caa a22 4500 605480044 CHVBK 20210128100413.0 cr unu---uuuuu 210128e20151001xx s 000 0 eng 10.1007/s10544-015-9994-1 doi (NATIONALLICENCE)springer-10.1007/s10544-015-9994-1 Static self-directed sample dispensing into a series of reaction wells on a microfluidic card for parallel genetic detection of microbial pathogens [Elektronische Daten] [Robert Stedtfeld, Yen-Cheng Liu, Tiffany Stedtfeld, Tanja Kostic, Maggie Kronlein, Onnop Srivannavit, Walid Khalife, James Tiedje, Erdogan Gulari, Mary Hughes, Brett Etchebarne, Syed Hashsham] A microfluidic card is described for simultaneous and rapid genetic detection of multiple microbial pathogens. The hydrophobic surface of native acrylic and a novel microfluidic mechanism termed "airlock” were used to dispense sample into a series of 64 reaction wells without the use of valves, external pumping peripherals, multiple layers, or vacuum assistance. This airlock mechanism was tested with dilutions of whole human blood, saliva, and urine, along with mock samples of varying viscosities and surface tensions. Samples spiked with genomic DNA (gDNA) or crude lysates from clinical bacterial isolates were tested with loop mediated isothermal amplification assays (LAMP) designed to target virulence and antibiotic resistance genes. Reactions were monitored in real time using the Gene-Z, which is a portable smartphone-driven system. Samples loaded correctly into the microfluidic card in 99.3% of instances. Amplification results confirmed no carryover of pre-dispensed primer between wells during sample loading, and no observable diffusion between adjacent wells during the 60 to 90min isothermal reaction. Sensitivity was comparable between LAMP reactions tested within the microfluidic card and in conventional vials. Tests demonstrate that the airlock card works with various sample types, manufacturing techniques, and can potentially be used in many point-of-care diagnostics applications. The Author(s), 2015 Static self-directed sample dispensing nationallicence Airlock mechanism nationallicence Laser etched nationallicence Disposable microfluidic card nationallicence Isothermal amplification nationallicence POC nationallicence Genetic diagnostics nationallicence Gene-Z nationallicence LAMP nationallicence Stedtfeld Robert Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut Liu Yen-Cheng Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut Stedtfeld Tiffany Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut Kostic Tanja Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut Kronlein Maggie Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut Srivannavit Onnop Department of Chemical Engineering, University of Michigan, 48109, Ann Arbor, MI, USA aut Khalife Walid Department of Microbiology, Sparrow Laboratories, Sparrow Health System, 48912, Lansing, MI, USA aut Tiedje James The Center for Microbial Ecology, Michigan State University, 48824, East Lansing, MI, USA aut Gulari Erdogan Department of Chemical Engineering, University of Michigan, 48109, Ann Arbor, MI, USA aut Hughes Mary Department of Osteopathic Medical Specialties, Section of Emergency Medicine, College of Osteopathic Medicine, Michigan State University, 48824, East Lansing, MI, USA aut Etchebarne Brett Department of Osteopathic Medical Specialties, Section of Emergency Medicine, College of Osteopathic Medicine, Michigan State University, 48824, East Lansing, MI, USA aut Hashsham Syed Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut Biomedical Microdevices Springer US; http://www.springer-ny.com 17/5(2015-10-01), 1-12 1387-2176 17:5<1 2015 17 10544 https://doi.org/10.1007/s10544-015-9994-1 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s10544-015-9994-1 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Stedtfeld Robert Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Liu Yen-Cheng Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Stedtfeld Tiffany Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Kostic Tanja Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Kronlein Maggie Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Srivannavit Onnop Department of Chemical Engineering, University of Michigan, 48109, Ann Arbor, MI, USA aut NATIONALLICENCE 700 1- Khalife Walid Department of Microbiology, Sparrow Laboratories, Sparrow Health System, 48912, Lansing, MI, USA aut NATIONALLICENCE 700 1- Tiedje James The Center for Microbial Ecology, Michigan State University, 48824, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Gulari Erdogan Department of Chemical Engineering, University of Michigan, 48109, Ann Arbor, MI, USA aut NATIONALLICENCE 700 1- Hughes Mary Department of Osteopathic Medical Specialties, Section of Emergency Medicine, College of Osteopathic Medicine, Michigan State University, 48824, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Etchebarne Brett Department of Osteopathic Medical Specialties, Section of Emergency Medicine, College of Osteopathic Medicine, Michigan State University, 48824, East Lansing, MI, USA aut NATIONALLICENCE 700 1- Hashsham Syed Civil and Environmental Engineering, Michigan State University, 48824, East Lansing, MI, USA aut NATIONALLICENCE 773 0- Biomedical Microdevices Springer US; http://www.springer-ny.com 17/5(2015-10-01), 1-12 1387-2176 17:5<1 2015 17 10544