Interference: A Much-Neglected Aspect in High-Throughput Screening of Nanoparticles

Area of Science:

• Nanotechnology
• Toxicology
• Biomedical Engineering

Background:

• Nanoparticle (NP) interference with conventional toxicity assays is a known issue, yet researchers continue to rely on these methods for high-throughput screening (HTS).
• Existing approaches often fail to address all NP interference mechanisms, which are assay-dependent.
• Intracellular NP interference with fluorescent dyes, crucial for HTS, has been largely neglected.

Purpose of the Study:

• To investigate the interference mechanisms of gold and silver NPs in HTS assay systems.
• To assess optical interference, adsorption, and catalysis as NP interference modes.
• To evaluate NP quenching of common fluorescent dyes used in HTS.

Main Methods:

• Investigated gold and silver NPs using absorbance (XTT assay), fluorescence (CytoTox-ONE), and luminescence (CellTiter Glo) assays.
• Assessed NP quenching effects on fluorescent dyes like 2",7"-dichlorofluorescein, propidium iodide, and TC-BI.
• Examined interference mechanisms including optical interference, adsorption, and catalysis.

Main Results:

• Gold and silver NPs demonstrated interference in absorbance, fluorescence, and luminescence-based HTS assays.
• Significant quenching of fluorescent dyes by NPs was observed.
• Different interference mechanisms were identified depending on the assay system and NP type.

Conclusions:

• Ignoring NP interference in HTS can compromise the reliability of big data for predictive toxicology.
• Comprehensive assessment of all NP interference mechanisms is mandatory for HTS.
• Validation of results using label-free methodologies is essential for accurate predictive toxicology.