Petri dish PCR: laser-heated reactions in nanoliter droplet arrays

Abstract

We report high-speed real-time PCR performed on an unmodified disposable polystyrene Petri dish. The reaction cycle relies solely on an infrared laser for heating; no conventional heater is required. Nanoliter droplets of PCR mixture as water-in-oil emulsions printed in an array format served as individual PCR microreactors. A simple contact printing technique was developed to generate a large array of uniform sized nanoliter droplets using disposable pipette tips. Printed droplets showed variation of less than 10% in volume and the oil/water/polystyrene interface formed a compact droplet microreactor approximately spherical in shape. The uniform droplet array was used to optimize the laser power required for the two heating steps of PCR, annealing/extension and melting, while the ambient conditions were at room temperature. The optical heating allows for an extremely fast heating rate due to the selective absorption of the infrared laser by PCR buffer only and not the oil or polystyrene Petri dish, allowing completion of 40 amplification cycles in ∼6 minutes. The quantitative assay capability of the system is also presented and discussed.