Estimation of subcellular organelle volume from ultrathin sections through centrioles with a discretized version of the vertical rotator

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Abstract

Summary

A method for the fast and efficient estimation of the volume (but not surface area) of subcellular organelles is presented. It consists of a rotator/coaxial-section approach based on the Pappus theorem and represents a discretized version of the vertical rotator where, instead of measuring intercept lengths, the points in distance classes are counted. Centrioles serve as a unique reference 'double-point' with constant size allowing unbiased cell selection from the whole population with equal probability and without the disector application. The sandwich-like method of sample preparation allows comparison of control and experimental cases with the same errors caused by overlapping and overprojection. Test experiments demonstrated that the vertical discretized rotator was an efficient and precise tool for the estimation of volume and that a few independent sections of unknown thickness were sufficient for the quantification of one experimental point.

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