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Boundary distance (BD) plotting is a technique for making orientationinvariant comparisons of the spatial distribution of biochemical markers withinand across cells/nuclei. Marker expression is aggregated over points with thesame distance from the boundary. We present a suite of tools for improved dataanalysis and statistical inference using BD plotting. BD is computed using theEuclidean distance transform after presmoothing and oversampling of nuclearboundaries. Marker distribution profiles are averaged using smoothing withlinearly decreasing bandwidth. Average expression curves are scaled andregistered by x-axis dilation to compensate for uneven lighting and errors innuclear boundary marking. Penalized discriminant analysis is used tocharacterize the quality of separation between average marker distributions. Anadaptive piecewise linear model is used to compare expression gradients inintra, peri and extra nuclear zones. The techniques are illustrated by thefollowing: (a) a two sample problem involving a pair of voltage gated calciumchannels (Cav1.2 and AB70) marked in different cells; (b) a paired sampleproblem of calcium channels (Y1F4 and RyR1) marked in the same cell.

Analysis of spatial distribution of marker expression in cells using boundary distance plots