Angiogenesis: quantitative assessment by the chick chorioallantoic membrane assay.

The aim of the present work was to improve quantitative assessment of angiogenesis by chick chorioallantoic membrane (CAM) assay based on measurement of DNA synthesis. Following incubation of [3H]thymidine (3H-T) with CAM in vivo, incorporation of 3H-T to DNA fraction was expressed as percent of total 3H-T present in the initial homogenate of CAM, regardless of CAM weight and full recovery of applied radioactivity. The assay required simple, partial isolation of DNA to remove traces of free or unspecifically bound 3H-T. These modifications gave a reproducible assay with adequate precision (10-20%). DNA content of CAM did not change between 10-15 days of embryo development and growth of CAM was completed on day 11. 10 to 12-day old CAMs were used for evaluation of the assay. Using a qualitative approach (visual scoring), tissues of several tumors were implanted on CAM. Extracts of tumors that produced the highest score stimulated DNA synthesis in CAM. A similar effect was induced by EGF while cytostatics inhibited DNA synthesis in CAM. Since stimulation of angiogenesis is not a cell type-specific phenomenon, the assay in the present modification should aid the studies on angiogenic factors. With the help of this assay the angiogenic activity of adenocarcinoma of human endometrium was described.