Antiviral neutralizing antibody to Hantaan virus as determined by plaque reduction technique

The 76--118 strain of Hantaan virus was titrated in E6 (Vero) cells by the plaque method using agarose overlay medium. Visible plaques, formed 10 days post-infection, were uniformly 2--3 mm in diameter. Dose-response experiments showed that a single infectious particle initiated the formation of a plaque. Infectivity titers by the plaque method were equivalent to those obtained by the endpoint method (TCID50) using the immunofluorescence antibody technique (IFA) for antigen detection. The single-cycle growth pattern of the virus showed an eclipse phase of 7 to 9 hours, with production of cell-free infectious virus 18 hours post-infection. Plaque reduction neutralization tests suggested that complement enhanced the neutralizing activity of sera; rat sera were particularly complement-dependent. The plaque reduction neutralization test was about 10 times more sensitive than the TCID50 neutralization test. Convalescent phase sera from patients with hemorrhagic fever with renal syndrome (HFRS) having higher IF antibody titers to Hantaan virus than to nephropathia epidemica (NE) virus were capable of neutralizing Hantaan virus, while sera from patients with higher IF antibody titers to NE virus than Hantaan virus did not contain neutralizing antibody to Hantaan virus.