Virus-specific, early appearing neutralizing activities (ENA) and interferon (IFN) were detected in tears collected from patients during epidemics of acute hemorrhagic conjunctivitis (AHC). In one study, ENA that neutralized enterovirus type 70 (EV70) was detected in tears collected from 114 of 130 AHC patients in Florida. In another study, ENA against coxsackievirus type A24 (CA24) was detected in tears collected from 39 of 57 patients in Singapore with AHC caused by CA24. No tear samples contained ENAs to both EV70 and CA24. Tear samples from uninfected eyes did not contain ENA to EV70 or CA24. ENA to EV70 was detected in 6 of 11 patients l-6h before the onset of AHC. In addition, tears of 68% of patients seen on the day of onset produced tears that contained ENA to EV70. Thus, ENA to EV70 may be detected less than 24h after infection (based on 24h incubation period). IFNβwas detected in 30% of tear samples collected from patients on the day of onset of AHC caused by EV70. This finding suggested that ENA and IFN could act together to inhibit primary infections of AHC. It was found that the combination of ENA and IFN inhibited virus replication synergistically (≥300 fold reduction) in preinfected cells. Our findings suggest that ENA represents a previously unreported early defense mechanism of the eye, that endogenous ENA and endogenous IFN could inhibit viruses synergistically in vivo, and that ENA in tears could be useful in identifying the agent causing AHC.