TY - JOUR
T1 - Fast prototyping microfluidics: Integrating droplet digital lamp for absolute quantification of cancer biomarkers
AU - Oliveira, Beatriz
AU - Veigas, Bruno
AU - Fernandes, Alexandra R.
AU - Águas, Hugo
AU - Martins, Rodrigo
AU - Fortunato, Elvira
AU - Baptista, Pedro Viana
N1 - UID/CTM/50025/2019
UID/Multi/04378/2019
Inn-INDIGO/0002/2015
PTDC/BTM-SAL/31201/2017
SFRH/BPD/124311/2016
PY - 2020/3/14
Y1 - 2020/3/14
N2 - Microfluidic (MF) advancements have been leveraged toward the development of state-of-the-art platforms for molecular diagnostics, where isothermal amplification schemes allow for further simplification of DNA detection and quantification protocols. The MF integration with loop-mediated isothermal amplification (LAMP) is today the focus of a new generation of chip-based devices for molecular detection, aiming at fast and automated nucleic acid analysis. Here, we combined MF with droplet digital LAMP (ddLAMP) on an all-in-one device that allows for droplet generation, target amplification, and absolute quantification. This multilayer 3D chip was developed in less than 30 minutes by using a low-cost and extremely adaptable production process that exploits direct laser writing technology in “Shrinky-dinks” polystyrene sheets. ddLAMP and target quantification were performed directly on-chip, showing a high correlation between target concentration and positive droplet score. We validated this integrated chip via the amplification of targets ranging from five to 500,000 copies/reaction. Furthermore, on-chip amplification was performed in a 10 µL volume, attaining a limit of detection of five copies/µL under 60 min. This technology was applied to quantify a cancer biomarker, c-MYC, but it can be further extended to any other disease biomarker.
AB - Microfluidic (MF) advancements have been leveraged toward the development of state-of-the-art platforms for molecular diagnostics, where isothermal amplification schemes allow for further simplification of DNA detection and quantification protocols. The MF integration with loop-mediated isothermal amplification (LAMP) is today the focus of a new generation of chip-based devices for molecular detection, aiming at fast and automated nucleic acid analysis. Here, we combined MF with droplet digital LAMP (ddLAMP) on an all-in-one device that allows for droplet generation, target amplification, and absolute quantification. This multilayer 3D chip was developed in less than 30 minutes by using a low-cost and extremely adaptable production process that exploits direct laser writing technology in “Shrinky-dinks” polystyrene sheets. ddLAMP and target quantification were performed directly on-chip, showing a high correlation between target concentration and positive droplet score. We validated this integrated chip via the amplification of targets ranging from five to 500,000 copies/reaction. Furthermore, on-chip amplification was performed in a 10 µL volume, attaining a limit of detection of five copies/µL under 60 min. This technology was applied to quantify a cancer biomarker, c-MYC, but it can be further extended to any other disease biomarker.
KW - Digital amplification
KW - Integrated microfluidics device
KW - Lab-on-a-chip
KW - Loop-mediated isothermal amplification
KW - Oncogenes
UR - http://www.scopus.com/inward/record.url?scp=85081608197&partnerID=8YFLogxK
U2 - 10.3390/s20061624
DO - 10.3390/s20061624
M3 - Article
C2 - 32183359
AN - SCOPUS:85081608197
SN - 1424-8220
VL - 20
JO - Sensors
JF - Sensors
IS - 6
M1 - 1624
ER -