Protocol for immunodot blot detection of UVB photoproducts in extracellular DNA

Abstract

Summary: UV radiation induces the formation of adducts in genomic DNA within cells that are later found to be present in cell-free fractions associated with extracellular vesicles (EVs) outside of cells. Here, we present a protocol for isolating UV photoproducts in extracellular DNA released from UVB-irradiated cells via differential centrifugation. We then detail steps for monitoring the DNA adducts using DNA immunoblotting. This protocol can be applied for detection of DNA adducts in EVs from cell culture and skin explant models.For complete details on the use and execution of this protocol, please refer to Carpenter et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Cell culture
• Cell isolation
• Cell-based Assays
• Cell separation/fractionation
• Antibody
• Signal Transduction