Posterior capsule opacification is the main complication of cataract surgery. Using adenovirus-mediated gene transfer, we recently reported that it was feasible to prevent PCO by overexpressing pro-apoptotic molecules such as pro-caspase 3 or Bax in the residual lens epithelial cells post-cataract surgery. However, this approach is feasible only if gene transfer can be restricted to the residual cells responsible for PCO. Initially, we tested an adenovirus (human serotype 5, HAd5), a lentivirus (HIV) and an oncoretrovirus (MLV) vector for the their in vivo transduction efficiency of rabbit lens cells. We found that HAd5 vectors were the most efficient (>90% of the cells could be transduced). Six potential lens-specific promoters were then cloned into HAd5 vectors and assayed for their ability to target expression to a specific population of cells, using in vitro, ex vivo and in vivo rabbit tissues and human lens capsular bags. We found that the LEP503, MIP and Filensin promoters induced strong lens-specific expression of a reporter gene, in human lens cells. Following this ex vivo assay, we showed in a rabbit PCO model that gene transfer could be spatially restricted to the capsular bag by confining the vector with Matrigel. Our combined approach using a lens-specific promoter and a biocompatible gel should render feasible a novel therapeutic strategy for PCO that targets the remaining lens cells.