Double-antigen sandwich ELISA for detecting dengue virus EDⅢ-specific antibody
DING Xi-xia1, CAI Jian-piao2, WEN Kun1, ZHAN Li-li1, SONG Jia-ling1, CHEN Man-jun1, CHE Xiao-Yan1
1. Division of Laboratory Medicine,Zhujiang Hospital,Southern Medical University,Guangzhou 510282,China; 2. Department of Microbiology, University of Hong Kong, HongKong 999077, China
Abstract:This study investigated whether the EDⅢ protein of DENV1-4 is suitable for the diagnosis of dengue virus (DENV) infection. Four serotypes of DENV recombinant EDⅢ protein were expressed in Pichia pastoris and labeled with Horseradis hperoxidase(HRP) through a modified sodium periodate method. A double-antigen sandwich enzyme-linked immunosorbent assay (ELISA) for detecting EDⅢ-specific antibodies was successfully established by determining the optimal coating concentration and proportion of EDⅢ protein and HRP-EDⅢ protein for the four serotypes of DENV. The assay showed specificity to DENV1-4 EDⅢ protein immunized animal serum, with no cross-reaction with Aspergillus fumigatus MP1 immunized animal serum. The sensitivity of the double-antigen sandwich ELISA was evaluated with the Australian Panbio MAC ELISA test in sera from 168 patients with confirmed DENV infection treated at Guangzhou Zhujiang Hospital in 2014. No significant difference was found between the double-antigen sandwich ELISA and Panbio MAC ELISA (57.1% vs 57.7%, P<0001). Combining the detection results of NS1 antigen and EDⅢ antibodies markedly increased the diagnostic sensitivity from 57.1% to 97.6%. Thus, our double-antigen sandwich ELISA has high specificity in detecting EDⅢ-specific antibodies in the sera of patients with DENV infection, and simultaneous detection of antigen and antibodies can increase the diagnostic sensitivity for DENV infection.
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