title: Spatiotemporal Rank Filtering Improves Image Quality Compared to Frame Averaging in 2-Photon Laser Scanning Microscopy.
abstract: Live imaging of biological specimens using optical microscopy is limited by tradeoffs between spatial and temporal resolution, depth into intact samples, and phototoxicity. Two-photon laser scanning microscopy (2P-LSM), the gold standard for imaging turbid samples in vivo, has conventionally constructed images with sufficient signal-to-noise ratio (SNR) generated by sequential raster scans of the focal plane and temporal integration of the collected signals. Here, we describe spatiotemporal rank filtering, a nonlinear alternative to temporal integration, which makes more efficient use of collected photons by selectively reducing noise in 2P-LSM images during acquisition. This results in much higher SNR while preserving image edges and fine details. Practically, this allows for at least a four fold decrease in collection times, a substantial improvement for time-course imaging in biological systems.
authors.affiliation: Department of Pathology, University of California San Francisco, San Francisco, California, United States of America.|Biological Imaging Development Center, University of California San Francisco, San Francisco, California, United States of America.|Computational Biology Graduate Group, University of California Berkeley, Berkeley, California, United States of America.
authors.forename: Henry
authors.initials: H
authors.lastname: Pinkard
authors.affiliation: Department of Pathology, University of California San Francisco, San Francisco, California, United States of America.|Biological Imaging Development Center, University of California San Francisco, San Francisco, California, United States of America.
authors.forename: Kaitlin
authors.initials: K
authors.lastname: Corbin
authors.affiliation: Department of Pathology, University of California San Francisco, San Francisco, California, United States of America.|Biological Imaging Development Center, University of California San Francisco, San Francisco, California, United States of America.
authors.forename: Matthew F
authors.initials: MF
authors.lastname: Krummel
mesh_terms: D019264:Adoptive Transfer; D000818:Animals; D018414:CD8-Positive T-Lymphocytes / Q000166:cytology*; D007091:Image Processing, Computer-Assisted / Q000706:statistics & numerical data*; D008198:Lymph Nodes / Q000166:cytology*; D051379:Mice; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal / Q000295:instrumentation / Q000379:methods*; D017785:Photons; D059629:Signal-To-Noise Ratio; D013997:Time Factors