ABSTRACT
DNA replication, similar to other cellular processes, occurs within dynamic macromolecular structures. Any comprehensive understanding ultimately requires quantitative data to establish and test models of genome duplication. We used two different super-resolution light microscopy techniques to directly measure and compare the size and numbers of replication foci in mammalian cells. This analysis showed that replication foci vary in size from 210 nm down to 40 nm. Remarkably, spatially modulated illumination (SMI) and 3D-structured illumination microscopy (3D-SIM) both showed an average size of 125 nm that was conserved throughout S-phase and independent of the labeling method, suggesting a basic unit of genome duplication. Interestingly, the improved optical 3D resolution identified 3- to 5-fold more distinct replication foci than previously reported. These results show that optical nanoscopy techniques enable accurate measurements of cellular structures at a level previously achieved only by electron microscopy and highlight the possibility of high-throughput, multispectral 3D analyses.
Subject(s)
DNA Replication , Microscopy/methods , Animals , Bromodeoxyuridine/analysis , Cell Line , Cell Nucleus Structures/ultrastructure , Image Processing, Computer-Assisted , Mice , Microscopy, Confocal , Proliferating Cell Nuclear Antigen/analysisABSTRACT
We analyze polarization switching in vertical-cavity surface-emitting lasers, taking into account a proper semiconductor frequency-dependent complex susceptibility and spin-flip processes. Thermal effects are included as a varying detuning, and gain differences arise from birefringence splitting. We find that, for large birefringence, gain differences between the two linearly polarized modes are preponderant, and switching occurs owing to thermal shift. For small birefringence polarization switching from the high- to the low-gain mode occurs owing to the combined effect of birefringence and semiconductor phase-amplitude-coupled dynamics for a finite value of the carrier spin-flip rate.
