Abstract
BackgroundYersinia pestis is a contributing agent to the epidemic disease, plague, which killed an estimated 200 million people during historical times. In this study, a rapid, cheap, sensitive, and specific technique, the lateral flow assay (F1 strips), has been successfully developed to detect this pathogen, by using paired monoclonal antibodies (MAbs) against Y. pestis capsule like fraction 1 (F1) protein. Compared with the polyclonal antibody (PAb) based F1 strips, the Mab-based F1 strips have a remarkable increased detection limitation (10 to 100 folds). Furthermore, besides the limitation and specificity evaluation, the application of this F1 strip on simulated clinical samples indicate the LFA can be a good candidate to detect plague.MethodsRecombinant F1 antigen was expressed and purified from a series of works. The various anti-F1 monoclonal antibodies generated from hybridoma cells were screened with the ELISA technique. To evaluate the feasibility of this Y. pestis F1 test strip, the F1 protein/Y. pestis was spiked into simulated clinical samples such as human serum, mouse bronchoalveolar lavage fluids, and mouse blood to mimic natural infection status. Additionally, this technique was applied to detect the Y. pestis in the environment-captured rats, to evaluate the practical usefulness of the strips.ResultsBy using this MAb-based-LFA technique, 4 ng/ml of recombinant F1-protein and 103 CFU/ml of Y. pestis could be detected in less than 10 mins, which is at least 10-folds than that of the PAb format. On the other hand, although various Yersinia strains were applied to the strips, only Y. pestis strain showed a positive result; all other Yersinia species did not produce a positive signal, indicating the high efficiency and specificity of the MAb-based F1-strips.ConclusionBased on our findings, we suggest that the MAb-format-LFA will be valuable as a diagnostic tool for the detection of Y. pestis. This report shows that the F1 strip is sufficient to support not only the detection of plague in simulated clinical samples, but also it may be a good candidate to meet the epidemiological surveillance during an outbreak of the biological warfare.
Highlights
Yersinia pestis is a contributing agent to the epidemic disease, plague, which killed an estimated 200 million people during historical times
The results demonstrated that Monoclonal antibody (MAb) format fraction 1 (F1)-strip is an ideal candidate for detection of Y. pestis in the environment and clinic
The caf gene cluster could express the F1 structural protein. This plasmid was transformed into E. coli for F1 protein production
Summary
Yersinia pestis is a contributing agent to the epidemic disease, plague, which killed an estimated 200 million people during historical times. Yersinia pestis is a slow-growing, non-motile, and non-spore-forming gram-negative coccobacillus (0.5 ~ 0.8 μm in diameter) of the family Enterobacteriaceae It is regarded as a facultative extracellular pathogen and is the causative agent of the notorious plague [1]. The bubonic and septicemic plague is not proven to be epidemical, it is possible for patients to transmit the pneumonic plague This is the most serious form that can produce highly contagious aerosols in the environment; the distance required for effective transmission is only 2-m [4, 5]. Owing to the high virulence and the high mortality rate due to Y. pestis infections, this species has the potential to be developed as a biological weapon [1, 11] It is currently categorized as a restricted agent (A-list) by the CDC (Centers for Disease Control and Prevention, Atlanta, USA). This study would meet the requirement of epidemiological surveillance on site and/or during biological warfare, and could reduce mortality, through early monitoring and adequate treatment, and effectively control distribution of the infectious agent
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