Environmental Health Institute, National Environment Agency (NEA), 11 Biopolis Way, Singapore, Singapore.
Infectious Diseases Translational Research Programme (ID TRP), Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.
Virol J. 2024 Sep 3;21(1):208. doi: 10.1186/s12985-024-02459-y.
Dengue is a global public health challenge which requires accurate diagnostic methods for surveillance and control. The gold standard for detecting dengue neutralizing antibodies (nAbs) is the plaque reduction neutralization test (PRNT), which is both labor-intensive and time-consuming. This study aims to evaluate three alternative approaches, namely, the MTT-based (or (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) microneutralization assay, the xCELLigence real-time cell analysis (RTCA), and the immuno-plaque assay-focus reduction neutralization test (iPA-FRNT).
Twenty-two residual serum samples were tested for DENV-2 nAbs using all four assays at three neutralization endpoints of 50%, 70% and 90% inhibition in virus growth. For each neutralization endpoint, results were compared using linear regression and correlation analyses. Test performance characteristics were further obtained for iPA-FRNT using 38 additional serum samples.
Positive correlation of DENV-2 neutralization titers for the MTT-based microneutralization assay and the PRNT assay was only observed at the neutralization endpoint of 50% (r = 0.690). In contrast, at all three neutralization end points, a linear trend and positive correlation of DENV-2 neutralization titers for the xCELLigence RTCA and the PRNT assays were observed, yielding strong or very strong correlation (r = 0.829 to 0.967). This was similarly observed for the iPA-FRNT assay (r = 0.821 to 0.916), which also offered the added advantage of measuring neutralizing titers to non-plaque forming viruses.
The xCELLigence RTCA and iPA-FRNT assays could serve as suitable alternatives to PRNT for dengue serological testing. The decision to adopt these methods may depend on the laboratory setting, and the utility of additional applications offered by these technologies.
登革热是一项全球性的公共卫生挑战，需要准确的诊断方法来进行监测和控制。检测登革热中和抗体（nAbs）的金标准是噬斑减少中和试验（PRNT），该方法既费时又费力。本研究旨在评估三种替代方法，即基于 MTT 的（或（3-（4,5-二甲基噻唑-2-基）-2,5-二苯基四唑溴盐）微量中和测定法、xCELLigence 实时细胞分析（RTCA）和免疫噬斑测定-焦点减少中和试验（iPA-FRNT）。
用所有四种方法在病毒生长抑制的 50%、70%和 90%三个中和终点检测 22 份残留血清样本中的 DENV-2 nAbs。对于每个中和终点，通过线性回归和相关分析比较结果。使用另外 38 份血清样本进一步获得 iPA-FRNT 的测试性能特征。
仅在 50%的中和终点观察到基于 MTT 的微量中和测定法和 PRNT 测定法的 DENV-2 中和效价的正相关（r=0.690）。相比之下，在所有三个中和终点，xCELLigence RTCA 和 PRNT 测定法的 DENV-2 中和效价呈线性趋势和正相关，相关性很强或非常强（r=0.829 至 0.967）。iPA-FRNT 测定法也观察到类似的情况（r=0.821 至 0.916），该测定法还具有测量非噬菌斑形成病毒的中和效价的额外优势。
xCELLigence RTCA 和 iPA-FRNT 测定法可作为 PRNT 的替代方法用于登革热血清学检测。采用这些方法的决定可能取决于实验室环境，以及这些技术提供的其他应用的实用性。
