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http://dx.doi.org/10.18419/opus-10791
Autor(en): | Katzmeier, Florian Aufinger, Lukas Dupin, Aurore Quintero, Jorge Lenz, Matthias Bauer, Ludwig Klumpe, Sven Sherpa, Dawafuti Dürr, Benedikt Honemann, Maximilian Styazhkin, Igor Simmel, Friedrich C. Heymann, Michael |
Titel: | A low-cost fluorescence reader for in vitro transcription and nucleic acid detection with Cas13a |
Erscheinungsdatum: | 2019 |
Dokumentart: | Zeitschriftenartikel |
Seiten: | 17 |
Erschienen in: | PLOS ONE 14 (2019), No. 12, e0220091 |
URI: | http://nbn-resolving.de/urn:nbn:de:bsz:93-opus-ds-108085 http://elib.uni-stuttgart.de/handle/11682/10808 http://dx.doi.org/10.18419/opus-10791 |
ISSN: | 1932-6203 |
Zusammenfassung: | Point-of-care testing (POCT) in low-resource settings requires tools that can operate independently of typical laboratory infrastructure. Due to its favorable signal-to-background ratio, a wide variety of biomedical tests utilize fluorescence as a readout. However, fluorescence techniques often require expensive or complex instrumentation and can be difficult to adapt for POCT. To address this issue, we developed a pocket-sized fluorescence detector costing less than $15 that is easy to manufacture and can operate in low-resource settings. It is built from standard electronic components, including an LED and a light dependent resistor, filter foils and 3D printed parts, and reliably reaches a lower limit of detection (LOD) of ≈ 6.8 nM fluorescein, which is sufficient to follow typical biochemical reactions used in POCT applications. All assays are conducted on filter paper, which allows for a flat detector architecture to improve signal collection. We validate the device by quantifying in vitro RNA transcription and also demonstrate sequence-specific detection of target RNAs with an LOD of 3.7 nM using a Cas13a-based fluorescence assay. Cas13a is an RNA-guided, RNA-targeting CRISPR effector with promiscuous RNase activity upon recognition of its RNA target. Cas13a sensing is highly specific and adaptable and in combination with our detector represents a promising approach for nucleic acid POCT. Furthermore, our open-source device may be used in educational settings, through providing low cost instrumentation for quantitative assays or as a platform to integrate hardware, software and biochemistry concepts in the future. |
Enthalten in den Sammlungen: | 04 Fakultät Energie-, Verfahrens- und Biotechnik |
Dateien zu dieser Ressource:
Datei | Beschreibung | Größe | Format | |
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9 iGem2017.pdf | Katzmeier et al. 2019 | 2,21 MB | Adobe PDF | Öffnen/Anzeigen |
pone.0220091.s001.pdf | Katzmeier et al. 2019 Appendix S1 | 128,81 kB | Adobe PDF | Öffnen/Anzeigen |
pone.0220091.s002.png | Katzmeier et al. 2019 Fig S1 | 1,59 MB | image/png | Öffnen/Anzeigen |
pone.0220091.s003.tif | Katzmeier et al. 2019 Fig S2 | 8,47 MB | TIFF | Öffnen/Anzeigen |
pone.0220091.s004.png | Katzmeier et al. 2019 Fig S3 | 803,78 kB | image/png | Öffnen/Anzeigen |
pone.0220091.s005.png | Katzmeier et al. 2019 Fig S4 | 373,06 kB | image/png | Öffnen/Anzeigen |
pone.0220091.s006.jpg | Katzmeier et al. 2019 Fig S5 | 8,34 MB | JPEG | Öffnen/Anzeigen |
pone.0220091.s007.zip | Katzmeier et al. 2019 File S1 | 1,55 MB | Unknown | Öffnen/Anzeigen |
pone.0220091.s009.pdf | Katzmeier et al. 2019 File S3 | 45,81 kB | Adobe PDF | Öffnen/Anzeigen |
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