This illustrates the need for a new vaccine concept that maintains high Mepiroxol protective efficacy regardless of the antigenic variations that arise frequently due to antigenic drift. Chemically and UV inactivated influenza virus preparations rely exclusively on antibody responses for protection and do not induce cytotoxic T cell responses. The Tc cell response to influenza is broadly cross-reactive between virus strains and is important in the recovery from primary infections. We have previously reported that c-Flu preparations can induce crossreactive Tc cell responses. Gamma-irradiation is the preferred method of inactivation of highly infectious agents for biochemical analysis, including Ebola, Marburg and Lassa viruses. It inactivates virus infectivity by generating strand-breaks in the genetic material and has the further advantage, compared with chemical agents, of high penetration into and through biological materials. In contrast to chemical treatment with Atropine sulfate formalin or b-propiolactone, which induces cross-linking of proteins, c-rays have little impact on the antigenic structure and biological integrity of proteins. The Manual on Radiation Sterilization of Medical and Biological Material of the International Atomic Energy Agency indicates that exposure to 0.65 kGy of c-rays causes a total loss of influenza virus infectivity, but disrupting the haemagglutinating activity requires an exposure to higher than 200 kGy. The reduced impact of c-irradiation on the antigenic structure of viral particles is therefore also expected to improve the magnitude and/or quality of humoral immunity elicited in vaccine recipients over that obtained by using present vaccine preparations. We have been investigating the ability of c-Flu preparations to induce heterotypic and cross-protective immunity against avian H5N1 influenza A virus. Our data clearly show that a single intranasal administration of c-A/PR8 protects mice against lethal H5N1 and other heterotypic infections. Human influenza A viruses bind to a2,6-linked sialic acid receptors expressed on epithelial cells of the upper respiratory tract of humans, whereas avian influenza H5N1 viruses bind to a2,3linked receptors expressed predominantly in the lower respiratory tract In general, binding of virus to a2,6-linked receptors is associated with low virulence but high transmissibility. In contrast, virus binding to a2,3-linked receptors is associated with low transmissibility but high virulence and often lethal influenza virus infections. Therefore, an avian influenza pandemic might be expected to be associated with a mutation in the haemagglutinin molecule that would allow H5N1 virus to bind to a2,6-linked receptors. Subunit vaccines based on currently identified H5 molecules cannot account for the antigenic variants that would result following such mutations and, consequently, neutralizing antibody-mediated protection induced by these vaccines would likely be limited. Tc cell responses to influenza infection, however, are mainly directed against internal viral proteins, which are highly conserved among all influenza A virus strains. Mutations of internal genes are not susceptible to antibodymediated selection and viral escape from Tc cell responses is curtailed by MHC class I polymorphism. Therefore, vaccines inducing Tc cell responses against the highly conserved internal proteins, such as the nucleoprotein, are expected to provide crossprotection against different influenza A virus strains. Our data indicate that the cross-reactive Tc cell responses induced by c-Flu are predominantly directed against the internal proteins, indicated by lysis of NPP-labeled targets.