Recombinant adenovirus (Ad) vectors can accomplish efficient in vivo gene transfer and thus are important in the context of a variety of gene therapy approaches. The cellular receptor for the Ad fiber knob is prevalent on a number of normal tissues which undermines the targeting of Ad to specific tumor cells. Therefore, the ablation of native Ad tropism and the introduction of novel Ad tropism are both necessary to target Ad vectors specifically to tumors. In this study, we have developed a flexible method for cross-linking the Fab fragment of a neutralizing anti-knob monoclonal antibody (1D6.14) to a cell receptor ligand. The cross-linking moieties are complementary low molecular weight recognition units, similar in concept to the avidin-biotin system. For proof of concept, we cross-linked 1D6.14 Fab to the basic fibroblast growth factor (FGF2). The Fab and FGF2 conjugates were synthesized and characterized both structurally and functionally. The conjugates were then complexed with an adenovirus vector carrying firefly luciferase (AdCMVLuc) and the resulting complex used to show infection of a number of tumor cell lines expressing FGF receptors. This cross-linking system should provide a rapid and convenient method of conjugating various ligands to the Fab fragment for targeting Ad vectors to different types of tumors.