Development of novel antibodies against non-structural proteins nsP1, nsP3 and nsP4 of chikungunya virus: potential use in basic research.
Chikungunya virus (CHIKV) has reemerged recently as an important pathogen, causing several large epidemics worldwide. This necessitates the development of better reagents to understand its biology and to establish effective and safe control measures. The present study describes the development and characterization of polyclonal antibodies (pAbs) against synthetic peptides of CHIKV non-structural proteins (nsPs; nsP1, nsP3 and nsP4). The reactivity of these pAbs was demonstrated by ELISA and Western blot. Additionally, in vitro infection studies in a mammalian system confirmed that these pAbs are highly sensitive and specific for CHIKV nsPs, as these proteins were detected very early during viral replication. Homology analysis of the selected epitope sequences revealed that they are conserved among all of the CHIKV strains of different genotypes, while comparison with other alphavirus sequences showed that none of them are 100% identical to the epitope sequences (except Onyong-nyong and Igbo Ora viruses, which show 100% identity to the nsP4 epitope). Interestingly, two different forms of CHIKV nsP1 and three different forms of nsP3 were detected in Western blot analysis during infection; however, further experimental investigations are required to confirm their identity. Also, the use of these antibodies demonstrated faster and enhanced expression profiles of all CHIKV nsPs in 2006 Indian outbreak strains when compared to the CHIKV prototype strain, suggesting the epidemic potential of the 2006 isolate. Accordingly, it can be suggested that the pAbs reported in this study can be used as sensitive and specific tools for experimental investigations of CHIKV replication and infection.
Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Bhubaneswar, 751023, India.
Sameer
S
Kumar
Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Bhubaneswar, 751023, India.
Prabhudutta
P
Mamidi
Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Bhubaneswar, 751023, India.
Abhishek
A
Kumar
Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Bhubaneswar, 751023, India.
Itishree
I
Basantray
Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Bhubaneswar, 751023, India.
Umarani
U
Bramha
Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Bhubaneswar, 751023, India.
Anshuman
A
Dixit
Imgenex India Pvt. Ltd., Bhubaneswar, India.
Prasanta Kumar
PK
Maiti
Imgenex India Pvt. Ltd., Bhubaneswar, India.
Sujay
S
Singh
Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Bhubaneswar, 751023, India.
Amol Ratnakar
AR
Suryawanshi
School of Biological Sciences, National Institute of Science Education and Research, Bhubaneswar, India.
Subhasis
S
Chattopadhyay
Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Bhubaneswar, 751023, India. sochat.ils@gmail.com.
Soma
S
Chattopadhyay
D000595:Amino Acid Sequence; D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis* / Q000276:immunology; D015153:Blotting, Western; D065632:Chikungunya Fever / Q000821:virology*; D002646:Chikungunya virus / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification* / Q000502:physiology; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000235:genetics / Q000276:immunology; D008969:Molecular Sequence Data; D011817:Rabbits; D016415:Sequence Alignment; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D014779:Virus Replication
D000914:Antibodies, Viral; D000939:Epitopes; D017361:Viral Nonstructural Proteins