Spatiotemporal Rank Filtering Improves Image Quality Compared to Frame Averaging in 2-Photon Laser Scanning Microscopy

PLoS One. 2016 Mar 3;11(3):e0150430. doi: 10.1371/journal.pone.0150430. eCollection 2016.

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adoptive Transfer
  • Animals
  • CD8-Positive T-Lymphocytes / cytology*
  • Image Processing, Computer-Assisted / statistics & numerical data*
  • Lymph Nodes / cytology*
  • Mice
  • Mice, Inbred C57BL
  • Microscopy, Confocal / instrumentation
  • Microscopy, Confocal / methods*
  • Photons*
  • Signal-To-Noise Ratio
  • Time Factors