Monoclonal Antibody Discovery Laboratory, Fondazione Toscana Life Sciences, Siena, Italy.
GlaxoSmithKline (GSK) Vaccines Institute for Global Health (GVGH), Siena, Italy.
Front Immunol. 2024 Apr 12;15:1374293. doi: 10.3389/fimmu.2024.1374293. eCollection 2024.
is the etiologic agent of a bacillary dysentery known as shigellosis, which causes millions of infections and thousands of deaths worldwide each year due to 's unique lifestyle within intestinal epithelial cells. Cell adhesion/invasion assays have been extensively used not only to identify targets mediating host-pathogen interaction, but also to evaluate the ability of -specific antibodies to reduce virulence. However, these assays are time-consuming and labor-intensive and fail to assess differences at the single-cell level.
Here, we developed a simple, fast and high-content method named visual Adhesion/Invasion Inhibition Assay (vAIA) to measure the ability of anti-antibodies to inhibit bacterial adhesion to and invasion of epithelial cells by using the confocal microscope Opera Phenix.
We showed that vAIA performed well with a pooled human serum from subjects challenged with and that a specific anti-IpaD monoclonal antibody effectively reduced bacterial virulence in a dose-dependent manner.
vAIA can therefore inform on the functionality of polyclonal and monoclonal responses thereby supporting the discovery of pathogenicity mechanisms and the development of candidate vaccines and immunotherapies. Lastly, this assay is very versatile and may be easily applied to other species or serotypes and to different pathogens.
是细菌性痢疾的病原体，又称志贺菌病，每年在全球导致数百万人感染和数千人死亡。由于其在肠上皮细胞内的独特生活方式，志贺氏菌能够逃避宿主的免疫防御机制。细胞黏附和侵袭试验已被广泛用于鉴定介导宿主-病原体相互作用的靶点，也用于评估针对的特异性抗体降低其毒力的能力。然而，这些试验耗时耗力，且无法评估单细胞水平的差异。
本研究开发了一种简单、快速和高通量的方法，称为可视化黏附和侵袭抑制试验（vAIA），使用共聚焦显微镜 Opera Phenix 来测量抗抗体抑制细菌黏附和侵袭上皮细胞的能力。
我们表明，vAIA 可用于评估来自志贺氏菌感染受试者的混合人血清，并表明一种特异性抗 IpaD 单克隆抗体可有效降低细菌的毒力，呈剂量依赖性。
因此，vAIA 可以反映多克隆和单克隆反应的功能，从而支持对致病性机制的发现以及候选疫苗和免疫疗法的开发。最后，该试验非常通用，可轻松应用于其他志贺氏菌种或血清型以及不同的病原体。
