Pseudomonas aeruginosa is an opportunistic pathogen that is highly resistant to antibiotics, especially when it grows in biofilms. As an alternative to antibiotic intervention, antimicrobial antibody drugs have drawn attention in recent years due to their immunotherapeutic functions. In this study, we designed a monoclonal scFv-Fc-form antibody, MFb, targeting P. aeruginosa antigen alginate and investigated its function against this bacterium in vitro. MFb was generated by transient gene expression in HEK293 cells and purified by one-step protein A affinity chromatography. Experiments showed that MFb could recognize alginate specifically based on enzyme-linked immunosorbent assays. Its KD value of 8.31 nM was determined by surface plasmon resonance, demonstrating its high affinity for alginate. Further detailed studies revealed that the antibody exerted antibacterial effects by three mechanisms: 1) MFb inhibited P. aeruginosa biofilm formation with an IC50 of 0.58 μg/mL; 2) MFb reduced P. aeruginosa adhesion to HeLa cells, and successfully prevented its invasion on epithelial cells; 3) based on an in vitro macrophage phagocytosis assay, MFb could enhance the phagocytotic capacity of macrophages for P. aeruginosa in a concentration-dependent manner. Taken together, our work demonstrated that the antimicrobial monoclonal antibody MFb has a protective effect on HeLa cells, and it may be a promising novel strategy to treat P. aeruginosa infection.

中文翻译：

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抗藻酸盐单克隆抗体对HeLa细胞铜绿假单胞菌感染的保护作用。

铜绿假单胞菌是机会性病原体，对抗生素具有高度抗性，尤其是当其在生物膜中生长时。作为抗生素干预的替代方法，近年来，抗微生物抗体药物因其免疫治疗功能而备受关注。在这项研究中，我们设计了针对铜绿假单胞菌抗原藻酸盐的单克隆scFv-Fc形式抗体MFb，并研究了其对这种细菌的体外功能。MFb由HEK293细胞中的瞬时基因表达产生，并通过一步蛋白A亲和层析纯化。实验表明，基于酶联免疫吸附试验，MFb可以特异性识别藻酸盐。通过表面等离子体激元共振测定其KD值为8.31 nM，表明其对藻酸盐的高亲和力。进一步的详细研究表明，该抗体通过三种机制发挥抗菌作用：1）MFb抑制铜绿假单胞菌生物膜形成，IC50为0.58μg/ mL；2）MFb减少了铜绿假单胞菌对HeLa细胞的粘附，并成功地阻止了其侵袭上皮细胞。3）基于体外巨噬细胞吞噬作用测定，MFb可以以浓度依赖性方式增强巨噬细胞对铜绿假单胞菌的吞噬能力。综上所述，我们的工作证明抗微生物单克隆抗体MFb对HeLa细胞具有保护作用，它可能是治疗铜绿假单胞菌感染的一种有前途的新策略。并成功地阻止了其侵袭上皮细胞；3）基于体外巨噬细胞吞噬作用测定，MFb可以以浓度依赖性方式增强巨噬细胞对铜绿假单胞菌的吞噬能力。综上所述，我们的工作证明抗微生物单克隆抗体MFb对HeLa细胞具有保护作用，它可能是治疗铜绿假单胞菌感染的一种有前途的新策略。并成功地阻止了其侵袭上皮细胞；3）基于体外巨噬细胞吞噬作用测定，MFb可以以浓度依赖性方式增强巨噬细胞对铜绿假单胞菌的吞噬能力。综上所述，我们的工作证明抗微生物单克隆抗体MFb对HeLa细胞具有保护作用，它可能是治疗铜绿假单胞菌感染的一种有前途的新策略。