While superresolution optical microscopy techniques afford enhanced resolution for biological applications, they have largely been used to study structures in isolated cells. We use the FDTD method to simulate the propagation of focused beams for STED microscopy through multiple biological cells. We model depletion beams that provide 2D and 3D confinement of the fluorescence spot and assess the effective PSF of the system as a function of focal depth. We compare the relative size of the STED effective PSF under one- and two-photon excitation. PSF calculations suggest that imaging is possible up to the maximum simulation depth if the fluorescence emission remains detectable.
A linear coherent superposition method for estimating the plane wave far-field scattering pattern from multiple biological cells computed by the finite-difference time-domain (FDTD) method is presented. The method enables the FDTD simulation results of scattering from a small number of complex scatterers, such as biological cells, to be used to estimate the far-field pattern from a large group of those same scatterers. The superposition method can be used to reduce the computational cost of FDTD simulations by enabling a single large scattering problem to be broken into smaller problems with more practical computational requirements. It is found that the method works best in cases where there is little multiple scattering interaction between adjacent cells, so the far-field pattern of multicell geometry can simply be calculated as a phase-adjusted linear superposition of the scattering from individual cells. A strategy is also presented for choosing the minimum number of cells in cases with significant multiple scattering interactions between cells.
A linear superposition method for estimating the plane wave farfield scattering pattern from multiple biological
cells computed by the finite difference time domain (FDTD) method is presented. The method allows the FDTD
simulation results of scattering from a small number of complex scatterers, such as biological cells, to be used
to estimate the farfield pattern from a large group of those same scatterers. This method can be used to reduce
the computational cost of FDTD simulations by allowing a single large scattering problem to be broken into
smaller problems with more practical computational requirements. It has been found that the method works
best in cases where there is little multiple scattering interaction between adjacent cells, so the farfield pattern
of multicell geometry can simply be calculated as a phase-adjusted linear superposition of the scattering from
individual cells. A strategy is also presented for choosing the minimum number of cells in cases with significant
multiple scattering interactions between cells.
The finite difference time domain method was used to compute scattering of a focused optical beam by multiple
heterogeneous biological cells. A perfectly matched layer boundary condition and the scattered-field-only method
were utilized in the simulation to increase accuracy and computational efficiency. A fifth-order approximation
to the focused Gaussian beam was used for the incident field. A parametric study was performed to determine
scattering effects of varying cellular fine structure, such as nuclear refractive index, organelle volume density,
cellular shape and the cell membrane on the point spread function of the beam. It was found that two-photon
PSF is largely unaffected by increasing numbers of scatterers within cells, while two-photon excitation signal
strength is dependent on both beam focal depth and the density of scatterers in tissue.
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