Three-Dimensional Maps of All Chromosomes in Human Male Fibroblast Nuclei and Prometaphase Rosettes
Figure 1
24-Color 3D FISH Representation and Classification of Chromosomes in a Human G0 Fibroblast Nucleus
(A) A deconvoluted mid-plane nuclear section recorded by wide-field microscopy in eight channels: one channel for DAPI (DNA counterstain) and seven channels for the following fluorochromes: diethylaminocoumarin (Deac), Spectrum Green (SG), and the cyanine dyes Cy3, Cy3.5, Cy5, Cy5.5, and Cy7. Each channel represents the painting of a CT subset with the respective fluorochrome. The combinatorial labeling scheme is described in Materials and Methods. RGB images of the 24 differently labeled chromosome types (1–22, X, and Y) were produced by superposition of the seven channels (bottom right).
(B) False color representation of all CTs visible in this mid-section after classification with the program goldFISH.
(C) 3D reconstruction of the complete CT arrangement in the nucleus viewed from different angles.
(D) Simulation of a human fibroblast model nucleus according to the SCD model (see Materials and Methods). The first image shows 46 statistically placed rods representing the 46 human chromatids. The next three images simulate the decondensation process and show the resulting CT arrangement obtained after different numbers of Monte Carlo relaxation steps (200, 1,000, and 400,000). This set of figures is taken from Video S1.