Developmental patterns of gene expression were determined following intravascular administration of adenovirus in utero, during sequential stages of murine development. Replication-deficient adenovirus (AdCMV.LacZ) was injected into yolk sac vessels of mouse embryos 12, 13, 15 and 18 days post-conception (d.p.c.). beta-Galactosidase (beta-gal) expression was evaluated 24-48 h after injection, at birth, and 5 weeks following normal delivery. Gene expression was detected in myocardial cells, endothelial cells of heart, lung, kidney, adrenal, gut, and in hepatocytes. The patterns of expression were distinct for each stage of virus administration and time-point of analysis. Intensity of individual organ expression varied with injection time-point, with the largest number of organs express- ing the transgene when embryos were injected at 15 d.p.c. beta-Gal activity was detected in only a subset of cells expressing the murine coxsackievirus and adenovirus receptor (CAR), indicating factors other than receptor distribution were responsible for the pattern of transgene expression observed. These studies begin to define critical parameters affecting intravascular gene delivery in utero and indicate that intrinsic developmental regulatory mechanisms may control exogenous gene expression. Intravenous administration of adenovirus provides a unique approach for in utero gene transduction and will be a useful adjunct in evaluating genes which have early lethal mutations.