Visualization of cellular aggregates cultured on a three dimensional collagen sponge matrix

A one-step vital stain is described for the macroscopic visualization of histotypic cell aggregates in fetal rat lung organotypic cultures. Organotypic cultures are incubated in 0.05%-0.1% 2,3,5'-triphenyl tetrazolium chloride (TTC) in culture medium (37 degrees C). Living cells reduce the tetrazole to a water-insoluble red colored formazan. Cell aggregates appear as densely stained foci against the lighter background of the Gelfoam substrate. Stained cultures may be scanned macroscopically to determine the degree of reaggregation and assess cell viability. Identification of aggregates by TTC staining improves the efficiency of tissue processing for electron microscopy and does not alter the ultrastructural appearance of the cultured cells.