Immunofluorescent staining of trypsinized formalin-fixed brain smears for rabies antigen: results compared with those obtained by standard methods for 221 suspect animal cases in Nigeria

Formalin-fixed samples from 221 animal brains received for rabies diagnosis in Nigeria were digested in 0.1% trypsin in phosphate buffered saline, pH 7.4, and smears stained for rabies antigen by direct immunofluorescence (IF). The results were compared with those obtained using fresh material from the same animals for Negri body staining, mouse inoculation (MI) and occasionally immunofluorescent staining. From 191 specimens examined for Negri bodies and by mouse inoculation 51 and 64 respectively proved positive. The IF smear technique under investigation failed to detect 5 of these but showed up as positive 30 which had been recorded as Negri-negative and 19 that had gone undetected by MI too. In a direct comparison with IF staining of fresh tissue from 23 known rabies positive animals the similar staining of trypsin-digested formalized smears failed to give a positive result in 2 out of 23 cases. Some problems were encountered with smears not sticking to slides. When gelatinized slides were used fluorescence was sometimes poorer. Where transport and refrigeration are difficult and section-cutting equipment is lacking the technique of IF staining of smears prepared from formalized brain tissue after treatment with trypsin can be a useful adjunct to other diagnostic methods. It also makes for safer working where special facilities are absent.