Verification of the disector method for counting neurons, with comments on the empirical method

The empirical and disector methods are unbiased sampling methods for determining numbers of neurons. The present study verifies and thus calibrates these methods by determining true numbers of ganglion cells in serial reconstructions and then using each method to estimate the same populations. The empirical method gives accurate counts but is laborious (inefficient). Five separate disector analyses, distinguished by height (h), were done for each ganglion. The findings are: (1) that the estimates are consistently low when h is minimal (reference and look‐up sections are adjacent), but (2) the estimates are accurate when h is greater (one to four sections intervene between reference and look‐up sections). We ascribe the difficulties with the first disector to “lost” or “invisible” caps. We emphasize that we would not have known of the problem unless we verified our counts. If there is suspicion that difficulties with profile recognition might occur, we recommend that serial sections of an appropriately chosen sample of tissue be prepared and 500–1,000 neurons (or, more generally, particles) be reconstructed. Then the method of choice can be used on the issue of choice to make certain of the necessary accuracy before proceeding with the main study.