Infection and Drug Resistance (Jul 2022)
Development of Highly Sensitive Sandwich ELISA for the Early-Phase Diagnosis of Chikungunya Virus Utilizing rE2-E1 Protein
Abstract
Mohammad Islamuddin,1,2 Abuzer Ali,3 Wajihul Hasan Khan,4 Amena Ali,5 Syed Kazim Hasan,1 Mohd Abdullah,6 Kentaro Kato,2 Malik Zainul Abdin,7 Shama Parveen1 1Molecular Virology Laboratory, Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, 110025, India; 2Laboratory of Sustainable Animal Environment, Graduate School of Agricultural Science, Tohoku University, Miyagi, Japan; 3Microbiology Laboratory, Ansari Health Center, Jamia Millia Islamia, New Delhi, 110025, India; 4Department of Pharmacognosy, College of Pharmacy, Taif University, Taif, 21944, Saudi Arabia; 5Molecular Virology Lab, Department of Microbiology, All India Institute of Medical Sciences (AIIMS), New Delhi, 110029, India; 6Department of Pharmaceutical Chemistry, College of Pharmacy, Taif University, Taif, 21944, Saudi Arabia; 7Department of Biotechnology, School of Chemical and Life Sciences, Hamdard University, New Delhi, 110026, IndiaCorrespondence: Mohammad Islamuddin; Shama Parveen, Email [email protected]; [email protected]: Chikungunya is caused by an alpha virus transmitted to humans by an infected mosquito. Infection is generally considered to be self-limiting and non-critical. Chikungunya infection may be diagnosed by severe joint pain with fever, but it is difficult to diagnose because the symptoms of chikungunya are common to many pathogens, including dengue fever. Diagnosis mainly depends on viral culture, reverse transcriptase polymerase chain reaction (RT-PCR), and IgM ELISA. Early and accurate diagnosis of the virus can be achieved by the application of PCR methods, but the high cost and the need for a thermal cycler restrict the use of such methods. On the other hand, antibody-based IgM ELISA is considered to be inexpensive, but antibodies against chikungunya virus (CHIKV) only develop after 4 days of infection, so it has limited application in the earlier diagnosis of viral infection and the management of patients. Because of these challenges, a simple antigen-based sensitive, specific, and rapid detection method is required for the early and accurate clinical diagnosis of chikungunya.Methods: The amino acid sequence of CHIKV ectodomain E1 and E2 proteins was analyzed using bioinformatics tools to determine the antigenic residues, particularly the B-cell epitopes and their characteristics. Recombinant E2-E1 CHIKV antigen was used for the development of polyclonal antibodies in hamsters and IgG was purified. Serological tests of 96 CHIKV patients were conducted by antigen-capture ELISA using primary antibodies raised against rCHIKV E2-E1 in hamsters and human anti-CHIKV antibodies.Results: We observed high specificity and sensitivity, of 100% and 95.8%, respectively, and these values demonstrate the efficiency of the test as a clinical diagnostic tool. There was no cross-reactivity with samples taken from dengue patients.Discussion: Our simple and sensitive sandwich ELISA for the early-phase detection of CHIKV infection may be used to improve the diagnosis of chikungunya.Graphical Abstract: Keywords: chikungunya, sandwich ELISA, recombinant E2-E1, B-cell epitopes, IgG, hamster antibodies
