Immunomagnetic Separation Improves the Detection of Mycobacteria by Paper-Based Lateral and Vertical Flow Immunochromatographic Assays

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.

Minimizing Cross-Reactivity for the Chemiluminescent Lateral Flow Immunoassay of Cardiac Troponin I Based on PEGylation of Gold Nanoparticles

“…Enhancing the sensitivity of a LFIA is critical for the detection of many biomarkers. In recent years, great efforts have been devoted to developing new strips with high sensitivity, mainly by conjugating antigens or antibodies to nanomaterials, such as gold nanoparticles (AuNPs), 3 magnetic nanoparticles, 6,7 quantum dots, 8 and fluorescent microspheres. 9,10 Among these methods, conjugating antibodies and horseradish peroxidase (HRP) to the AuNPs simultaneously can lead to effective and ultrasensitive chemiluminescent (CL) immunoassays (CL-LFIAs).…”

“…Enhancing the sensitivity of a LFIA is critical for the detection of many biomarkers. In recent years, great efforts have been devoted to developing new strips with high sensitivity, mainly by conjugating antigens or antibodies to nanomaterials, such as gold nanoparticles (AuNPs), magnetic nanoparticles, , quantum dots, and fluorescent microspheres. , Among these methods, conjugating antibodies and horseradish peroxidase (HRP) to the AuNPs simultaneously can lead to effective and ultrasensitive chemiluminescent (CL) immunoassays (CL-LFIAs). , Compared with other optical methods, the CL-LFIAs are highly specific and ultrasensitive with high signal-to-noise ratios . However, these assays usually suffer from cross-reactivity issues that would cause false-positive errors to further lower the detection accuracy.…”

Rapid and simultaneous detection of SARS-CoV-2 and influenza A using vertical flow assay based on AAO and SERS nanotags

“…In consequence, a series of strategies have been integrated with VFA for sensitivity improvement. [20][21][22] Among these technologies, surface enhanced Raman scattering (SERS) has impressive advantages for VFA, including distinctive fingerprint signals, excellent multiplexing, and ultrasensitivity. 23 Recently, SERS nanotags have been introduced into immunoassay for improving the LOD, enriching the quantitative ability and multiplex detection.…”

Osmotic Processor for Enabling Sensitive and Rapid Biomarker Detection via Lateral Flow Assays

“…Techniques have been developed to improve the sensitivities of LFAs, including kinetics and transport control ( Yang et al, 2013 ; Rivas et al, 2014 ; Tang et al, 2017 ; Ishii et al, 2018 ), biochemical signal amplification ( Hu et al, 2013 ; Parolo et al, 2013 ; Panferov et al, 2016 ), improved labeling ( Choi et al, 2010 ), and sample enrichment. Sample enrichment techniques include centrifugal filtration ( Corstjens et al, 2015 ), immunomagnetic separation ( Panferov et al, 2017 ; Ben Aissa et al, 2021 ), electrophoretic and phasic separation ( Wu et al, 2014 ; Chiu et al, 2015 ; Kim et al, 2017 ), isotachophoresis ( Moghadam et al, 2015 ), dialysis ( Tang et al, 2016 ), and test-zone pre-enrichment ( Zhang et al, 2020 ). These systems are not suitable for low-resource, point-of-care settings as they require expensive reagents, equipment, and/or complex procedures.…”