Spatially sculpted laser scissors for study of DNA damage and repair

Abstract

Abstract. We present a simple and efficient method for controlled linear induction of DNA damage in live cells. By passing a pulsed laser beam through a cylindrical lens prior to expansion, an elongated elliptical beam profile is created with the ability to expose controlled linear patterns while keeping the beam and the sample stationary. The length and orientation of the beam at the sample plane were reliably controlled by an adjustable aperture and rotation of the cylindrical lens, respectively. Localized immunostaining by the DNA double strand break ͑DSB͒ markers phosphorylated H2AX ͑␥H2AX͒ and Nbs1 in the nuclei of HeLa cells exposed to the "line scissors" was shown via confocal imaging. The line scissors method proved more efficient than the scanning mirror and scanning stage methods at induction of DNA DSB damage with the added benefit of having a greater potential for high throughput applications.

Light-induced injury in mouse embryos revealed by single-cell RNA sequencing

“…Light-induced injury in cells is often associated with development arrest, in which apoptosis is induced by damage to the chromosome [15–17]. Consistent with those findings, differential transcription levels, more cell apoptosis, and greater cell responses to DNA damage stimuli were found in embryos after light exposure (Fig.…”

Analysis of DNA double-strand break response and chromatin structure in mitosis using laser microirradiation

“…Therefore, a variety of approaches to induce DNA damage and study the subsequent response have been employed. One of the more recent and growing approaches to study DNA repair factor recruitment uses lasers to produce spatially defined DNA damage in interphase nuclei ( 1–20 ). These studies have used a wide variety of laser systems and dosimetry, often making it difficult to compare and interpret results ( 19 ).…”

Photothermal nanoblade for patterned cell membrane cutting

“…Optoporation [ 7 , 8 ] ablates a single hole in the cell membrane at the focal point of a tightly focused pulsed laser beam. Two dimensional patterned membrane poration can also be achieved by rapidly scanning the laser focus within a field of view [ 9 ] or by using beam-shaping optics [ 10 ].…”