A Model-based approach for microvasculature structure distortion correction in two-photon fluorescence microscopy images

This paper investigates a postprocessing approach to correct spatial distortion in two-photon fluorescence microscopy images for vascular network reconstruction. It is aimed at in vivo imaging of large field-of-view, deep-tissue studies of vascular structures. Based on simple geometric modelling of the object-of-interest, a distortion function is directly estimated from the image volume by deconvolution analysis. Such distortion function is then applied to subvolumes of the image stack to adaptively adjust for spatially varying distortion and reduce the image blurring through blind deconvolution. The proposed technique was first evaluated in phantom imaging of fluorescent microspheres that are comparable in size to the underlying capillary vascular structures. The effectiveness of restoring three-dimensional (3D) spherical geometry of the microspheres using the estimated distortion function was compared with empirically measured point-spread function. Next, the proposed approach was applied to in vivo vascular imaging of mouse skeletal muscle to reduce the image distortion of the capillary structures. We show that the proposed method effectively improve the image quality and reduce spatially varying distortion that occurs in large field-of-view deep-tissue vascular dataset. The proposed method will help in qualitative interpretation and quantitative analysis of vascular structures from fluorescence microscopy images. Lay Description Image distortion in fluorescence microscopy affects qualitative interpretation and particularly quantitative comparison of biological structures. Conventional image deconvolution approaches using empirically measured point-spread function (PSF) from submicron beads have limitations in correcting for large structural distortion in in vivo deep-tissue scans that require spatially varying models of the PSF. In this paper, we present a model-based image restoration approach to reduce spatial distortion and image blurring of microvascular structures in two-photon fluorescence microscopy images. Our results showed significant improvement in quantitative geometry measurements of the microsphere phantom compared to conventional measured PSF approach. We also showed that the method improve the image quality of vessel structures in mouse skeletal muscle. This image restoration approach can assist in qualitative and quantitative analysis of large field-of-view deep-tissue two photon fluorescence microscopy images.