Three‐dimensional counting

An accurate and direct method to estimate numbers of cells in sectioned material

Robert Williams, Pasko Rakic

Research output: Contribution to journalArticle

365 Citations (Scopus)

Abstract

We introduce a way to count and measure cells in an optically defined volume of tissue called a counting box. This method—direct three‐dimensional counting (3DC)—eliminates the need for correction factors, such as that introduced by Abercrombie (Anat. Rec. 94:239–247, '46), to determine the number of cells per unit volume (Nv). Problems caused by irregular cell shape and cell size, nonrandom orientation, and splitting of cells by the knife during sectioning are overcome. Furthermore, 3DC is insensitive to large variations in section thickness. The innovative feature of 3DC is the definition of a counting box with top and bottom sides located inside the section a precise distance away from the cut surfaces of the tissue. The positions of the top and bottom sides of the counting box are delimited by using a digital length gauge and a Z‐axis control unit. Sections of tissue between 8 and 100 μ thick are examined with a high numerical aperture objective in combination with video‐enhanced differential interference contrast optics (DIC). Cells are marked on a television screen while the microscopist scans systematically from the top to the bottom of the counting box. Cells that are located completely inside the box and cells that only cross through its top, right, or back sides are counted. All cells that cross the planes that define the bottom, left, and front sides of the counting box are not counted. Direct 3DC provides an accurate, simple, and reliable way to count cells, nuclei, nucleoli, or other objects in sectioned material. To build the counting system requires a standard transmitted light microscope equipped with DIC optics, a video camera, a position sensor, and a Z‐axis control unit.

Original languageEnglish (US)
Pages (from-to)344-352
Number of pages9
JournalJournal of Comparative Neurology
Volume278
Issue number3
DOIs
StatePublished - Jan 1 1988

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Cell Count
Cell Nucleolus
Cell Shape
Television
Cell Nucleus
Cell Size
Light

All Science Journal Classification (ASJC) codes

  • Neuroscience(all)

Cite this

Three‐dimensional counting : An accurate and direct method to estimate numbers of cells in sectioned material. / Williams, Robert; Rakic, Pasko.

In: Journal of Comparative Neurology, Vol. 278, No. 3, 01.01.1988, p. 344-352.

Research output: Contribution to journalArticle

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