Correlative 3D cryo X-ray imaging reveals intracellular location and effect of designed antifibrotic protein–nanomaterial hybrids

J. Groen; A. Palanca; A. Aires; J. J. Conesa; D. Maestro; S. Rehbein; M. Harkiolaki; A. V. Villar; A. L. Cortajarena; E. Pereiro

doi:10.1039/D1SC04183E

Correlative 3D cryo X-ray imaging reveals intracellular location and effect of designed antifibrotic protein–nanomaterial hybrids†

J. Groen,

^a A. Palanca,

^bc A. Aires,^d J. J. Conesa,^ae D. Maestro,^b S. Rehbein,^f M. Harkiolaki,^g A. V. Villar,

^bh A. L. Cortajarena

*^di and E. Pereiro

*^a

Author affiliations

* Corresponding authors

^a MISTRAL Beamline, Experiments Division, ALBA Synchrotron Light Source, Cerdanyola del Valles, 08290 Barcelona, Spain
E-mail: epereiro@cells.es

^b Instituto de Biomedicina y Biotecnologia de Cantabria (IBBTEC), University of Cantabria, CSIC, 39011 Santander, Spain

^c Department of Anatomy and Cell Biology, University of Cantabria, 39011 Santander, Spain

^d Center for Cooperative Research in Biomaterials (CIC biomaGUNE), Basque Research and Technology Alliance (BRTA), Paseo de Miramón 194, Donostia San Sebastian, Spain
E-mail: alcortajarena@cicbiomagune.es

^e National Center for Biotechnology CSIC (CNB-CSIC), Department of Macromolecular Structures, Cantoblanco, 28049 Madrid, Spain

^f Helmholtz-Zentrum Berlin für Materialien und Energie, Bessy II, D-12489 Berlin, Germany

^g Beamline B24, Diamond Light Source, Harwell Science and Innovation Campus, Didcot, Oxfordshire, UK

^h Department of Physiology and Pharmacology, University of Cantabria, Avd. Herrera Oria s/n, Santander, Spain

ⁱ Ikerbasque, Basque Foundation for Science, 48009 Bilbao, Spain

Abstract

Revealing the intracellular location of novel therapeutic agents is paramount for the understanding of their effect at the cell ultrastructure level. Here, we apply a novel correlative cryo 3D imaging approach to determine the intracellular fate of a designed protein–nanomaterial hybrid with antifibrotic properties that shows great promise in mitigating myocardial fibrosis. Cryo 3D structured illumination microscopy (cryo-3D-SIM) pinpoints the location and cryo soft X-ray tomography (cryo-SXT) reveals the ultrastructural environment and subcellular localization of this nanomaterial with spatial correlation accuracy down to 70 nm in whole cells. This novel high resolution 3D cryo correlative approach unambiguously locates the nanomaterial after overnight treatment within multivesicular bodies which have been associated with endosomal trafficking events by confocal microscopy. Moreover, this approach allows assessing the cellular response towards the treatment by evaluating the morphological changes induced. This is especially relevant for the future usage of nanoformulations in clinical practices. This correlative super-resolution and X-ray imaging strategy joins high specificity, by the use of fluorescence, with high spatial resolution at 30 nm (half pitch) provided by cryo-SXT in whole cells, without the need of staining or fixation, and can be of particular benefit to locate specific molecules in the native cellular environment in bio-nanomedicine.

Chemical Science

Correlative 3D cryo X-ray imaging reveals intracellular location and effect of designed antifibrotic protein–nanomaterial hybrids†

Abstract

Supplementary files

Article information

Download Citation

Permissions

Correlative 3D cryo X-ray imaging reveals intracellular location and effect of designed antifibrotic protein–nanomaterial hybrids

Social activity

Search articles by author

Spotlight

Advertisements