Immunofluorescence staining for detection of variola virus

In September 1975 Bangladesh was the only country in the world with endemic variola major, and the eradication of the disease was imminent. A rapid and accurate laboratory diagnostic method was required to supplement immunodiffusion in agar gel and culture on chorioallantoic membrane of embryonated egg available at the Institute of Public Health in Dacca, Bangladesh. To determine its effectiveness, a new, improved immunofluorescence (IF) staining technique was introduced. Laboratory specimens (scabs or vesicular or pustular impressions) were collected from patients who had, or were suspected of having, smallpox. Seventy-eight of 144 specimens collected were found to be IF positive for smallpox. As the number of laboratory-positive cases far exceeded the number of clinically diagnosed smallpox cases, IF-positive cases were reinvestigated and subsamples of the IF-positive specimens were tested at a World Health Organization poxvirus reference laboratory at the Centers for Disease Control in Atlanta, Ga. The results indicated 100% sensitivity for the IF technique (no false-negative results) in diagnosing variola major but also showed a high rate of false-positive results. Consequently, IF could not be recommended as a routine screening test for smallpox.