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Published on: January 25, 2017
Serodiagnosis of human plague by a combination of immunomagnetic separation and flow cytometry
W D Splettstoesser1, R Grunow, L Rahalison
1Department of Microbiology, Central Institute of the Federal Armed Forces Medical Services-Koblenz, Koblenz, Germany. SanAkBw_IMB@t-online.de
Background:
Plague is a severe, highly communicable bacterial disease caused by Yersinia pestis. It is still endemic in more than 20 countries worldwide. Although known as a devastating disease for centuries, laboratory confirmation of clinical suspected cases is still problematic. No standardized and internationally approved test system is commercially available. The aim of this study was the introduction and evaluation of a combination of immunomagnetic separation and flow cytometry for the serodiagnosis of human plague.
Methods:
Paramagnetic polystyrene beads were coated with purified F1 capsular antigen (F1 CA) and reacted with sera from plague patients, from 26 laboratory personnel vaccinated against plague and from 102 healthy blood donors (HBD). After incubation with fluorescein isothiocyanate-conjugated anti-human rabbit IgG, particle-associated fluorescence was detected by flow cytometry.
Results:
Anti-F1 CA antibodies could be demonstrated in all patients with bacteriologically confirmed plague and in 22 sera (84.6%) from vaccinees. Only one serum in the HBD group showed a weakly positive reaction. The total assay time was less than 2 h.
Conclusions:
Compared with a recently published combination of an anti-F1 CA enzyme-linked immunosorbent assay (ELISA) and immunoblot, the new assay showed the same sensitivity as the ELISA and almost the same specificity (99.0 versus 100%) as the immunoblot. Allowing a rapid, reliable, and quantitative analysis, immunomagnetic separation combined with flow cytometry might replace both conventional immunoassays.
Insights
A new assay using immunomagnetic separation and flow cytometry offers rapid and reliable serodiagnosis for plague, a severe bacterial disease caused by Yersinia pestis. This method accurately detects anti-F1 CA antibodies, potentially replacing traditional immunoassays.
Area of Science:
- Medical Microbiology
- Immunology
- Diagnostic Technology
Background:
- Plague, a severe bacterial disease caused by Yersinia pestis, remains endemic globally.
- Current laboratory confirmation of plague cases is problematic due to a lack of standardized diagnostic tests.
- There is a need for a reliable and internationally approved test system for human plague serodiagnosis.
Purpose of the Study:
- To introduce and evaluate a novel diagnostic approach for human plague serodiagnosis.
- To assess the efficacy of combining immunomagnetic separation (IMS) with flow cytometry (FC) for plague detection.
- To provide a rapid, reliable, and quantitative method for identifying anti-F1 CA antibodies.
Main Methods:
- Paramagnetic beads coated with F1 capsular antigen (F1 CA) were used.
- Sera from plague patients, vaccinated individuals, and healthy blood donors were analyzed.
- Particle-associated fluorescence was measured using flow cytometry after antibody incubation.
Main Results:
- Anti-F1 CA antibodies were detected in all confirmed plague patients and 84.6% of vaccinees.
- Only one healthy blood donor showed a weak positive reaction, indicating high specificity.
- The entire assay procedure was completed in under 2 hours.
Conclusions:
- The IMS-FC assay demonstrates comparable sensitivity to ELISA and high specificity, similar to immunoblotting.
- This method offers a rapid, reliable, and quantitative alternative for plague serodiagnosis.
- Immunomagnetic separation combined with flow cytometry shows potential to replace conventional immunoassays for plague detection.
