The interaction between heparin, a polyanion, and a polycationic dendrimer with a glycine core and lysine branches Gly–Lys63(NH2)64 has been investigated. Complexation was assessed by transmission electron microscopy, size and zeta potential measurements, methylene blue spectroscopy, and measuring the anti-coagulant activity of heparin in vitro and in vivo. Complete association between the heparin and the dendrimer occurred a 1:1 mass ratio (2:1 molar ratio or +/−charge ratio) with formation of quasi-spherical complexes in the size range of 99–147 nm with a negative zeta potential (−47 mV). Heparin–dendrimer (dendriplex) formation led to a concentration-dependent neutralization of the anticoagulant activity of heparin in human plasma in vitro, with complete loss of activity at a 1:1 mass ratio. The anticoagulant activity of the dendriplexes in Sprague-Dawley rats was also evaluated after subcutaneous administration with uncomplexed heparin as a comparator. The in vivo anticoagulant activity of heparin in plasma, evaluated using an antifactor Xa assay, was abolished after complexation. Measurement of [3H]-heparin showed that both free heparin and dendriplexes were present in plasma and in organs. Such data conﬁrmed stably the formation of dendriplexes, which could be essential in developing novel dendrimer-based anti-angiogenic therapeutics suitable in combinatory therapeutics and theranostics.