The extensive circulation of Highly Pathogenic (HP) H5N1 Avian Influenza in Egypt in poultry since 2006 resulted in the emergence of distinct clades with the recent identification of a further clade: 2.2.1. 1. The aim of this study was to characterize for the first time the antigenic profile of an extensive collection of genetically diverse Egyptian H5N1 HP viruses isolated between 2007 and 2010 applying antigenic cartography and principal component analysis to serological data. We identified that Egyptian H5N1 viruses have undergone significant antigenic diversification between 2007 and 2010 and two distinct antigenic clusters co-circulated in 2010. Such clusters correlated with 2.2.1 and 184.108.40.206 clades, showing for the first time that the new emerging clade 220.127.116.11 is antigenically distinct. This study highlights that the antigenic diversity of H5N1 HP Egyptian viruses may represent a potential challenge for the development of an effective vaccination programme for animal and human health in Egypt.