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The abacus is a counting frame designed to facilitate the process of arithmetic calculation.
The surprising creative re-working, occurs in Luci's action of using the felt bead necklace like a counting frame or abacus.
A counting frame (1612 µm2) was placed randomly on the first counting area and systemically moved through all counting areas until the entire delineated area was sampled.
A counting frame of 10.000 µm2 (Region of Interest; ROI) was superimposed on the calibrated images and cells were counted within this area using the image processing and analysis program ImageTool (UTHSCSA, USA) as described before [73] [75].
Fiber intersects were counted using unbiased, systematic sampling using a 60× objective with a counting frame size of 50 µm×50 µm, grid size of 130 µm×200 µm, and virtual plane separation of 7.0 µm [30].
NeuN-positive cells were counted in all cornu ammonis (CA) fields and dentate gyrus (DG) in 2 sections through the anterior hippocampus and Iba-1 positive cells in the right hemisphere in 4 sections, using stereological principles (Stereo investigator 7, System Inc, Magdeburg, Germany), with a counting frame of 40×40 µm per section for NeuN, and 150×150 µm per section for Iba-1.
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The numerical densities were determined: (1) Densities of DAPI-labeled cell nuclei (readout for the cellular density) were determined by counting the numbers of DAPI-stained cell nuclei which were within a counting-frame of 10.000 µm2 (ROI).
Cell profiles were counted at a final screen magnification of × 1024 using a 2D counting frame and a simple, unbiased counting rule (Gundersen, 1977).
Using this random and systematic sampling technique with an unbiased counting frame, a high level of reproducibility was found, as has also been demonstrated by others (Gundersen et al, 1988; Hansen et al, 1998).
We therefore used the SMI32 staining pattern to identify the boundaries between the different regions of the EG. Estimates of the number of fibres in each region were made using the optical fractionator (Stereo Investigator software) by scanning every second or third AChE section using a square grid (0.09 mm), oriented randomly, with a square counting frame of 400 μm.
In general, we used a 40×40 µm counting frame, a 1 µm guard, 100×100 µm sampling grid, and a dissector height of 8 µm.
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