Specific nuclear domains are nonrandomly positioned within the nuclear space, 
and this preferential positioning has been shown to play an important role in 
genome activity and stability. Well-known examples include the organization of 
repetitive DNA in telomere clusters or in the chromocenter of Drosophila and 
mammalian cells, which may provide a means to control the availability of 
general repressors, such as the heterochromatin protein 1 (HP1). We have 
specifically characterized the intranuclear positioning of in vivo fluorescence 
of the Caenorhabditis elegans HP1 homologue HPL-2 as a marker for 
heterochromatin domains in developing embryos. For this purpose, the wavelet 
transform modulus maxima (WTMM) segmentation method was generalized and adapted 
to segment the small embryonic cell nuclei in three dimensions. The 
implementation of a radial distribution algorithm revealed a preferential 
perinuclear positioning of HPL-2 fluorescence in wild-type embryos compared with 
the diffuse and homogeneous nuclear fluorescence observed in the lin-13 mutants. 
For all other genotypes analyzed, the quantitative analysis highlighted various 
degrees of preferential HPL-2 positioning at the nuclear periphery, which 
directly correlates with the number of HPL-2 foci previously counted on 2D 
projections. Using a probabilistic 3D cell nuclear model, we found that any two 
nuclei having the same number of foci, but with a different 3D probabilistic 
positioning scheme, can have significantly different counts in the 2D maximum 
projection, thus showing the deceptive limitations of using techniques of 2D 
maximum projection foci counts. By this approach, a strong perinuclear 
positioning of HPL-2 foci was brought into light upon inactivation of conserved 
chromatin-associated proteins, including the HAT cofactor TRAPP.