cells and tissues using a denaturing filter retardation assay (FRA). Here, we describe step-by-step protocols and experimental procedures for the investigation of mHTT aggregates in complex biosamples using FRAs. The methods are illustrated with examples from studies in cellular, transgenic fly, and mouse models of HD, but can be adapted for any disease-relevant protein with amyloidogenic polyQ tracts.


Filter retardation assay Protein misfolding Protein aggregation Coaggregation Insoluble aggregates Amyloidogenesis Seeding and spreading of protein aggregates 



This study received funding from the EC funding initiative ERA-NET NEURON, consortium “ABETA ID” funded by the German Federal Ministry for Education and Research (BMBF), grant no. 01W1301, the Berlin Institute of Health Collaborative Research Grant no. 1.1.2.a.3 “Elucidating the proteostasis network to control Alzheimer’s disease” funded by the German Federal Ministry for Education and Research (BMBF), the Helmholtz Validation Fund grant no. HVF-0013 “Enabling Technologies for Drug Discovery against Protein Misfolding Diseases” funded by the Helmholtz Association, Germany, (to E.E.W.), and the Max Delbrück Center for Molecular Medicine in the Helmholtz Association for application-oriented research (to E.E.W.). Anne Ast and Franziska Schindler contributed equally to this work.


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Anne Ast
    • 1
  • Franziska Schindler
    • 1
  • Alexander Buntru
    • 1
  • Sigrid Schnoegl
    • 1
  • Erich E. Wanker
    • 1
    Email author
  1. 1.NeuroproteomicsMax Delbrueck Center for Molecular MedicineBerlinGermany

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