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Review
. 2017 Jan 25:8:55.
doi: 10.3389/fmicb.2017.00055. eCollection 2017.

xMAP Technology: Applications in Detection of Pathogens

Nikol Reslova  1 , Veronika Michna  2 , Martin Kasny  3 , Pavel Mikel  2 , Petr Kralik  4
Affiliations
Review

xMAP Technology: Applications in Detection of Pathogens

Nikol Reslova et al. Front Microbiol. .

Abstract

xMAP technology is applicable for high-throughput, multiplex and simultaneous detection of different analytes within a single complex sample. xMAP multiplex assays are currently available in various nucleic acid and immunoassay formats, enabling simultaneous detection and typing of pathogenic viruses, bacteria, parasites and fungi and also antigen or antibody interception. As an open architecture platform, the xMAP technology is beneficial to end users and therefore it is used in various pharmaceutical, clinical and research laboratories. The main aim of this review is to summarize the latest findings and applications in the field of pathogen detection using microsphere-based multiplex assays.

Keywords: diagnostics; immunoassay; magnetic microspheres; multiplex detection; nucleic acid detection; pathogen identification; xMAP.

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Figures

FIGURE 1
FIGURE 1
The xMAP Technology based on internally dyed microspheres. Different concentrations of red and infrared fluorophores were used to create 100 distinct microsphere sets. Each set is able to conjugate to a specific _target molecule (yellow and orange lines = nucleic acid; green star = fluorescent reporter).
FIGURE 2
FIGURE 2
Microsphere architecture. The polystyrene divinylbenzene core is surrounded by a polymer layer, which is formed by polystyrene methacrylic acid (infusion of dyes). The surface of each microsphere is irregular, porous and carboxylated. Magnetic microspheres have an additional layer of magnetite within the polymer layer and so differ also in size.
FIGURE 3
FIGURE 3
Principle of analysis by the MAGPIX fluorescent imager. Magnetic microspheres immobilized on a magnet are recognized by LEDs and a CCD camera records the picture (LED, light-emitting diode; CCD, charge-coupled device).
FIGURE 4
FIGURE 4
Direct DNA hybridization (DDH, yellow lines = capture oligonucleotide; orange line = amplified _target sequence; green star = fluorescent reporter). _target DNA sequence is amplified, while one of the primers is fluorescently labeled. Amplicons are then specifically hybridized (according to complementarity) to capture oligonucleotides on the microsphere surface.
FIGURE 5
FIGURE 5
Principle of Allele-specific primer extension (ASPE) (red and green line = anti-TAGs; green star = fluorescent reporter). Allele-specific detection probes, differing in one nucleotide on the polymorphic side, hybridize to amplified _target sequence. After addition of DNA polymerase and dNTPs (one of which is fluorescently labeled), molecules are extended according to complementarity. Products are captured by anti-TAGs on the specific microsphere set.
FIGURE 6
FIGURE 6
Principle of Single base chain extension (SBCE) (red dot = dideoxynucleotide; green star = fluorescent reporter; red line = anti-TAG). Specific detection probes are terminated one base before the polymorphic site. Utilization of fluorescently labeled dideoxynucleotides necessitates a separate reaction for each nucleotide in focus (minimally two). _target DNA hybridizes with probes after amplification but only the mix with the proper ddNTP leads ultimately to the synthesis of a labeled product, which is captured by anti-TAG on the microsphere surface.
FIGURE 7
FIGURE 7
Principle of Oligonucleotide ligation assay (OLA) (green star = fluorescent reporter; Pho = phosphate group; red line = anti-TAG). The _target DNA sequence is PCR-amplified prior to the ligation step of the annealed probes. One of the detection probes consists of a sequence complementary to the _target sequence (polymorphic site at the 3′ end if SNP identification is needed) and also an additional TAG tail sequence. The second detection probe is fully complementary to the _target sequence and serves as a reporter due to its fluorescent label at the 3′ end. Detection probes bind next to each other, DNA ligase recognizes the nick and makes a bond. The product is captured by anti-TAG on the microsphere surface.
FIGURE 8
FIGURE 8
Principle of Multiplex oligonucleotide ligation PCR assay (MOL-PCR) (orange line = detection probe 1; green line = detection probe 2; blue lines = universal PCR primers; burgundy line = amplified negative strand; green star = fluorescent reporter; Pho = phosphate group; red line = anti-TAG). Specific detection probes bind next to each other to _target sequence via complementary parts, while the parts including the TAG sequence and binding sites for PCR primers form tails sticking out into space. DNA ligase recognizes the nick and makes a bond. The complex sequence of ligated probes becomes a template for singleplex PCR with universal primers; one of the primers is fluorescently labeled. Labeled amplicon hybridizes via its TAG sequence to capture anti-TAG on the microsphere.
FIGURE 9
FIGURE 9
Principle of microsphere-based multiplex immunoassays. (A) Capture sandwich (CS; yellow hexagon = _target; blue Y = capture antibodies; green Y = detection antibody; green star = fluorescent reporter); (B) Indirect serological assay (ISA; yellow hexagon = capture antigen; blue Y = specific _target antibody; green Y = detection anti-antibody; green star = fluorescent reporter).
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References

    1. Anderson J. P., Rascoe L. N., Levert K., Chastain H. M., Reed M. S., Rivera H. N., et al. (2015). Development of a luminex bead based assay for diagnosis of toxocariasis using recombinant antigens Tc-CTL-1 and Tc-TES-26. PLoS Negl. Trop. Dis. 9:e0004168 10.1371/journal.pntd.0004168 - DOI - PMC - PubMed
    1. Angeloni S., Cordes R., Dunbar S., Garcia C., Gibson G., Martin C., et al. (2014). xMAP Cookbook: A Collection of Methods and Protocols for Developing Multiplex Assays with xMAP Technology, 2nd Edn (Austin, TX: Luminex; )
    1. Anonymous. (2014). Multicriteria-Based Ranking for Risk Management of Food-Borne Parasites: Microbiological Risk Assessment Series (MRA) 23. Rome: FAO, 324.
    1. Anonymous. (2015). Regulation (EU) 2015/1375, laying down specific rules on official controls for Trichinella in meat. Official J. Eur. Union 28 7.
    1. Babady N. E., Miranda E., Gilhuley K. A. (2011). Evaluation of Luminex xTAG fungal analyte-specific reagents for rapid identification of clinically relevant fungi. J. Clin. Microbiol. 49 3777–3782. 10.1128/JCM.01135-11 - DOI - PMC - PubMed
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