Gold-Conjugated Nanobodies for Targeted Imaging Using High-Resolution Secondary Ion Mass Spectrometry.

Paola Agüi-Gonzalez,Silvio O Rizzoli,Nhu T N Phan,Tal M Dankovich

doi:10.3390/nano11071797

Abstract

Nanoscale imaging with the ability to identify cellular organelles and protein complexes has been a highly challenging subject in the secondary ion mass spectrometry (SIMS) of biological samples. This is because only a few isotopic tags can be used successfully to target specific proteins or organelles. To address this, we generated gold nanoprobes, in which gold nanoparticles are conjugated to nanobodies. The nanoprobes were well suited for specific molecular imaging using NanoSIMS at subcellular resolution. They were demonstrated to be highly selective to different proteins of interest and sufficiently sensitive for SIMS detection. The nanoprobes offer the possibility of correlating the investigation of cellular isotopic turnover to the positions of specific proteins and organelles, thereby enabling an understanding of functional and structural relations that are currently obscure.

Highlights

Nanoscale secondary ion mass spectrometry (NanoSIMS), an imaging technique characterized by high spatial resolution and high sensitivity, has become a valuable analytical tool for molecular imaging in biological research [1]
NPswith witha afunctional functionalgroup group maleimide were conjugated to the nanobodies havthe cysteine residues via the cysteine maleimide reaction
The indirect immunostaining was by the Au NPs coupled to the nanobody against a light chain of the mouse antibody facilitated by the Au NPs coupled to the nanobody against a light chain of the mouse (Au anti-mouse secondary nanobody), which allowed us to label any proteins of interest antibody (Au anti-mouse secondary nanobody), which allowed us to label any proteins recognized beforehand with a mouse antibody

Summary

Introduction

Nanoscale secondary ion mass spectrometry (NanoSIMS), an imaging technique characterized by high spatial resolution and high sensitivity, has become a valuable analytical tool for molecular imaging in biological research [1]. Across the broad range of achievable lateral resolutions with SIMS instruments, NanoSIMS allows the detection of elemental and small ion species at a spatial resolution down to 50 nm. The lateral resolution achieved with NanoSIMS is relatively comparable to super-resolution light microscopy and electron microscopy techniques. In SIMS, the spatial resolution depends both on the type of the primary ion beam and the abundance of the analytes. In ToF-SIMS, with a bismuth liquid metal ion gun, a spatial resolution down to 500 nm could be achieved. Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations

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Results

Discussion

Conclusion