Label-Free Digital Holographic Microscopy for In Vitro Cytotoxic Effect Quantification of Organic Nanoparticles.

Kai Moritz Eder,Álvaro Barroso,Björn Kemper,Anne Marzi,Steffi Ketelhut,Jürgen Schnekenburger

doi:10.3390/cells11040644

Abstract

Cytotoxicity quantification of nanoparticles is commonly performed by biochemical assays to evaluate their biocompatibility and safety. We explored quantitative phase imaging (QPI) with digital holographic microscopy (DHM) as a time-resolved in vitro assay to quantify effects caused by three different types of organic nanoparticles in development for medical use. Label-free proliferation quantification of native cell populations facilitates cytotoxicity testing in biomedical nanotechnology. Therefore, DHM quantitative phase images from measurements on nanomaterial and control agent incubated cells were acquired over 24 h, from which the temporal course of the cellular dry mass was calculated within the observed field of view. The impact of LipImage™ 815 lipidots® nanoparticles, as well as empty and cabazitaxel-loaded poly(alkyl cyanoacrylate) nanoparticles on the dry mass development of four different cell lines (RAW 264.7, NIH-3T3, NRK-52E, and RLE-6TN), was observed vs. digitonin as cytotoxicity control and cells in culture medium. The acquired QPI data were compared to a colorimetric cell viability assay (WST-8) to explore the use of the DHM assay with standard biochemical analysis methods downstream. Our results show that QPI with DHM is highly suitable to identify harmful or low-toxic nanomaterials. The presented DHM assay can be implemented with commercial microscopes. The capability for imaging of native cells and the compatibility with common 96-well plates allows high-throughput systems and future embedding into existing experimental routines for in vitro cytotoxicity assessment.

Highlights

Nanotechnology is an emerging research field in medicine and pharmaceutical sciences
We summarize that the calculations of 24-h dry mass increments retrieved from digital holographic microscopy (DHM) quantitative phase imaging (QPI) images allowed an endpoint quantification of proliferation and nanoparticle cytotoxicity
As proposed by earlier research [14,29,30,52], the data from our study suggests that DHM as a label-free quantitative imaging tool is suitable for in vitro cytotoxicity studies but with the promising capability to be applied in the field of medical nanotechnology

Summary

Introduction

Nanotechnology is an emerging research field in medicine and pharmaceutical sciences. The physico-chemical advantages of nanoparticles are utilized for diagnosis of diseases, enhanced drug delivery, as well as for support in regenerative medicine [1,2]. Inorganic nanoparticles like iron oxide and gold are applied as contrast agents [4,5] while, organic nanomaterials like polymeric or liposomal, as investigated in this study, can fulfill various pharmaceutical and medical functions, such as drug carriers, contrast agents, immunotherapies or gene editors [6,7]. The prominent role of medical nanotechnology in current science and medicine was highlighted by the lipid nanoencapsulation of the mRNA-based COVID-19 vaccine [8,9]. In contrast to industrial nanomaterials, for which primarily risks to humans upon accidental exposure must be considered, for medical applications, the understanding of the entire impact of nanoparticles on cellular processes is essential for both effectiveness and safety [10,11]

Results

Discussion

Conclusion