Determination of cell quantity in three-dimensional culture systems, such as the fibroblast-populated collagen matrix, can be difficult secondary to the abundance of extracellular collagen. The WST-1 assay, which can quantitate cellular processes in monolayer culture, was tested as a measure of relative cell quantity in a three dimensional culture system, the fibroblast-populated collagen matrix. The assay had acceptable accuracy and precision, and was not affected by the extracellular matrix. It was concluded that the assay was a reasonable alternative to other measures of cell quantity in the fibroblast-populated collagen matrix.