High throughput toxicity screening and intracellular detection of nanomaterials

Abstract

With the growing numbers of nanomaterials (NMs), there is a great demand for rapid and reliable ways of testing NM safety—preferably using in vitro approaches, to avoid the ethical dilemmas associated with animal research. Data are needed for developing intelligent testing strategies for risk assessment of NMs, based on grouping and read‐across approaches. The adoption of high throughput screening (HTS) and high content analysis (HCA) for NM toxicity testing allows the testing of numerous materials at different concentrations and on different types of cells, reduces the effect of inter‐experimental variation, and makes substantial savings in time and cost. HTS/HCA approaches facilitate the classification of key biological indicators of NM‐cell interactions. Validation of in vitro HTS tests is required, taking account of relevance to in vivo results. HTS/HCA approaches are needed to assess dose‐ and time‐dependent toxicity, allowing prediction of in vivo adverse effects. Several HTS/HCA methods are being validated and applied for NM testing in the FP7 project NANoREG, including Label‐free cellular screening of NM uptake, HCA, High throughput flow cytometry, Impedance‐based monitoring, Multiplex analysis of secreted products, and genotoxicity methods—namely High throughput comet assay, High throughput in vitro micronucleus assay, and γH2AX assay. There are several technical challenges with HTS/HCA for NM testing, as toxicity screening needs to be coupled with characterization of NMs in exposure medium prior to the test; possible interference of NMs with HTS/HCA techniques is another concern. Advantages and challenges of HTS/HCA approaches in NM safety are discussed. WIREs Nanomed Nanobiotechnol 2017, 9:e1413. doi: 10.1002/wnan.1413For further resources related to this article, please visit the WIREs website.

Immunotoxicity, genotoxicity and epigenetic toxicity of nanomaterials: New strategies for toxicity testing?

“…Within the EC FP7 NANoREG project (www.nanoreg.eu/) HTS/HCA, methods have been reviewed and their adaptation for NM testing was thoroughly assessed. Advantages and challenges pertaining to HTS/HCA approaches in NM safety evaluation have been identified and are summarised in the paper by Collins et al (2017).…”

“…The challenges in evaluating the large number of NMs and their complex variations using resourceintensive and time-consuming in vivo assays, are a motivating factor for developing faster, more economical and reliable assays (Lan et al, 2014;Collins et al, 2017). Several methods have been developed and adapted for HTS applications during recent years.…”

Comet Assay on Toxicogenetics; Several Studies in Recent Years on Several Genotoxicological Agents

“…Lysis allaow the removing membranes, releases the soluble components of the cell, strips histones from DNA, and sheets of compact structures that are nucleoids wherein the DNA is attached at intervals to the nuclear matrix [13].…”

“…After treatment with lysis buffer, slides were incubated for 20 min in a freshly prepared alkaline buffer (300 mM NaOH, 1 mM EDTA; pH>13) [12], before DNA was submitted to electrophoresis (20 min; 25 V; 0.90 V/cm; 300 mA) [14]. Under electrophoresis, DNA is attracted to the anode, only the loops containing breaks, relaxing supercoiling, able to significantly expand [13].…”

Quantifying Nanoparticle Cellular Uptake: Which Method is Best?

“…Various cutting-edge techniques have emerged during the last years which can quantify the NPs based on their distinctive characteristics, for example -chemical composition, optical properties, magnetic properties or electron density. Comprehensive reviews and in-depth discussions are readily available [1][2][3][4] and are not reiterated herein. For the purpose of this commentary, we have selected some of the most widely used analytical techniques in the nanoscientific biomedical community based on a review of the existing nanotoxicology studies published within the scope of the last 5 years.…”