Quantitative Detection of Biological Nanoparticles Using Twilight Off-Axis Holographic Microscopy: Insights on Complex Formation between PEGylated Gold Nanoparticles and Lipid Vesicles
Artikel i vetenskaplig tidskrift, 2025

The detection of biological nanoparticles (NPs), such as viruses and extracellular vesicles (EVs), plays a critical role in medical diagnostics. However, these particles are optically faint, making microscopic detection in complex solutions challenging. Recent advancements have demonstrated that distinguishing between metallic and dielectric signals with twilight off-axis holographic microscopy makes it possible to differentiate between metal and biological NPs and to quantify complexes formed from metal and biological NPs binding together. Here, this method is employed to investigate complex formation through specific interactions between streptavidin (StrAv)-modified gold NPs (StrAv-AuNPs) and large biotin-containing unilamellar lipid vesicles (biotin-LUVs), serving as virus and EV mimics. To minimize AuNP self-aggregation during functionalization of PEGylated 25 nm radius AuNPs with tetrameric StrAv, 0.06% biotin-PEG (similar to 5 biotin per AuNP) was used, which also serves to ensure that aggregation involving multiple LUVs is effectively prevented. While the StrAv-biotin ratio did not significantly affect AuNP self-aggregation upon fabrication of StrAv-AuNPs, a 1000-fold StrAv excess with respect to biotin-PEG on the AuNPs was required to fabricate StrAv-AuNPs with the anticipated reactivity with biotin-LUVs. Through a combination of waveguide scattering microscopy, surface plasmon resonance, and twilight off-axis holographic microscopy, we demonstrate that this likely stems from a dramatic reduction in the association rate constant between StrAv and biotin within the PEG layer. Furthermore, by using a mixture of 3 kDa nonbiotinylated PEG and 5 kDa biotin-PEG, functional StrAv-AuNPs were successfully fabricated at an orders of magnitude lower StrAv-to-biotin ratio, enabling a sub-pM limit of detection of biotin-LUVs using off-axis holography.

Författare

Julia Andersson

Chalmers, Fysik, Nano- och biofysik

Anders Lundgren

Göteborgs universitet

Institutionen för kemi, GU

Erik Olsén

University of British Columbia (UBC)

Institutionen för fysik, GU

Petteri Parkkila

Chalmers, Fysik, Nano- och biofysik

Daniel Midtvedt

Göteborgs universitet

Institutionen för fysik, GU

Fredrik Höök

Chalmers, Fysik, Nano- och biofysik

Journal of Physical Chemistry B

1520-6106 (ISSN) 1520-5207 (eISSN)

Vol. 129 37 9506-9516

Funktionell leverans av nukleotid-baserade läkemedel

Stiftelsen för Strategisk forskning (SSF) (IRC15-0065), 2017-03-01 -- 2024-12-31.

Ämneskategorier (SSIF 2025)

Atom- och molekylfysik och optik

Biofysik

Annan fysik

DOI

10.1021/acs.jpcb.5c04228

PubMed

40924948

Mer information

Senast uppdaterat

2025-09-26