Development and evaluation of antigen capture ELISA for early clinical diagnosis of chikungunya.
The resurgence of chikungunya (CHIK) in the form of unprecedented explosive epidemic after a gap of 3 decades in India and Indian Ocean islands is a point of major public health concern. The laboratory diagnosis is essentially based on virus isolation, IgM ELISA, and reverse transcriptase polymerase chain reaction (RT-PCR). Although PCR-based methods are used for early and accurate diagnosis, the high cost of the assay and requirement of thermal cycler limit its application only to referral laboratories. The antibody-based IgM ELISA is found to be cost-effective, but it takes 5 to 6 days for the patient to develop antibody and, thus, has less implication for early clinical diagnosis and patient management. Therefore, a simple rapid, sensitive, and specific antigen detection system is reported for early and reliable clinical diagnosis as well as effective surveillance of CHIK. A double antibody sandwich system was designed for antigen capture ELISA, employing rabbit and mouse anti-CHIK IgG antibodies as capture and detector antibodies, respectively. An optimal assay condition with 0 background was established having no reactivity with healthy human serum and Cerebro spinal fluid (CSF) samples. The comparative evaluation with SYBR Green I-based real-time RT-PCR revealed an accordance of 96% with a sensitivity and specificity of 95% and 97%, respectively. The specificity of this assay was confirmed through cross-reactivity studies with confirmed dengue and Japanese encephalitis (JE) patient serum and CSF samples. The antigen capture ELISA reported in this study was able to detect the presence of viral antigen as early as the second day of fever and, thus, can be very useful for early clinical diagnosis of CHIK with acute phase patient serum and CSF samples. This can also be used for rapid screening of large numbers of clinical samples in endemic areas during epidemics.
Shukla
Jyoti
J
Division of Virology Division, Defence Research and Development Establishment, Gwalior-474002, India.
Khan
Mohsin
M
Tiwari
Mugdha
M
Sannarangaiah
Santhosh
S
Sharma
Shashi
S
Rao
Putcha Venkata Lakshmana
PV
Parida
Manmohan
M
D018354:Alphavirus Infections / Q000175:diagnosis*; D000818:Animals; D000914:Antibodies, Viral; D000956:Antigens, Viral / Q000097:blood* / Q000134:cerebrospinal fluid*; D002646:Chikungunya virus / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D051379:Mice; D011817:Rabbits; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity
D000914:Antibodies, Viral; D000956:Antigens, Viral