Das Biswadeep, Samal Sagnika, Hamdi Hamida, Dash Manoj Kumar, Singh Gyanraj
School of Biotechnology, Kalinga Institute of Industrial Technology, Deemed to Be University, Bhubaneswar, Odisha 751024, India.
School of Biotechnology, Kalinga Institute of Industrial Technology, Deemed to Be University, Bhubaneswar, Odisha 751024, India.
Diagn Microbiol Infect Dis. 2025 Mar;111(3):116679. doi: 10.1016/j.diagmicrobio.2025.116679. Epub 2025 Jan 6.
Dengue virus (DENV) is an important arthropod-borne viral disease, with four antigenically and genetically diverse serotypes (DENV-1, DENV-2, DENV-3, and DENV-4). Timely and accurate diagnosis of dengue virus serotypes is crucial for the management of outbreaks. This study focussed on the development of a RT-PCR based lateral flow strip assay to detect DENV serotypes in a dual detection manner without using gel electrophoresis. The assay uses anti-biotin/streptavidin colloidal gold conjugates with fluorescent/enzymatic tagged DENV serotype specific antibodies for the direct detection of DENV infected serum samples on a nitrocellulose membrane using biotin-BSA as control line. The detection limit of the assay was up to 10 copies of cDNA for DENV-1 and 100 copies for DENV-2, DENV-3, and DENV-4. In house evaluation of DENV LFIA demonstrated 100 % sensitivity in all the serotypes compared to conventional RT-PCR, 100 % specificity for DENV-1, DENV-2, DENV-3, and 95 % specificity for DENV-4 detection. DENV serotyping was assessed in a dual detection manner (DENV-1/DENV-3 and DENV-2/DENV-4 at two test lines) on the strip. The limitation of the assay is the requirement of PCR for initial amplification and confirmation of individual serotype in case of DENV-1/DENV-3 and DENV-2/DENV-4 detection, besides the field evaluation of the assay detected DENV-2 and DENV-3 serotypes, and no other serotype was detected in line with RT-PCR findings.
登革病毒（DENV）是一种重要的节肢动物传播的病毒性疾病，有四种抗原性和基因不同的血清型（DENV-1、DENV-2、DENV-3和DENV-4）。及时准确地诊断登革病毒血清型对于疫情管理至关重要。本研究聚焦于开发一种基于逆转录聚合酶链反应（RT-PCR）的侧向流动试纸条检测法，以双重检测方式检测登革病毒血清型，且无需使用凝胶电泳。该检测法使用抗生物素蛋白/链霉亲和素胶体金缀合物与荧光/酶标记的登革病毒血清型特异性抗体，以生物素化牛血清白蛋白（biotin-BSA）作为对照线，在硝酸纤维素膜上直接检测登革病毒感染的血清样本。该检测法的检测限对于DENV-1的cDNA高达10个拷贝，对于DENV-2、DENV-3和DENV-4为100个拷贝。与传统RT-PCR相比，对登革病毒侧向流动免疫分析（LFIA）进行的内部评估显示，所有血清型的灵敏度均为100%，DENV-1、DENV-2、DENV-3的特异性为100%，DENV-4检测的特异性为95%。在试纸条上以双重检测方式（在两条测试线处检测DENV-1/DENV-3和DENV-2/DENV-4）评估登革病毒血清分型。该检测法的局限性在于，在检测DENV-1/DENV-3和DENV-2/DENV-4时，需要进行PCR进行初始扩增和确认单个血清型，此外，该检测法的现场评估检测到了DENV-2和DENV-3血清型，且未检测到其他血清型，这与RT-PCR结果一致。
