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Updated: Mar 30, 2026
Biomolecular Imaging of Cellular Uptake of Nanoparticles using Multimodal Nonlinear Optical Microscopy
Published on: May 16, 2022
Exploring the cellular and tissue uptake of nanomaterials in a range of biological samples using multimodal nonlinear
Helinor J Johnston1, Rabah Mouras, David M Brown
1Nano Safety Research Group, School of Life Sciences, Heriot-Watt University, Edinburgh EH14 4AS, UK.
Abstract:
The uptake of nanomaterials (NMs) by cells is critical in determining their potential biological impact, whether beneficial or detrimental. Thus, investigation of NM internalization by cells is a common consideration in hazard and efficacy studies. There are currently a number of approaches that are routinely used to investigate NM-cell interactions, each of which have their own advantages and limitations. Ideally, imaging modalities used to investigate NM uptake by cells should not require the NM to be labelled (e.g. with fluorophores) to facilitate its detection. We present a multimodal imaging approach employing a combination of label-free microscopies that can be used to investigate NM-cell interactions. Coherent anti-Stokes Raman scattering microscopy was used in combination with either two-photon photoluminescence or four-wave mixing (FWM) to visualize the uptake of gold or titanium dioxide NMs respectively. Live and fixed cell imaging revealed that NMs were internalized by J774 macrophage and C3A hepatocyte cell lines (15-31 μg ml(-1)). Sprague Dawley rats were exposed to NMs (intratracheal instillation, 62 μg) and NMs were detected in blood and lung leucocytes, lung and liver tissue, demonstrating that NMs could translocate from the exposure site. Obtained data illustrate that multimodal nonlinear optical microscopy may help overcome current challenges in the assessment of NM cellular uptake and biodistribution. It is therefore a powerful tool that can be used to investigate unlabelled NM cellular and tissue uptake in three dimensions, requires minimal sample preparation, and is applicable to live and fixed cells.
Insights
This study introduces a label-free multimodal imaging technique to track nanomaterial (NM) uptake in cells and tissues. This advanced method visualizes NM internalization and biodistribution without requiring NM labeling, overcoming current limitations in hazard and efficacy studies.
Area of Science:
- Nanotechnology and Materials Science
- Cell Biology and Imaging
- Toxicology and Biodistribution Studies
Background:
- Cellular uptake of nanomaterials (NMs) is crucial for assessing their biological impact, influencing both hazard and efficacy.
- Current methods for studying NM-cell interactions often require NM labeling, limiting their applicability and potentially altering NM behavior.
- There is a need for advanced imaging techniques that can visualize unlabeled NM uptake and biodistribution in complex biological systems.
Purpose of the Study:
- To develop and validate a multimodal, label-free imaging approach for investigating nanomaterial cellular uptake.
- To assess the capability of this technique in visualizing the internalization of gold and titanium dioxide NMs by cell lines.
- To evaluate the biodistribution of NMs in vivo following exposure, demonstrating translocation from the exposure site.
Main Methods:
- Employed a multimodal imaging strategy combining label-free microscopy techniques: Coherent Anti-Stokes Raman Scattering (CARS) microscopy.
- Utilized two-photon photoluminescence or Four-Wave Mixing (FWM) microscopy to visualize gold or titanium dioxide NMs, respectively.
- Applied the technique to both live and fixed J774 macrophage and C3A hepatocyte cell lines, and to Sprague Dawley rats exposed via intratracheal instillation.
Main Results:
- Successfully visualized the internalization of gold and titanium dioxide NMs into J774 macrophage and C3A hepatocyte cell lines at concentrations of 15-31 μg ml⁻¹.
- Detected translocated NMs in blood and lung leukocytes, as well as in lung and liver tissues of Sprague Dawley rats after intratracheal instillation (62 μg).
- Demonstrated the technique's ability to provide 3D visualization of NM cellular and tissue uptake with minimal sample preparation.
Conclusions:
- Multimodal nonlinear optical microscopy offers a powerful, label-free solution for assessing nanomaterial cellular uptake and biodistribution.
- This approach overcomes limitations of current imaging modalities by enabling visualization of unlabeled NMs in live and fixed cells and tissues.
- The technique facilitates a more accurate and comprehensive understanding of NM-cell interactions and in vivo translocation, crucial for safety and efficacy assessments.
