Towards many colors in FISH on 3D-preserved interphase nuclei

The article reviews the existing methods of multicolor FISH on
nuclear targets, first of all, interphase chromosomes. FISH proper
and image acquisition are considered as two related components of a
single process. We discuss (1) M-FISH (combinatorial labeling +
deconvolution + widefield microscopy); (2) multicolor labeling +
SIM (structured illumination microscopy); (3) the standard approach
to multicolor FISH + CLSM (confocal laser scanning microscopy; one
fluorochrome - one color channel); (4) combinatorial labeling +
CLSM; (5) non-combinatorial labeling + CLSM + linear unmixing. Two
related issues, deconvolution of images acquired with CLSM and
correction of data for chromatic Z-shift, are also discussed. All
methods are illustrated with practical examples. Finally, several
rules of thumb helping to choose an optimal labeling + microscopy
combination for the planned experiment are suggested. Copyright (c)
2006 S. Karger AG, Basel.