Fast prototyping microfluidics: integrating Digital LAMP for evaluation of gene expression

Microfluidics (MF) is becoming the next step of integrated platforms for molecular diagnostics, where isothermal schemes allow further simplification of DNA detection and quantification protocols. MF for loop-mediated isothermal amplification (LAMP) is today the focus of a new generation of chip-based devices for molecular detection towards fast and automated nucleic acid quantification. Here, we integrated MF and digital droplet LAMP (ddLAMP) on a chip that allows droplet generation, amplification and target quantification. This multilayer 3D chip is produced using a low-cost and extremely adaptable production process using direct laser writing technology in Shrinky-dinks polystyrene plastic sheets in less than 30 minutes. ddLAMP and target quantification were performed directly on chip showing a high correlation between target concentration and positive droplet score. We validate this ddLAMP integrated chip via the amplification of targets between 5 and 500,000 copies/reaction under 60 min. Moreover, on-chip ddLAMP was performed in a 10 µL volume, with a limit of detection of 5 copies/µL of target. This technology was applied to quantify a cancer biomarker, c-MYC, but it can be further extended to any other disease biomarker.