The Quantitation of Cells: Optical Enumeration of Cells

Fundamental to any quantitative study of the kinetics of cell population growth and to the accurate mea­surement of changes in the cell’s physical or chemical properties are methods for determining either the mass or the number of cells present in the population.

A variety of techniques are available to do this; among those that are easily used are estimations of cell mass based on the total fresh (i.e., wet) weight or dry weight of the sample and estimation of cell numbers from measurements of the turbidity of the cell suspen­sion.

However, these methods lack the level of accu­racy that is generally considered necessary for quanti­tative studies.

More frequently, the precise number of cells present in the sample is determined by the direct optical examination of an aliquot of cell suspension drawn from the culture, by electronic gating, or by flow cytometry. Although the latter two procedures are far more accurate than weight determinations, turbidity methods, or optical enumeration, they re­quire that the cells occur separately and, therefore, cannot be applied directly to tissues.

Optical Enumeration of Cells:

The direct microscopic enumeration of cells requires a glass counting chamber (often referred to as a hema­cytometer because of its historical and widespread use for counting various kinds of blood cells). This de­vice consists of a thick glass microscope slide, the sur­face of which has been etched with a number of squares of known dimensions.

A drop of cell suspen­sion is placed between the counting surface and an overlying cover slip, and because the distance between the undersurface of the cover slip and the surface of the counting area is known, the total number of cell present in a known volume of sample can be determined.

Then, by extrapolation, the number of cells in the entire cell culture (or in any portion of it) can eas­ily be calculated. The enumeration of cells by direct optical examination is tedious but extremely useful, especially if the suspensions contain different types of cells (as in the case of blood) that may be distinguished visually. Hemacytometric methods are still used in many laboratories.