In view of their promising potential we have evaluated recombinant soluble HA trimers in chickens and mice for their ability to induce protective immunity against infection with HPAI H5N1. To this end, recombinant soluble H5 proteins provided with a GCN4 trimerization motif and a STREP-tag II, the latter for purification purposes, were expressed in mammalian cells. The recombinant soluble H5 trimers (sH5 3 ) were purified from the culture supernatants using a simple one-step purification protocol and characterized with respect to their oligomeric state and bioactivity. Subsequently, vaccination with the sH5 3 preparation was shown to provide complete protection against challenge with HPAI H5N1 both in mice and in chickens, in the latter already after a single immunization.