Reproducible Formation of Insulin Superstructures: Amyloid-Like Fibrils, Spherulites, and Particulates

Research output: Chapter in Book/Report/Conference proceedingBook chapterResearchpeer-review

  • Camilla Thorlaksen
  • Martin Busch Neergaard
  • Minna Groenning
  • Foderà, Vito

Inducing protein aggregation in vitro under various formulation and stress conditions may lead to an increased understanding of the different association routes a protein can undergo. However, a range of factors can affect the aggregation process, often leading to heterogenous samples and experimental irreproducibility between labs. Here, we present detailed methods to reproducibly form homogenous samples of superstructures: amyloid-like fibrils, spherulites, and particulates from human insulin. We discuss pitfalls and good practice in the lab, with the aim of creating awareness on the potential sources of artefacts for protein stability and aggregation studies.

Original languageEnglish
Title of host publicationProtein Aggregation : Methods in Molecular Biology
EditorsA. Stanisław Cieplak
PublisherHumana Press
Publication date2023
ISBN (Print)978-1-0716-2596-5
ISBN (Electronic)978-1-0716-2597-2
Publication statusPublished - 2023
SeriesMethods in Molecular Biology

Bibliographical note

Funding Information:
The work was funded, and all protein material was provided by Novo Nordisk A/S. For use of the TEM, we acknowledge the Core Facility for Integrated Microscopy, Faculty of Health and Medical Sciences, University of Copenhagen. For use of the MFI image evaluation software, we acknowledge Jesper Søndergaard Marino from Novo Nordisk A/S. Illustrations presented in the chapter were created with V.F. also acknowledges the VIL-LUM FONDEN for the Villum Young Investigator Grant “Protein Superstructures as Smart Biomaterials (ProSmart)” 2018–2023 (project number: 19175). The authors acknowledge Marco van de Weert (University of Copenhagen) for inspiring discussions on reproducibility in protein stability and aggregation studies.

Publisher Copyright:
© 2023, Springer Science+Business Media, LLC, part of Springer Nature.

    Research areas

  • Aggregation, Amyloid-like fibrils, Insulin, Micro-Flow Imaging, Particulates, Spherulites, Superstructures, Transmission electron microscopy

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