A simple, inexpensive and multi-scale 3-D fluorescent test sample for optical sectioning microscopies
Fluorescence standards allow for quality control and for the comparison of data sets across instruments and laboratories in applications of quantitative fluorescence. For example, users of microscopy core facilities expect a homogenous and time-invariant illumination and a uniform detection sensitivity, which are prerequisites for quantitative imaging analysis, particle tracking or fluorometric pH or Ca2+-concentration measurements. Similarly, confirming the three-dimensional (3-D) resolution of optical sectioning micro-scopes prior to volumetric reconstructions calls for a regular calibration with a standardised point source. Typically, the test samples required for such calibration measurements are different ones, and they depend much on the very microscope technique used. Also, the ever-increasing choice among these techniques increases the demand for comparison and metrology across instruments. Here, we advocate and demonstrate the multiple uses of a surprisingly versatile and simple 3-D test sample that can complement existing and much more expensive calibration samples: simple commercial tissue paper labelled with a fluorescent highlighter pen. We provide relevant sample characteristics and show examples ranging from the sub-µm to cm scale, acquired on epifluorescence, confocal, image scanning, two-photon (2P) and light-sheet microscopes.
A simple, inexpensive and multi-scale 3-D fluorescent test sample for optical sectioning microscopies.
Olevsko I, Szederkenyi K, Corridon J, Au A, Delhomme B, Bastien T, Fernandes J, Yip C, Oheim M, Salomon A : Microsc Res Tech, 2021