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Immunomagnetic Separation Improves the Detection of Mycobacteria by Paper-Based Lateral and Vertical Flow
Alejandra Ben Aissa1, Barbara Araújo1,2, Esther Julián3
1Departament de Química, Universitat Autònoma de Barcelona, 08193 Bellaterra, Spain.
Insights
A new method combines immunomagnetic separation and paper-based assays for sensitive detection of Mycolicibacterium fortuitum. This technique offers rapid screening of hemodialysis water, protecting patients from contamination.
Area of Science:
- Microbiology
- Analytical Chemistry
- Biotechnology
Background:
- Mycolicibacterium fortuitum poses a risk in healthcare settings, particularly in hemodialysis water.
- Sensitive and rapid detection methods are crucial for preventing patient exposure.
Purpose of the Study:
- To develop and evaluate a combined immunomagnetic separation (IMS) and paper-based nucleic acid immunochromatographic assay for detecting Mycolicibacterium fortuitum.
- To compare the analytical performance of nucleic acid vertical flow (NAVF) and nucleic acid lateral flow (NALF) platforms.
Main Methods:
- Immunomagnetic separation using antibody-specific magnetic particles for bacterial preconcentration.
- Double-tagging PCR amplification of the 16S rRNA gene, labeling amplicons with biotin and digoxigenin.
- Comparative analysis of NAVF and NALF paper-based platforms utilizing streptavidin gold nanoparticles for signal generation.
Main Results:
- Both NAVF and NALF assays, when coupled with IMS, achieved the required limit of detection in hemodialysis water.
- Nucleic acid lateral flow (NALF) demonstrated lower limits of detection compared to NAVF.
- Visual readout was successfully achieved through the capture of gold-modified amplicons.
Conclusions:
- The combined IMS and paper-based assay (NAVF and NALF) are promising for rapid screening of hemodialysis water.
- These methods can help prevent immunosuppressed patients from exposure to contaminated water sources.
- The study highlights the utility of these techniques in clinical and public health settings.
Abstract:
This work addresses a method that combines immunomagnetic separation (IMS) and paper-based nucleic acid immunochromatographic assay for the sensitive detection of Mycolicibacterium fortuitum (basonym Mycobacterium fortuitum) In particular, the preconcentration of the bacteria was achieved by using magnetic particles modified with an antibody specific towards mycobacteria. Following the IMS, the bacteria were lysed, and the genome was amplified by double-tagging PCR, using a set of primers specific for the 16S rRNA gene for Mycobacterium. During the amplification, the amplicons were labeled with biotin and digoxigenin tags. Moreover, a comparative study of paper-based immunochromatographic platforms, relying on vertical and lateral flow and on the use of streptavidin gold nanoparticles as a signal generating system, was also performed. The visual readout was achieved when the gold-modified amplicons were captured by the anti-DIG antibody in the test line. The analytical performance of both methods, nucleic acid vertical flow (NAVF) and nucleic acid lateral flow (NALF), is also discussed. Although NALF showed lower limit of detections (LODs), both NALF and NAVF combined with IMS were able to detect the required LOD in hemodialysis water, becoming two promising and useful techniques for the rapid screening of water supplies in hemodialysis centers, to prevent the exposure of immunosuppressed patients to contaminated sources.
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