WHAT WAS WRONG WITH THE ABERCROMBIE AND EMPIRICAL CELL COUNTING METHODS - A REVIEW

When a tissue volume is sectioned, cells or other objects are cut into segments by the sectioning process. The Abercrombie and empirical met hods count object segments in histological sections and then apply a c orrection formula to convert the segment count to object number There has been considerable recent controversy over whether these methods sh ould be abandoned (in favor of the disector). Although both methods ap pear unbiased as thought experiments on paper, regardless of variation in object size, shape, or orientation, in practice two problems are i nherent in the segment-counting approach: the practical problem of los t caps and a conceptual flaw that becomes apparent only when a need fo r unbiased estimation of certain factors in the correction formulae is seriously addressed. The Abercrombie method is inevitably biased by l ost caps, whereas in the empirical method, this potential bias can be avoided. In the Abercrombie formula, the relevant factor to be estimat ed (aside from section thickness) is H, mean object height in the axis perpendicular to the section plane, and in the empirical method, it i s the ratio of segment number to object number. In both methods, the f actor in question should be estimated from an unbiased sample of the t otal population of objects. But unbiased selection of a statistically adequate number of objects for this estimation constitutes an unbiased , statistically adequate count. Once this is done, there is no reason to complete the steps for estimation of the factor; the count in this specimen is finished. It is shown that the empirical method's serial s ection procedure can be used to estimate H. This estimate is more sens itive to lost caps than the Abercrombie equation, but, when the formul a for H is substituted into the Abercrombie equation, the lost caps er ror disappears. However, this approach is useless, as making this subs titution transforms the Abercrombie equation into the empirical method equation. (C) 1998 Wiley-Liss, Inc.