Future microbiology · 2016
Development of a rapid diagnostic assay based on magnetic bead purification of OXA-β-lactamase mRNA.
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Brandt Christian, Zander Esther, Pfeifer Yvonne, Braun Sascha D, Ehricht Ralf, Makarewicz Oliwia, Pletz Mathias W
Аннотация
AIM: To countermeasure the global spread of β-lactamases, we developed a rapid molecular test for the highly variable OXA-β-lactamases that allows minimizing the time to effective treatment.
METHODS: OXA-mRNA was specifically enriched from total RNA using group-specific biotinylated DNA probes and streptavidin-coated magnetic beads. Phylogenetic OXA groups were distinguished by PCR product size.
RESULTS: This mRNA fishing method is highly sensitive, yielding specific results from as little as 1 ng total RNA. It enables discrimination of OXA-extended substrate spectrum β-lactamases and carbapenemases and the semi-quantitative detection of highly expressed ISAba1-controlled variants.
CONCLUSION: Targeting mRNA with specific probes on magnetic beads will allow for adaptation to automated systems, such as point-of-care diagnostics.
METHODS: OXA-mRNA was specifically enriched from total RNA using group-specific biotinylated DNA probes and streptavidin-coated magnetic beads. Phylogenetic OXA groups were distinguished by PCR product size.
RESULTS: This mRNA fishing method is highly sensitive, yielding specific results from as little as 1 ng total RNA. It enables discrimination of OXA-extended substrate spectrum β-lactamases and carbapenemases and the semi-quantitative detection of highly expressed ISAba1-controlled variants.
CONCLUSION: Targeting mRNA with specific probes on magnetic beads will allow for adaptation to automated systems, such as point-of-care diagnostics.