Optical Filters and Light Sources for FISH

Brilliant fluorescence signals with almost no background and cross talk are the aim of FISH analysis in imaging systems. The precise selection of hardware components like optical filters and light sources plays a major role. Considering fluorescent dye characteristics is the base of configuring perfectly matched multicolor-FISH (mFISH) filters, which allow the simultaneous application of up to seven dyes. The spectral characteristics of filters are here explained with respect to microscope setups. Spectral cross talk, pixel-shift effects, and stable energy output will be the main issues in daily work. Specific hard-coated single-band filters with small bandwidth but maximum transmission avoid cross talk to a high degree; the use of multiband filters allows simultaneous imaging of up to four dyes; multiband systems with separate exciters and emitters in filter wheels can be controlled by software. The comparison of mercury and metal halide light sources to new light-emitting diode (LED)-based light sources is shown, which reveals the chance of stable and long-term light output of the new LED light sources. White-light LED sources are nowadays a perfect choice for replacing mercury-based lamps. The combination of hard-coated mFISH filters with stable LED light sources is a very helpful tool in daily work.

Source: Springer protocols feed by Cell Biology - October 20, 2016 Category: Cytology Source Type: news