3D morphological analysis of the mouse cerebral vasculature: Comparison of in vivo and ex vivo methods
Fig 2
FWHM of signal along optical axis and x-axis versus depth for beads embedded in agar.
The beads were 0.5 μm diameter yellow-green fluorescent beads (excitation peak 505 nm; emission peak 515 nm) and were embedded in fructose-cleared 1% low melting point agar. Imaging was performed using 2PFM at an excitation wavelength of 800 nm. The FWHM was calculated by fitting a Gaussian to the signal profile along either the optical or x-axis for these beads. Prior to fitting the Gaussian, the image of the beads was blurred by a Gaussian with FWHM 1.5 μm, as per the vascular images on which vessel tracking was performed. Since the slope of the x-axis was not statistically different from 0 (p = 0.8136), only the PSF along the optical axis was assumed to change with depth when performing vessel tracking. The ribbons surrounding the straight lines represent the 95% confidence interval.