Rapid diagnosis of rabies in humans and animals by a dot blot enzyme immunoassay.
OBJECTIVE
The presently advocated tests for rapid diagnosis of rabies, such as the fluorescent antibody test (FAT) are expensive and require expertise to carry out and interpret the results. In this study, a simple direct dot blot enzyme immunoassay (DIA) has been developed and evaluated to detect the rabies antigen in brain specimens of animals and humans. The utility of this test in the ante-mortem diagnosis of human rabies has also been evaluated.


METHODS
Brain homogenates of suspected rabid animals (n = 250), humans (n = 16) and clinical samples like saliva (n = 12) and cerebrospinal fluid (CSF) (n = 12) were directly spotted on polyvinylidene difluoride membrane (PVDF) and the absorbed rabies nucleoprotein antigen was detected using biotinylated antinucleoprotein antibody followed by treatment with streptavidin peroxidase conjugate and color development with diamino benzedine (DAB). Rabies-infected and normal mouse brain homogenates were used as positive and negative controls, respectively. The results of this test were evaluated with fluorescent antibody technique (for brain samples) and mouse inoculation test (for saliva and CSF samples).


RESULTS
A distinct dark brown color was seen in the positive control and all positive samples, while there was no color development with either the negative control or the negative samples. The concordance between the fluorescent antibody test (FAT) and dot immunoassay was 98.4% for brain samples, 83.3% for saliva and 91.6% for CSF samples. The specificity of the test was found to be 100%.


CONCLUSIONS
The dot blot enzyme immunoassay (DIA) test described here is a sensitive, specific and rapid test for the post-mortem diagnosis of rabies in animals and humans. The utility of this test for the ante-mortem diagnosis of rabies needs to be further evaluated.
Madhusudana
Shampur Narayan
SN
Department of Neurovirology, National Institute of Mental Health and Neurosciences, PB No. 2900, Hosur Road, Bangalore 560029, India. mshampur@hotmail.com
Paul
Joel Prem Vasanth
JP
Abhilash
Venugopal Karavattu
VK
Suja
Mooriyath Sukumaran
MS
D000818:Animals; D000956:Antigens, Viral / Q000032:analysis; D001921:Brain / Q000821:virology*; D002415:Cats; D002417:Cattle; D002555:Cerebrospinal Fluid / Q000821:virology; D004285:Dogs; D005455:Fluorescent Antibody Technique; D006801:Humans; D015151:Immunoblotting / Q000379:methods*; D007124:Immunoenzyme Techniques / Q000379:methods; D051379:Mice; D011818:Rabies / Q000175:diagnosis* / Q000662:veterinary / Q000821:virology; D011820:Rabies virus / Q000302:isolation & purification*; D012463:Saliva / Q000821:virology; D012680:Sensitivity and Specificity; D013997:Time Factors
D000956:Antigens, Viral