Pinhole Closure Improves Spatial Resolution in Confocal Scanning Microscopy

Methods Mol Biol. 2021:2274:385-389. doi: 10.1007/978-1-0716-1258-3_31.

Abstract

Confocal microscopy is a simple, super-resolution technique, which does not produce a marked increase in resolution compared to other advanced techniques, such as super-resolution nanoscopy. Here, we present a simple protocol to acquire "slightly, but easily resolved" images by pinhole closure (<1 airy unit) in a conventional confocal scanning microscope equipped with an avalanche photodiode, a detector with high sensitivity. We use murine neuroblastoma Neuro2a cells to demonstrate the image resolution obtained via this protocol without the use of any special software to enhance image quality.

Keywords: Confocal microscopy; Fluorescence microscopy; Laser scanning microscopy; Live-cell imaging; Super-resolution microscopy.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Image Processing, Computer-Assisted / methods*
  • Mice
  • Microscopy, Confocal / instrumentation*
  • Microscopy, Confocal / methods
  • Microscopy, Fluorescence / instrumentation*
  • Microscopy, Fluorescence / methods
  • Molecular Imaging / methods*
  • Neuroblastoma / pathology*
  • Software*
  • Tumor Cells, Cultured