GC-MS/MS Measurement of Nanomaterial Induced Genotoxicity in Isolated DNA
Bryant C. Nelson, Elijah J. Petersen, Pawel Jaruga, M Miral Dizdar
Making accurate measurements of the environmental fate and environmental and biological effects of engineered nanomaterials (ENMs) is critical for reliable risk assessment of these materials. ENM induced oxidative stress and/or direct binding of ENMs to DNA can result in genotoxicity. However, the unique behaviors of ENMs may cause measurement artifacts during the determination of their genotoxic potential. For example, results from several DNA damage studies using the traditional Comet assay have reported artifacts from the presence of ENMs. Potential explanations for this phenomenon are that ENMs may induce DNA damage during processing after the exposure period has concluded or that the ENMs may be in the Comet tail and mistaken for DNA. These types of measurement artifacts preclude the accurate measurement of DNA damage and prevents researchers from clarifying the fundamental mechanisms of ENM induced toxicity. In this document, we describe a protocol to quantitatively measure a range of DNA lesions using isotope-dilution gas chromatrography/tandem mass spectrometry (GC-MS/MS) with isotope-dilution. This approach utilizes NIST standard reference material (SRM) 2396 (Oxidative DNA Damage Mass Spectrometry Standards). This method circumvents many of the artifacts observed in the Comet assay in nanogenotoxicity tests by directly quantifying molecular level DNA damage (i.e., DNA lesion) for a range of oxidatively induced damage products rather than making a non-specific measurement of DNA damage (i.e., Comet tail length). Isotope-dilution mass spectrometry methods have been recently utilized by our laboratory to successfully measure DNA damage both in vitro and in vivo caused by gold nanoparticles, copper oxide nanoparticles, single-wall carbon nanotubes, iron oxide nanoparticles, titanium dioxide nanoparticles and silver nanoparticles.
, Petersen, E.
, Jaruga, P.
and , M.
GC-MS/MS Measurement of Nanomaterial Induced Genotoxicity in Isolated DNA, Special Publication (NIST SP), National Institute of Standards and Technology, Gaithersburg, MD, [online], https://doi.org/10.6028/NIST.SP.1200-19
(Accessed February 22, 2024)